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Progression of subclinical atherosclerosis in type 2 diabetes: Therapeutic and metabolic influences
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Progression of subclinical atherosclerosis in type 2 diabetes: Therapeutic and metabolic influences
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Content
PROGRESSION OF SUBCLINICAL ATHEROSCLEROSIS
IN TYPE 2 DIABETES:
THERAPEUTIC AND METABOLIC INFLUENCES
Copyright 2005
by
Ling Zheng
A Dissertation Presented to the
FACULTY OF THE GRADUATE SCHOOL
UNIVERSITY OF SOUTHERN CALIFORNIA
In Partial Fulfillment of the
Requirements for the Degree
DOCTOR OF PHILOSOPHY
(EPIDEMIOLOGY)
December 2005
Ling Zheng
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UMI Number: 3220174
Copyright 2005 by
Zheng, Ling
All rights reserved.
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ACKNOWLEDGEMENTS
I am very grateful for getting the best doctoral training possible from the Department
of Preventive Medicine at the University of Southern California. I cannot thank Dr.
Wendy Mack enough as my educational American dream would not have come true
without her support and guidance. Dr. Mack encouraged and challenged me
throughout my doctoral training. Her great editorial comments have been extremely
helpful. I thank Dr. Preston-Martin and Dr. Azen as they patiently guided me to
organize and present my dissertation better, and thank Dr. Buchanan and Dr. Hodis
for their great input in terms of understanding the fundamental pathophysiology of a
disease and interpreting statistical results from clinicians’ perspectives. In a word, I
appreciate all my committee members for sharing their time and knowledge with me
to help me grow in the past few years.
I am very thankful for the staff of three research teams - the Atherosclerosis
Research Unit, the General Clinical Research Center and the Statistical Consultation
Research Center at USC. Their contributions made TART data available and were
most appreciated. Also, I thank Drs. Hodis and Buchanan, and Dr. Wendy Mack for
their permission to use TART data for my dissertation.
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I am indebted to my parents and brothers for their loving support and being proud of
me as who I am. A very hearty thank you goes to my better half - Steven. It is not
easy to push husband through, while it is harder to push wife through. Thank you
Steven for cooking delicious meals, taking care of Harmony, and giving me great
feedback on my dissertation. My little sweetheart, Harmony, has grown with my
dissertation in the past three years. Many thanks for this special gift from the above
as I could not have accomplished this work without the joy she brings in my life.
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TABLE OF CONTENTS
ACKNOWLEDGEMENTS............................................................................................. ii
LIST OF TABLES..........................................................................................................vii
LIST OF FIGURES..........................................................................................................ix
ABBREVIATIONS...........................................................................................................x
ABSTRACT................................................................................................................... xiii
PREFACE.........................................................................................................................xv
I. Literature Review
1. Pathogenesis and Descriptive Epidemiology of Type 2 Diabetes Mellitus.
1
1.1 Definition and Diagnostic Criteria of Type 2 Diabetes Mellitus............. 1
1.2 Pathogenesis of Type 2 Diabetes Mellitus..................................................3
1.3 Descriptive Epidemiology of Type 2 Diabetes Mellitus .......................... 4
1.4 Vascular Complication of Type 2 Diabetes Mellitus ................................6
2. Descriptive Epidemiology of Cardiovascular Disease in Persons with
Type 2 Diabetes Mellitus............................................................. 8
2.1 Definition of Cardiovascular Disease......................................................... 8
2.2 Data Sources and Limitations......................................................................9
2.3 Prevalence.....................................................................................................10
2.4 Age................................................................................................................ 10
2.5 Race............................................................................................................... 11
2.6 Gender........................................................................................................... 11
2.7 Mortality....................................................................................................... 12
2.8 Time Trends..................................................................................................13
2. 9 Other Risk Factors ...................................................................................13
3. Pathogenesis of Atherosclerosis in Persons with Type 2 Diabetes Mellitus
3.1 Definition of Atherosclerosis......................................................................16
3.2 Structure of a Normal Arterial Wall........................................................... 17
3.3 Stages of Atherosclerosis Progression....................................................... 17
3.4 The Response-to-injury Hypothesis........................................................... 18
3.5 Function of the Normal Endothelium....................................................... 19
3.6 Mechanisms of Atherosclerosis in Persons with Type 2 Diabetes
Mellitus....................................................................................... 20
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4. Ultrasonographic Measurements of Atherosclerosis
-Common Carotid Intima-Media Thickness.............................24
4.1 Introduction...................................................................................................24
4.2 CIMT Measured by Ultrasonography versus Other Atherosclerosis
Imaging Methods.........................................................................25
4.3 Association of Atherosclerosis in Coronary and Carotid Arteries...........26
4.4 Methods for CIMT Measurement.............................................................. 27
4.5 IMT Measured in the Common Carotid Arteries versus Internal
Carotid Arteries and Bifurcation................................................28
4.6 Measurement of IMT in Near Wall versus Far Wall of the Carotid
Artery............................................................................................ 30
4.7 CVD Risk Factors and CIMT......................................................................31
4.8 CIMT and CVD............................................................................................32
4.9 Limitations of CIMT Measurements.......................................................... 35
5. Randomized Clinical Trials of Antihypertensive and Lipid Lowering
Therapy and CVD in Persons with Type 2 Diabetes M ellitus.................37
5.1 Introduction...................................................................................................37
5.2 Trials Testing the Effect of Antihypertensive Therapy on CVD Risk
.................................................................................. 38
5.3 Trials Testing the Effect of Lipid Lowering Therapy on CVD Risk
.................................................................................. 42
6. Randomized Clinical Trials of the Effects of Antihypertensive Agents
and Lipid Lowering Medications on CIMT Progression.......................... 49
6.1 Introduction................................................................................................... 49
6.2 Antihypertensive Agents and CIMT Progression...................................... 50
6.3 Trials Testing the Effects of Lipid Lowering Therapy on CIMT
Progression................................................................................... 57
6.4 Conclusions.................................................................................................. 64
n. Data Analysis and Planned Publications
7. Antihypertensive Therapy Reduces Progression of Subclinical
Atheroslcerosis among Patients with Type 2 Diabetes Mellitus...............66
7.1 Introduction.................................................................................................. 67
7.2 Methods........................................................................................................ 68
7.3 Results...........................................................................................................71
7.4 Discussion.....................................................................................................75
7.5 Conclusion....................................................................................................78
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8. Lipid Modifying Therapy Reduces Progression of Subclinical
Atheroslcerosis among Patients with Type 2 Diabetes Mellitus............... 79
8.1 Introduction...................................................................................................79
8.2 Methods.........................................................................................................81
8.3 Results........................................................................................................... 85
8.4 Discussion..................................................................................................... 88
8.5 Conclusion.................................................................................................... 91
III. Grant Proposal
9. Pooled Analysis of Metabolic Factors and Atherosclerosis
Progression....................................................................................................92
9.1 Specific Aims................................................................................................ 92
9.2 Backgroud and Signficance......................................................................... 95
9.3 Preliminary Studies.....................................................................................104
9.4 Research Design and Methods..................................................................114
9.5. Human Subjects Research....................................................................... 130
9.6. Study Timeline........................................................................................... 133
References...................................................................................................................... 134
Bibliography.................................................................................................................. 157
v i
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LIST OF TABLES
Table 4.1: Summary of Prospective Studies Examining the Association
between CIMT and Incident CVD Risk.......................................................................34
Table 5.1: Randomized Trials Testing the Effect of Specific Antihypertensive
Therapies on CVD Risk................................................................................................. 39
Table 5.2: Randomized Trials Testing the Effect of Lipid Lowering Therapy
on CVD Risk................................................................................................................... 43
Table 6.1: Summary of Randomized Clinical Trials of the Effect of
Antihypertensive Agents on CIMT Progression........................................................... 51
Table 6.2: Summary of Randomized Trials of the Effect of Lipid Lowering
Intervention on CIMT Progression................................................................................ 58
Table 7.1: Characteristics of TART Subjects at the Initial Screening Visit
and at Randomization..................................................................................................... 72
Table 7.2: Parameter Estimates Related to the CIMT Progression Rate from the
Multivariable Mixed Effect Model for Systolic Blood Pressure, Duration of
Antihypertensive Therapy and CIMT Progression.......................................................73
Table 8.1: Characteristics of TART Subjects at the Initial Screening Visit
and at Randomization..................................................................................................... 85
Table 8.2: Parameter Estimates Related to the CIMT Progression Rate from the
Multivariable Mixed Effect Model for LDL-C levels, Duration of LMT and
CIMT Progression...........................................................................................................87
Table 9.1: Design and Baseline Characteristics of Five Randomized
Atherosclerosis Regression Trials............................................................................... 107
Table 9.2: Baseline Characteristics of 199 Evaluable Subjects in EPAT................ 111
Table 9.3: Adjusted Comparisons of Inflammatory and Hemostatic Factors
by MetS group: EPAT.................................................................................................. 112
Table 9.4: Partial Spearman Correlation Coefficients of Inflammatory and
Hemostatic Factors and Other Components of the MS (N=199)..............................113
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Table 9.5: Pearson Partial Correlation Coefficients of hsCRP and sCIAMl
with hemostatic factors: EPAT.................................................................................113
Table 9.6: Baseline and ontrial Measurements of Five Atherosclerosis
Progression Trials.......................................................................................................122
Table 9.7: New Laboratory Data to be Collected in the Proposed Work............... 122
Table 9.8: Proposed statistical modeling using multivariate linear mixed
effects models............................................................................................................. 125
Table 9.9: Power Estimations (two-sided significance level = 0.05)...................... 129
Table 9.10: Proposed Study Timeline......................................................................... 133
v iii
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LIST OF FIGURES
Figure 3.1: Endothelial Dysfunction in Diabetes........................................................ 21
Figure 4.1: Diagrammatic representation of right carotid artery scan..................... 29
Figure 7.1: Predicted annual CIMT progression rates in relation to SBP and the
duration of antihypertensive therapy from the multivariable mixed-effects
model................................................................................................................................ 74
Figure 8.1: Predicted annual CIMT rates (using Table 8.2 parameter estimates)
in relation to LDL-C levels and the duration of LMT from the multivariate
mixed-effects model (assuming a lOmg/dL reduction of LDL-C during the
trial).................................................................................................................................. 88
ix
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ABBREVIATIONS
4S Scandinavian Simvastatin Survival Study
ACAPS Asymptomatic Carotid Artery Progression Study
ACAPS Asymptomatic Carotid Artery Progression Study
ADA American Diabetes Association
AFC APS/T exC APS
Air Force Coronary Atherosclerosis Prevention
Study/Texas Coronary Atherosclerosis Prevention Study
ALLHAT-LLT
Antihypertensive and Lipid-Lowering Treatment to
Prevent Heart Attack Trial ~ Lipid Lowering Therapy
ARBITER
Arterial Biology for the Investigation of the Treatment
Effects of Reducing Cholesterol
ARIC Atherosclerosis Risk in Communities Study
ASAP
Atorvastatin vs. Simvastatin on Atherosclerosis
Progression
ASCOT-LLA
Anglo-Scandinavian Cardiac Outcomes Trial -Lipid
Lowering Arm
BMI body mass index
BRFSS the Behavioral Risk Factor Surveillance System
BVAIT the B-Vitamin Atherosclerosis Intervention Trail
CAD coronary artery disease
CARE Cholesterol and Recurrent Events
CCA common carotid artery
CDC Centers for Disease Control and Prevention
CHD coronary heart disease
CHS Cardiovascuar Health Study
CIMT carotid intima-media thickness
CLAS Cholesterol Lowering Atherosclerosis Study
CVD cardiovascular disease
DBP diastolic blood pressure
DM diabetes mellitus
ELAM-1 endothelial cell adhesion molecule-1
EPAT the Estrogen in the Prevention of Atherosclerosis Trial
HDL-C high density lipoprotein cholesterol
HHS Helsinki Heart Study
HOPE Heart Outcomes and Prevention Evaluation
HOT Hypertension Optimal Treatment
HPS Heart Protection Study
ICA internal carotid artery
IDNT Irbesartan Diabetic Nephropathy Trial
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IFN-g interferon gamma
IGT impaired glucose tolerance
IL-1 interleukin-1
IMT intima-media thickness
INSIGHT
Intemaltional Nifedipine GITS Study: Intervention as a
Goal in Hypertension Treatment
KIHD Kuopio Ischemic Heart Diseae study
LDL low density lipoprotein
LDL-C low density lipoprotein cholesterol
LIPID
Long-term Intervention with Pravastatin in Ischemic
Disease
LPS Lescol Intervention Prevention Study
LMT Lipid modifying therapy
MARS Monitored Atherosclerosis Regression Study
MIDAS Multicenter Isradipine Diuretic Atherosclerosis Study
MetS the Metabolic Syndrome
NCRR National Center for Research Resources
NDDG National Diabetes Data Group
NHANESm
the Third National Health and Nutrition Examination
Survey
NHIS National Health Interview Survey
NIH National Institutes of Health
OGTT oral glucose tolerance test
PLAC Pravastin, Lipids, and Atheroscleoris in the Carotids trial
Post-CABG Post-Coronary Artery Bypass Graft
PREVENT
Prospective Randomized Evaluation of the Vascular
Effects of Norvasc Trial
PROSPER Prospective Study of Pravastatin in the Elderly at Risk
REGRESS Regression Growth Evaluation Statin Study
RENAAL
Reduction of Endpoints in NIDDM with the Angiotensin II
Antagonist Losartan Study
SBP systolic blood pressure
SECURE
Study to Evaluate Carotid Ultrasound changes in patients
treated with Ramipril and vitamin E
SHEP Systolic Hypertension in the Elderly Program
Syst-Eur Systolic Hypertension in Europe
T2DM type 2 diabetes mellitus
TART the Troglitazone Atherosclerosis Regression Trial
TNF-a tumor necrosis factor-alpha
UKPDS-HDS
United Kingdom Prospective Diabetes Study -
Hypertension in Diabetes Study
x i
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VCAM-1 vascular cell adhesion molecule-1
VEAPS the Vitamin E Atherosclerosis Prevention Study
VLDL very low density lipoprotein
WELL-HART
the Women’s Estrogen-Progestin Lipid-Lowering
Hormone Atherosclerosis Regression Trial
WHO World Health Organization
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ABSTRACT
Diabetes is a major risk factor for cardiovascular disease (CVD). As a
pathological pathway of CVD, atherosclerosis progression is accelerated in diabetics.
In nondiabetic populations, the beneficial effect of lipid modifying therapy is
conclusive while the antiatherogenic effects of antihypertensive agents remain
controversial. Few such data are available in persons with type 2 diabetes mellitus
(T2DM). To investigate the association between antihypertensive and lipid
modifying therapy and CIMT progression among diabetic patients, we conducted
post hoc cohort analyses using longitudinal data from the Troglitazone
Atherosclerosis Regression Trial, a randomized trial designed to evaluate the impact
of troglitazone on CIMT progression in adults with insulin-requiring T2DM.
Multivariate mixed-effects models were used to evaluate annual rate of change in
CIMT in relation to blood pressure and the duration of antihypertensive therapy, and
in relation to low-density lipoprotein cholesterol and duration of lipid modifying
therapy, respectively. Our data indicate antihypertensive and lipid modifying
therapy are associated with a reduction in CIMT progression in a duration dependent
manner in type 2 diabetes. Moreover, we propose to pool data from five completed
clinical trials to evaluate the contributions of the metabolic syndrome (MetS) to
CIMT progression. Approximately one fourth of American adults have the MetS
and these individuals confer greater risk of cardiovascular morbidity and mortality.
We will investigate the underlying structures of the MetS before and after
incorporating proinflammatory and prothrombotic variables in the factor analysis.
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We will also evaluate the value of adding inflammatory and hemostatic factors to the
definition of MetS. Furthermore, we will examine the potential risks of core factors
of the MetS on atherosclerosis progression among subjects with CVD or diabetes. In
conclusion, this dissertation provides important insights on therapeutic and metabolic
influences on progression of subclinical atherosclerosis in type 2 diabetes. These
findings may improve treatment, cardiovascular risk prediction and further
evaluation of antiatherogenic interventions in this high-risk population.
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PREFACE
Diabetes is a major risk factor for cardiovascular disease (CVD). Type 2
diabetes mellitus (T2DM) comprises approximately 95% of all diabetes cases. CVD
is the most prevalent complication in persons with T2DM. Over the past three
decades, diabetic populations in the United States have not experienced the same
improvements in cardiovascular event survival as nondiabetic populations. The
current and projected continuing epidemic of T2DM and associated prevalence of
CVD imply an extensive health care burden in the next two to three decades.
As a pathological pathway of CVD, atherosclerosis is characterized as a
progressive disease with structural change in the intimal and medial layers of the
vascular system. Atherosclerosis progression is accelerated in persons with T2DM.
Common carotid intima-media thickness (CIMT) assessed with high resolution B-
mode ultrasonography is a well-established noninvasive measure of subclinical
atheroscleoris and is positively associated with elevated levels of conventional
cardiovascular risk factors in both diabetic and nondiabetic populations. Increased
CIMT and accelerated progression of CIMT are also associated with increased risk
for cardiovascular morbidity and mortality.
In nondiabetic populations, the beneficial effect of lipid lowering therapy is
conclusive while the antiatherogenic effects of antihypertensive agents remain
controversial. Few such data are available in persons with T2DM. The coexisting
conditions of hypertension and dyslipidemia in persons with T2DM define a high-
risk group for cardiovascular disease and warrants intensive prevention strategies.
xv
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The therapeutic efficacy of different strategies remains to be addressed by
interventional trials in an efficient and cost-effective manner. Clinical trials and
observational studies using CIMT progression as an endpoint require much smaller
sample sizes, shorter duration of follow-up and also allow evaluation of efficacy
among relatively young adults or even adolescents. The latter advantage is critical
because of the increasing incidence of T2DM and prevalence of known CVD risk
factors in American youth.
This dissertation investigates the association between antihypertensive and
lipid lowering therapy and progression of CIMT among persons with T2DM, using
data from the Troglitazone Atherosclerosis Regression Trial (TART), a randomized
clinical trial designed to determine whether troglitazone (400mg daily) reduces the
progression of subclinical atherosclerosis in persons with insulin-requiring T2DM.
In the literature review chapter (focusing on persons with T2DM), we review
the pathogenesis and descriptive epidemiology of T2DM (Section 1), and
cardiovascular disease (Section 2), and pathogenesis of atherosclerosis (Section 3).
Ultrasonographic measurements of atherosclerosis with CIMT are presented in
Section 4. We then summarize randomized clinical trials of antihypertensive and
lipid lowering therapy and CVD (Section 5), and trials examining the effects of
antihypertensive agents and lipid lowering medications on CIMT progression
(Section 6).
In the data analysis chapter, the effects of antihypertensive therapy on
progression of subclinical atherosclerosis among hypertensive patients with type 2
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diabetes mellitus using TART data are presented in Section 1. The association of
lipid modifying therapy and progression of subclinical atherosclerosis using the same
study cohort will be presented in Section 2.
In the grant proposal chapter, we propose to perform a pooled analysis of the
data from the Estrogen in the Prevention of Atherosclerosis Trial (EPAT), the
Vitamin E Atherosclerosis Prevention Study (VEAPS), the Women’s Estrogen-
Progestin Lipid-Lowering Hormone Atherosclerosis Regression Trial (WELL-
HART), TART and the B-vitamin Atherosclerosis Intervention Trail (BVAIT).
These trials were all conducted in the past decade at the Atherosclerosis Research
Unit of University of Southern California, using similar trial and data collection
protocols. The wide variety of populations sampled, including persons with and
without CVD, and with and without diabetes, will allow us to evaluate our
hypotheses among these subgroups.
The specific aims are to pool data from five completed clinical trials using
CIMT as a trial endpoint in order to evaluate the contributions of the metabolic
syndrome (MS) to progression of subclinical atherosclerosis. By principal
component analysis, we will identify principal components of the MS that are
statistically independent of one another using two sets of original variables: (1) the
standard 5 metabolic factors used in the NCEP ATP III definition of the MS, and (2)
the standard 5 metabolic factors plus inflammatory and hemostatic variables. Then
we will investigate the associations between the two sets of principle components of
the MS and progression of CIMT in the pooled sample, and determine the best fitting
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model. Based on the best fitting model, we will further examine whether the
associations between principal components of the MS and CIMT progression vary by
CVD and diabetes status.
Ling Zheng
x v iii
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Literature Review
Chapter 1: Pathogenesis and Descriptive Epidemiology of Type 2 Diabetes
Mellitus
Summary of Key Points
• Insulin resistance and consequent P-cell exhaustion are two key steps in
the pathogenesis of type 2 diabetes mellitus
• Current epidemic of type 2 diabetes mellitus in the United States
• The projected continuing epidemic of type 2 diabetes mellitus implies
extensive health care burden in the United States
1.1 Definition and Diagnostic Criteria of Type 2 Diabetes Mellitus
Diabetes Mellitus (DM) is a group of metabolic diseases characterized by
hyperglycemia resulting from defects in insulin secretion, insulin action, or both.
There are three major types of DM - type 1 DM, type 2 DM and gestational DM.
Type 1 DM was previously referred to as insulin-dependent diabetes mellitus
because pancreatic beta-cell destruction in these patients usually leads to insulin
deficiency. Gestational DM is defined as hyperglycemia first recognized during
pregnancy. Type 2 diabetes mellitus (T2DM) comprises approximately 95% of all
DM cases. T2DM was previously referred to as non-insulin-dependent diabetes
mellitus or adult-onset diabetes mellitus as exogenous insulin is not always required
in this population (1).
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The first generally accepted systematic classification and diagnosis of
diabetes was developed by the National Diabetes Data Group (NDDG) in 1979. In
1985, the World Health Organization (WHO) study group on diabetes mellitus
endorsed the substantive recommendations of the NDDG and based on age of onset
distinguished two major forms of DM - type 1 DM and type 2 DM (1). According
to the NDDG/WHO criteria, diabetes is defined by fasting plasma glucose >140
mg/dl or 2-hour plasma glucose > 200 mg/dl; impaired glucose tolerance is defined
by fasting plasma glucose < 140 mg/dl and 2-hour plasma glucose 140 - 199 mg/dl.
In 1997, the Expert Committee of the American Diabetes Association (ADA)
recommended several changes to the above diagnostic criteria, including the
definition of normal fasting plasma glucose as <110 mg/dl, and the definition of
diabetes as having fasting plasma glucose >126 mg/dl (1).
Thus, the ADA specifies three criteria to diagnose diabetes mellitus: 1)
symptoms of diabetes (polyuria, polydipsia and unexplained weight loss) plus a
casual plasma glucose concentration > 200 mg/dl, 2) fasting plasma glucose >126
mg/dl, 3) 2-hour plasma glucose > 200 mg/dl during a 75-g oral glucose tolerance
test (OGTT). If any one of the above three criteria is confirmed in a subject twice, a
clinical diagnosis can be made. Impaired fasting glucose is defined as fasting plasma
glucose 110 -125 mg/dl. For epidemiologic studies, a single fasting plasma glucose
concentration is typically used to estimate the prevalence of diabetes in a population
( 1).
2
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1.2 Pathogenesis of Type 2 Diabetes Mellitus
The specific etiologies of T2DM are not known, although it is widely
accepted that T2DM results from an interaction between genetic and environmental
factors. The specific etiologic genes are still unknown but are under intense
investigation. Environmental factors include increasing age, obesity, high caloric
intake, physical inactivity, low birthweight and family history of T2DM.
The underlying metabolic causes of T2DM are a combination of impairment
in insulin-mediated glucose disposal (insulin resistance) and defective secretion of
insulin by pancreatic beta-cells. Insulin resistance in insulin sensitive tissues
(muscle, liver and fat) develops from obesity and physical inactivity, acting on a
substrate of genetic susceptibility (2). Insulin secretion declines with advancing age
(3), and this decline may be accelerated by genetic factors (4).
Based on longitudinal data on a cohort of Pima Indians, a two-step
development of T2DM was proposed and widely accepted (5). During the first step,
individuals with normal glucose tolerance progress to impaired glucose tolerance
(IGT) with the development and worsening of insulin resistance. The possible
mechanisms of insulin resistance include diminished numbers of insulin receptors on
target cells, diminished capacity of insulin receptors on target cells and problems in
the translocation of glucose transporters.
The second step in the development of T2DM consists of a worsening of
impaired glucose tolerance until beta cells become exhausted. In response to insulin
resistance, the pancreatic islet initiates a prolonged period of insulin hypersecretion,
3
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which produces fi-cell exhaustion and results in insulin deficiency. Insulin deficiency
leads to increased hepatic glucose output and results in fasting hyperglycemia.
1.3 Descriptive Epidemiology of Type 2 Diabetes Mellitus
1.3.1 Data Sources and Limitations
In epidemiologic studies, a single fasting plasma glucose concentration is
often used to estimate the prevalence of diabetes in a population (1). T2DM is a
chronic disease with a long asymptomatic stage (4 - 7years). It is difficult to assess
the incidence rate due to the unclear timing of disease onset. Therefore, the
prevalence of T2DM is often used as the measure of disease occurrence.
The Centers for Disease Control and Prevention (CDC) has compiled data on
diabetes in the United States obtained from several surveys, including the National
Health Interview Survey (NHIS), the third National Health and Nutrition
Examination Survey (NHANES HI), the National Hospital Discharge Survey, and
surveys conducted through the Behavioral Risk Factor Surveillance System
(BRFSS). The ADA diagnostic criteria are used for estimation.
1.3.2 Prevalence and Incidence
Based on data from these national surveys, the CDC estimates approximately
18.2 million people (6.3% of the population, 13 million diagnosed and 5.2 million
undiagnosed) in the United States had diabetes in 2002. Among them, about 95%
had T2DM. Approximately 1.3 million new cases of DM are diagnosed every year
(6; 7).
4
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According to the NHANES III (1988-1994), the prevalence rate of diabetes
(diagnosed and undiagnosed) increases with age. Among persons aged 20 to 39
years, about 1-2 % have this disease, while 18-20 % of persons aged 60 to 74 years
have diabetes (8). Prevalence is similar for men and women in each age group (20-
39, 40-49, 50-59, 60-74 and > 75 years). The age-standardized prevalence rates are
84 and 77 per 1,000 in men and women, respectively.
The prevalence of diagnosed diabetes varies greatly among ethnic groups.
The age- and sex- standardized prevalence rates are 48, 82 and 93 per 1,000 for non-
Hispanic whites, non-Hispanic blacks and Mexican-Americans, respectively (8).
The prevalence in American Indians is high (around 20%) with the Pima tribe of
Arizona having the highest prevalence (> 50%) in the US and in the world (9).
1.3.3 Time Trends
The NHANES HI also reported trend of increasing diabetes prevalence in the
United Sates over the past three decades (10). Using ADA diagnostic criteria, the
prevalence of diabetes, among persons who are 40-74 years of age, increased from
8.9% in the years 1976-1980 to 12.3% by 1988-1994. Using WHO diagnostic
criteria in the same sample, a similar increasing trend was found with 11.4% in the
years 1976-1980 to 14.3% by 1988-1994. Increasing prevalence reflects both the
declining diabetes-related mortality (11) and increasing incidence of diabetes in all
ethnic groups, especially in minority populations (Pima Indians, African Americans
and Hispanic Americans) (12; 13) and American youth (14; 15).
5
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The prevalence rates for diabetes are predicted to grow substantially over the
next several decades (16). The projected increase in the prevalence of T2DM raises
major concerns regarding morbidity and mortality associated with this disease. As
the population of the US grows older, more sedentary, and obese, the risk of
developing diabetes and its complications will increase. The continuing epidemics
of obesity and diabetes convey increased risk for CVD and premature mortality,
which implies a potentially large health care burden in the United States.
1.4 Vascular Complications of Type 2 Diabetes Mellitus
T2DM is frequently not diagnosed until complications appear. The
complications of T2DM primarily affect the vascular system. Microvascular
complications result from changes in the small blood vessels, and include diabetic
nephropathy, neuropathy and retinopathy. Macrovascular complications result from
changes in the large blood vessels, and include diseases of coronary arteries,
peripheral arteries, and carotid vessels. These vascular complications increase
morbidity and mortality in T2DM and can be prevented or delayed with
multifactorial intervention (17).
Diabetic nephropathy refers to elevated urinary albumin or protein excretion
in the absence of other renal disease. It is the single leading cause of end-stage renal
disease (18). About 20-40% of persons with diabetes have nephropathy.
Microalbuminuria (persistent albuminuria in the range of 30-299 mg/24h) is not only
a marker for development of nephropathy, but also a marker of increased CVD risk
among individuals with T2DM.
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Diabetic retinopathy refers to damage of the small blood vessels in the retina.
It is the leading cause of blindness in persons aged 20 to 74 years in the United
States (18). About 15-28% of persons with T2DM have retinopathy at the time of
diabetes diagnosis. By twenty years after the initial diagnosis of diabetes, more than
60% of persons with T2DM have developed some degree of retinopathy (19).
Diabetic neuropathy is the disease of nerves. It can occur in both the
peripheral and autonomic nervous systems. It causes pain, paraesthia, hyperesthia,
dysesthia, muscle weakness, atrophy and loss of sensation of the extremities.
Amputation and foot ulceration are the most common consequences of diabetic
neuropathy. The prevalence of neuropathy ranges from 17 to 26% in persons with
T2DM (18).
Macrovascular complications, including coronary heart disease (CHD),
stroke, and peripheral vascular disease, are the major causes of morbidity and
mortality in persons with T2DM (20). The combination of CHD and stroke are often
referred to as cardiovascular disease. Compared to nondiabetic persons, iindividuals
with T2DM have a 2 to 4-fold increase in the risk of CVD (21), a 1.5 to 3.0-fold
increase in CHD mortality and about a 2 fold increase in stroke mortality (22). The
prevalence of peripheral vascular disease is about 22% among persons with T2DM,
and is higher than that among nondiabetic persons (23). Overall, persons with
T2DM and CVD have a poorer prognosis, poorer short-term survival, increased risk
of recurrent disease, poorer response to surgery and increased risk of congestive
heart failure relative to nondiabetic persons with CVD (24).
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Literature Review
Chapter 2: Descriptive Epidemiology of Cardiovascular Disease in Persons with
Type 2 Diabetes Mellitus
Summary of Key Points
• CVD is the most prevalent complication in persons with T2DM, thus is
the largest cost component in management of medical complications
• Over the past 30 years, diabetic populations have not experienced the
same improvements in cardiovascular event survival as persons without
DM
• The current and projected continuing epidemic of T2DM and associated
prevalence of CVD imply an extensive health care burden in the next two
to three decades
2.1 Definition of Cardiovascular Disease
Cardiovascular disease (CVD) is a disease of the heart (cardio) and blood
vessels (vascular) often caused by a narrowing of the blood vessels, which is often
due to accumulation of plaque in the lining of these vessels. It includes coronary
heart disease, stroke, congestive heart failure, peripheral vascular disease, congenital
heart disease, endocarditis, and many other conditions.
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2.2 Data Sources and Limitations
The majority of data regarding the prevalence of CVD in this section derive
from the National Health Interview Survey (NHIS) of the National Center for Health
Statistics, CDC (25). The NHIS is a health survey of the civilian,
noninstitutionalized, household population of the United States, conducted annually
since 1957. All participants are 35 years of age or older. Diabetes and CVD is self-
reported by asking whether a health professional had ever told them they had
diabetes or CVD (including coronary heart disease, any other kind of heart condition
or heart disease and stroke). Respondents who refused to answer or did not know
were excluded from data analyses. Estimates were age-adjusted using estimates of
the 2000 U.S. population as the standard. Because approximately one third of
persons with diabetes are undiagnosed, this survey cannot reflect the burden of CVD
among persons with undetected diabetes.
Data regarding hospital discharge in persons with diabetes are available from
the National Hospital Discharge Survey (NHDS) of the National Center for Health
Statistics, CDC (26), which collected data annually from a sample of short-stay,
nonfederal hospitals in the United States. Hospital discharges for which diabetes
was listed as the first diagnosis or as any of six secondary diagnoses were examined.
Rates were calculated using resident population estimates and estimates of the
population with diabetes. Rates were adjusted to the 2000 U.S. Standard Population
using four age groups (0-44, 45-64, 65-74, 75+). Because long-term and federal
hospitals are not included in this survey, hospitalizations involving persons with
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diabetes are likely underestimated. In addition, persons who are hospitalized more
than once in a year are counted more than once. Therefore, discharge rates might not
reflect rates per person.
2.3 Prevalence
From these data, it is estimated that CVD accounted for 77% of
hospitalizations for chronic complications in diabetic populations in the United
States in 1989 (27). According to the NHIS, 5 million reported receiving a
diagnosis of CVD among diagnosed diabetic patients (excluding gestational
diabetes) aged 35 years or older in 2002 (28). Among these 5 million diabetic
patients, 3.4 and 1.3 million reported receiving a diagnosis of coronary heart disease
and stroke, respectively.
2.4 Age
The prevalence of ischemic heart disease among persons with diabetes is
about 14, 3 and 2 times the rate among nondiabetics in age groups of 18-44 years,
45-64 years and 65 years or older, respectively (29). In 2002, among diagnosed
diabetic patients (excluding gestational diabetes) aged 35 years or older, the
prevalence of any self-reported CVD was 288, 464, and 580 per 1,000 among
diabetics aged 35-64, 65-74 and > 75 years, respectively (28). The rates for CVD as
a first-listed diagnosis among diabetic populations increases with age. In 2001, age-
specific rates were 21.5, 83.5, 145.7, and 240.2 per 1,000 among diabetics aged 0-44,
46-64, 65-74 and > 75 years, respectively (30).
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2.5 Race
The extent to which diabetes accounts for CVD in the United States varies in
different ethnic groups, largely because of the differences in the frequency of
diabetes among them. The proportion of CVD attributable to diabetes or IGT ranges
from about 14% in the white population to 50% to 80% among the American Indian
population (10). Due to the increasing prevalence of diabetes in all ethnic groups, the
absolute risk of CVD will continue to increase.
Among diagnosed diabetic patients (excluding gestational diabetes) aged 35
years or older in 2002, the age-adjusted prevalence of CVD were 359, 305 and 256
per 1,000 among whites, blacks and Hispanics, respectively (31). Age-standardized
rates for major CVD as a first-listed diagnosis among diabetic populations increased
slightly in whites but dramatically in blacks in the past two decades. In 2001, the
rate for whites was similar to the rate in 1980 (40.9 vs. 43.8 per 1,000), while the
2001 rate for blacks was 77.6% higher than in 1980 (62.9 vs. 35.2 per 1,000 diabetic
population) (30). No similar information is available for Hispanics.
2.6 Gender
For people with diabetes, the prevalence rates of CVD increase with age and
are higher in men than in women. Among diagnosed diabetic patients (excluding
gestational diabetes) aged 35 years or older in 2002, the age-adjusted prevalence of
any self-reported CVD was 399 per 1,000 for white males, 315 per 1,000 for white
females, 277 per 1,000 for black males, 323 per 1,000 for black females, 298 per
1,000 for Hispanic males, and 210 pre 1,000 for Hispanic females (31). From 1980
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to 2001, age-standardized rates of hospital discharge for major CVD as a first-listed
diagnosis among diabetic populations increased slightly in men but greatly in
women. This age-standardized rate for males in 2001 was similar to the rate in 1980
(56.4 vs. 56.0 per 1,000 diabetic population), while the rate for females was 31.7%
higher than in 1980 (57.3 vs. 43.5 per 1,000) (28).
2.7 Mortality
As the most prevalent complication of diabetes mellitus, CVD is the cause of
more than 80% of deaths in patients with T2DM (32). Mortality from stroke is
increased almost 3-fold when patients with diabetes are matched to those without
diabetes (33). Diabetic populations have not experienced the same improvements in
cardiovascular event survival as persons without diabetes mellitus over the past 30
years. In non-diabetic populations, the age-adjusted heart disease mortality rate
declined 36.4% in men and 27% in women from the years 1971-1975 to the years
1982-1984 (34). Over the same period in diabetic populations, this rate declined
13.1% in men and increased 23% in women. The marked decline in mortality from
CVD in the overall US population has been attributed to reduction in risk factors
(high blood pressure, hypercholesterolemia and cigarette smoking) for CVD and
improvement in acute treatment intervention (35; 36). However, the smaller decline
in men and greater increase in women observed in the diabetic population suggest
that more aggressive treatment interventions should be implemented in patients with
diabetes mellitus.
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2.8 Time Trends
The prevalence of CVD in persons with T2DM has increased in the past two
decades. According to the NHDS, between 1980 and 2001 the number of hospital
discharges with CVD as the first-listed diagnosis and diabetes as a secondary
diagnosis increased from about 549,000 (2.74 per 1,000) to 1,404,000 (4.96 per
1,000).
2.9 Other Risk Factors
As an underlying etiology of CVD, the development of atherosclerosis is
accelerated among persons with T2DM (37). The exact causes are not well
understood. However, conventional risk factors for the development of
atherosclerosis are well established, such as, hypertension, abnormal lipid profile
(elevated total cholesterol, low-density lipoprotein cholesterol (LDL-C) and
triglycerides, and decreased high-density lipoprotein cholesterol (HDL-C)), and
smoking. Prospective studies indicate that all of these traditional cardiovascular risk
factors continue to act as independent contributors to CVD in persons with T2DM
(9). The role of other non-conventional risk factors, such as altered lipoprotein
concentrations, inflammatory markers, fibrinogen, physical inactivity and high fat
diet, is not conclusive and needs more investigation.
2.9.1 Prevalence of Hypertension
Hypertension is about twice as prevalent among persons with diabetes than
among those without (38). According to the Third National Health and Nutrition
Examination survey (1988-1994), 71% of U.S. adults with diabetes have elevated
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blood pressure (> 130/85 mmHg) or are taking prescription drugs for hypertension
(39). In this cohort of diabetics, the prevalence of elevated blood pressure increased
with age (39.6%, 71.5% and 83.5% among peopled aged 18 to 44 years, aged 45 to
64 years and aged > 65 years, respectively) and was higher in women than men
across the three age groups. Age-adjusted prevalence rates are uniformly high for
both genders and for Mexican Americans, non-Hispanic blacks, and non-Hispanic
whites. Among hypertensive patients with T2DM, 71% had hypertension, 57% were
treated while only 12% had a blood pressure < 130/85 mmHg and 45% had a blood
pressure < 140/90 mmHg (39).
Moreover, individuals with T2DM who are hypertensive have an
approximate 2-fold higher risk of cardiovascular events than individuals with T2DM
who are normotensive (40). The mortality rate for hypertensive diabetics is three
times that of normotensive diabetics (41). Therefore, aggressive blood pressure
control (<130/80 mmHg) is recommended for persons with T2DM (17).
2.9.2 Prevalence of Dyslipidemia
Population-based data on the prevalence of dyslipidemia (disorders of
lipoprotein metabolism, including lipoprotein overproduction or deficiency) in
persons with T2DM are scant. The form of dyslipidemia that is most characteristic
of type 2 diabetes is increased triglyceride and decreased HDL-C levels (42; 43). In
many studies, the dyslipidemia in type 2 diabetic subjects relative to nondiabetic
subjects is more severe in women than in men (42; 43), which is consistent with the
relatively greater risk of CHD in diabetic women than in diabetic men. In the
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Framingham Study cohort, prevalence of hypertriglyceridemia (>203 mg/dl) in type
2 diabetic and nondiabetic men was 19% and 9%, respectively. In type 2 diabetic
and nondiabetic women, the prevalence of hypertriglyceridemia (>173 mg/dl) was
17% and 8%, respectively. The corresponding prevalence for low HDL-C was 21%
(vs. 9%) in men (<30 mg/dl) as opposed to 25% (vs. 10%) in women (<38 mg/dl).
The prevalence of high LDL cholesterol was 9% in men with T2DM, 11% in
nondiabetic men, 15% in women with T2DM and 16% in nondiabetic women (43).
Of note, most patients with T2DM do not have marked elevations of LDL
cholesterol, but these patients nonetheless carry high enough levels to support the
development of atherosclerosis (44). Recent lipid-lowering trials demonstrated the
role of LDL cholesterol in patients with T2DM. Even borderline-high-risk LDL
cholesterol level (130-159 mg/dl) are of concern in T2DM and a target goal of < 100
mg/dl is recommended by the Third Report of the National Cholesterol Education
Program’s Adult Treatment Panel (45).
In summary, CVD is the most prevalent complication in persons with T2DM,
thus is the largest cost component in the management of clinical complications (46).
Hypertension and dyslipedemia are two common coexisting conditions in persons
with T2DM. Aggressive and early antiatherogenic interventions in this particular
population can be more cost-effective relative to interventions in groups with any
single cardiovascular risk factor.
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Literature Review
Charpter 3: Pathogenesis of Atherosclerosis in Persons with Type 2 Diabetes
Mellitus
Summary of Key Points
• Atherosclerosis is a progressive inflammatory disease
• Atherosclerosis progression is accelerated in persons with T2DM
• Endothelial dysfunction plays a central role in the pathogenesis of
atherosclerosis in persons with T2DM
3.1 Definition of Atherosclerosis
“Atherosclerosis comes from the Greek words athero (meaning gruel or
paste) and sclerosis (hardness)”. It is a progressive disease characterized by the
accumulation of lipids and fibrous elements in the wall of the large and medium
sized arteries, resulting in narrowing of the arterial lumen and eventual impairment
in blood flow (47). Atherosclerosis is the primary cause of heart disease and stroke.
Atherosclerosis used to be considered a degenerative process that inevitably
progresses with age and results simply from the accumulation of lipids. During the
past two decades, the concepts behind the pathogenic mechanisms of atherosclerosis
have changed dramatically. Currently, atherosclerosis is viewed as a chronic
inflammatory process because monocyte-derived macrophages and specific subtypes
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of T lymphocytes are present in every stage of the disease process (48). This chronic
condition can lead to an acute clinical event by plaque rupture and thrombosis.
3.2 Structure of a Normal Arterial Wall
The arterial wall consists of three layers: intima, media and adventitia. The
intima is composed of a single continuous layer of endothelial cells and internal
elastic lamina. Arterial endothelial cells regulate a number of important vascular
functions, including vasomotor tone, coagulation, inflammation, and vascular
growth. The medial layer consists predominantly of smooth muscle cells, embedded
in extracellular matrix, that regulate vascular tone in response to local and humoral
stimuli. The adventitia is composed mainly of fibroelastic tissue and collagen (49).
3.3 Stages of Atherosclerosis Progression
Six types of histological intimal lesions appear sequentially in the
progression of atherosclerosis, including foam cells, fatty streaks, intermediate
lesions, atheromas, fibrous plaques, and complicated lesions (50). The growth of the
lesion is caused mainly by lipid accumulation in the first four stages. Foam cells and
fatty streaks usually occur in infants and children and therefore also are referred to as
early lesions. Intermediate lesions may evolve soon after puberty and are the
histologic bridge between early and advanced lesions. Atheromas consist of
intracellular lipid accumulation and a core of extracellular lipid and usually occur
from the third decade on. Fibrous plaques appear from the fourth decade on and are
formed by smooth muscle proliferation and collagen. Complicated lesions are
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characterized by combined components of fatty streaks, a lipid core and a fibrous
cap.
3.4 The Response-to-injury Hypothesis
In response to mechanical injury or exposure to atheogenic stimuli, such as
atherogenic lipoproteins, hyperglycemia, hypertension, and smoking, low density
lipoprotein (LDL) is transported across the injured endothelium and binds to the
extracellular matrix and lipid oxidation is initiated. The accumulated oxidized LDL
stimulates endothelial cells to express adhesion molecules and elaborate growth
factors that lead to recruitment of leukocytes. Leukocytes adhere and migrate into
the vessel wall. Oxidized LDL and leukocytes are then taken up by macrophage
scavenger receptors and develop into foam cells. In turn, foam cells release growth
factors and cytokines (including tumor necrosis factor-alpha (TNF-a), interleukin-
l(IL-l), and interferon gamma (IFN-y)) that promote recruitment of smooth muscle
cells and stimulate neointimal proliferation, continue to accumulate lipid, and further
injure the endothelium (47; 51).
Cycles of accumulation of mononuclear cells, migration and proliferation of
smooth-muscle cells, and formation of fibrous tissue lead to further enlargement and
restructuring of the lesion. Eventually a plaque is formed with a fibrous cap that
overlies a core of lipid and necrotic tissue. At some point, the artery can no longer
compensate by dilation. Plaque rupture or erosion leads to acute cardiovascular
events (47; 51).
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3.5 Function of the Normal Endothelium
Normal endothelial function plays a critical role in regulating vascular
functions as the endothelium is a continuous, single cell membrane that tightly lines
the entire circulatory system. In response to biochemical and mechanical stimuli,
endothelial cells synthesize and elaborate a number of factors that modulate vascular
tone, inflammation, thrombosis, and vascular growth. Factors involved include
vasodilators vs. vasoconstrictors, profibrinolytic factors vs. antifibrinolytic factors,
anticoagulants vs. procoagulants, platelet inhibitors vs. platelet activators, cell
growth inhibitors vs. cell growth promoters and anti-inflammatory factors vs.
proinflammatory factors. A dynamic balance of factors is critical to maintain normal
endothelial function (52).
The normal endothelium provides an antiatherogenic environment that
inhibits platelet and leukocyte adhesion, prevents vasospasm, promotes fibrinolysis,
and inhibits vascular smooth muscle cell growth (53). Among various endothelium-
derived substances, nitric oxide (NO) is one of the most import substances due to its
multi-antiatherogenic functions. By releasing nitric oxide, the endothelium protects
the blood vessel by 1) maintaining vasodilation, 2) inhibiting platelet aggregation
and adhesion, 3) diminishing smooth muscle cell proliferation and migration, and 4)
preventing monocyte adhesion to endothelium and migration into the vessel wall
(37).
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3.6 Mechanisms of Atherosclerosis in Persons with Type 2 Diabetes Mellitus
Atherosclerotic CVD is accelerated in persons with T2DM, resulting in a 2-
to 4-fold increase in the incidence of cardiovascular events (21). The pathogenesis
of atherosclerosis in this particular population is complex and is not fully understood.
Endothelial dysfunction is associated with traditional cardiovascular risk factors,
including hypertension, diabetes mellitus, smoking and dyslipidemia that precede the
onset of atherosclerosis (37). As noted, endothelial dysfunction underlies many
stages in the progression of atherosclerosis (50). Therefore, it is also believed to
play a central role in the pathogenesis of atherosclerosis in persons with T2DM (37).
3.6.1 Endothelial Cell Dysfunction
In persons with T2DM, hyperglycemia, excess free fatty acid release, and
insulin resistance promote adverse metabolic events within the endothelial cell.
Activation of these systems impairs endothelial function, leading to vasoconstriction,
increased inflammation and promotion of thrombosis, eventually resulting in
accelerated atherosclerosis. Details of a proposed model of endothelial dysfunction
in diabetes are shown in Figure 3.1 (adapted from Beckman JA et al.(37)).
In summary, diabetes causes endothelial cell dysfunction by the following
possible mechanisms: 1) reduction of the concentration and bioavailability of nitric
oxide; 2) increase in vasoconstrictors such as endothelin-1 and angiotensin II; 3)
activatation of the transcription factors nuclear factor kB and activator protein 1; 4)
promotion of nonenzymatic glycation and oxidation of lipoproteins; 5) reduction of
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the synthesis of prostacyclin; 6) increase in the expression of endothelial cell
adhesion molecule-1 (ELAM-1) and vascular cell adhesion molecule-1 (VCAM-1)
(54).
3.6.2 Co-existing Conditions
Hypertension and dyslipidemia are two common co-existing conditions
among persons with T2DM. These conditions exacerbate the progression of
atherosclerosis and worsen the prognosis of diabetes in persons with T2DM (38).
Figure 3.1 Endothelial Dysfunction in Diabetes
Hyperglycemia'
, Diabetes Mellitus...
Excess Free Fatty Acids
.
Oxidative Stress
Protein Kinase C Activation
Receptor for Advanced Giycation
End Product (RAGE) Activation
Insulin Resistance
v Nitric Oxide
1 Endothelin-1
t Angiotensin I I
ENDOTHELIUM
t Nitric Oxide
* Activation of NF-kB
* Angiotensin I I
* Activation of
Activator Protein-1
Vasoconstriction
I iypertens'Ori
Vascular Smooth
Muscle Cell Growth
Inflammation
Release of Chemokines
Release of Cytokines
Expression of Cellular
Adhesion Molecules
t Nitric Oxide
f Tissue Factor
♦ Plasminogen Activator
inhibitor-1
t Prostacyclin
Thrombosis
Hypercoagulation
Platelet Activation
Decreased Fibrinolysis
Atherogenesis
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Hypertensive patients with T2DM usually have elevated concentrations of
angiotensin II (an active form of angiotensin that causes profound vasoconstriction
with resulting increase in blood pressure). Angiotensin II binds to specific receptors
on smooth muscle and activates phospholipase C, which leads to increases in
intracellular calcium concentrations and in smooth muscle cell contraction, increased
protein synthesis, and smooth muscle cell hypertrophy and lipoxygenase (an enzyme
that catalyses the oxidative conversion of arachidonic acid to the hydroxyeicosenoic
acid) activity. Moreover, hypertension increases the formation of hydrogen peroxide
and free radicals, which reduce the formation of nitric oxide, increase leukocyte
adhesion and increase peripheral resistance (the resistance to flow of blood in the
systemic circuit) (55; 56).
Abnormal lipid profiles in diabetic patients with CAD are characterized by
elevated triglyceride levels, decreased HDL-C levels, and increased levels of small
dense LDL (57). Overproduction of very low density lipoprotein (VLDL)-
triglyceride may lead to increased uptake of VLDL into the arterial wall (58).
Decreased production of HDL-C may promote LDL oxidation. Small dense LDL
particles are proatherogenic and more readily undergo oxidation, which accelerate
their uptake by monocytes and vascular smooth muscle cells in the vessel wall (59).
In summary, atherosclerosis progression may be accelerated in persons with
T2DM by this cluster of risk factors. Screening and risk factor modification with
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therapeutic interventions can be of great importance in modifying atherosclerosis
progression in this population.
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Literature Review
Chapter 4: Ultrasonographic Measurements of Atherosclerosis
- Common Carotid Artery Intima-Media Thickness
Summary of Key Points
• Common carotid artery intima-media thickness assessed with high
resolution B-mode ultrasonography is a well-established noninvasive
measure of subclinical atherosclerosis
• The automated edge tracking method minimizes measurement error in
CIMT measurement
• Increased carotid intima-media thickness and accelerated progression of
carotid intima-media thickness are associated with increased risk for
cardiovascular morbidity and mortality in nondiabetic populations
• The associations between CIMT, progression of CIMT and the risk of
CVD have not been evaluated in the diabetic population
4.1 Introduction
As a pathological pathway of CVD, atherosclerosis is characterized as a
progressive disease with structural change in the initimal and medial layer of the
vascular system. High resolution B-mode ultrasonography can noninvasively and
precisely quantify structural change in the arterial wall. The validity and reliability
of quantifying intima-media wall thickness of the carotid artery (CIMT) had been
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sufficiently tested to warrant its use as a surrogate endpoint for cardiovascular
morbidity. More importantly, as a reproducible direct measure of degree of
atherosclerosis, CIMT is a well-established noninvasive measure of subclinical
atherosclerosis (60).
4.2 CIMT Measured by Ultrasonography versus Other Atherosclerosis Imaging
Methods
Atherosclerosis can be assessed either invasively by coronary angiography or
intravascular ultrasound, or noninvasively by CT measures of coronary artery
calcification, or B-mode ultrasound imaging of the extracranial carotid arteries (61).
Coronary angiography is most suitable for the assessment of the late stages of
atherosclerosis when stenosis of the lumen becomes apparent. Because of its
invasive characteristics, intravascular ultrasound is usually done at the same time
that a coronary angiogram or angioplasty is being performed. Coronary artery
calcification is a computed tomographic technique to measure the degree of
calcification in atherosclerotic plaques in the coronary arteries. It is noninvasive but
relatively expensive. The associations between coronary calcification and other
measures of subclinical atherosclerosis, and the risk of CVD are under investigation.
Although angiography provides excellent resolution for evaluation of the
vascular lumen, it has major limitations in terms of generalizability to population
samples: 1) it can only ethically be used in person with known or suspected CHD; 2)
cannot assess early stages of atherosclerosis because the growth of atherosclerotic
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lesions initially affects the vessel wall or external arterial diameter without
encroachment into the arterial lumen (50).
In contrast, high resolution B-mode ultrasonography can noninvasively and
precisely quantify structural change in the arterial wall over short repeated intervals
in asymptomatic subjects with low cost. The use of quantitative outcome variables
and repeated measurements over time greatly reduces the necessary sample size for
both epidemiological studies and clinical trials (62; 63). Of the various noninvasive
imaging methods available, only carotid ultrasound is recommended by the
American Heart Association for detection of subclinical atherosclerosis (64).
4.3 Association of Atherosclerosis in Coronary and Carotid Arteries
Atherosclerosis is a systemic disease of the arterial system. Clinical
manifestations of atherosclerosis occur primarily in three large vascular beds:
coronary arteries, lower extremities, and extracranial carotid arteries (37). It is
difficult to image coronary arteries with B-mode ultrasonography because of the
deep anatomical location and constant movement. Therefore, an easily accessible
and representative vascular bed is needed to measure atherosclerosis by noninvasive
ultrasonography.
The carotid arteries are most suitable for study among asymptomatic subjects
because of their superficial localization, size and limited movement. Moreover,
autopsy data from the International Atherosclerosis Project show that the occurrence
of lesions across the major arterial beds parallel each other. The degree of
atherosclerosis in the carotid arteries correlates with that in the coronary arteries and
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the abdominal aorta (65). In addition, carotid and coronary arteries share many risk
factors that contribute to the progression of atherosclerosis and CVD (66; 67).
CIMT is strongly related to atherosclerosis in the coronary arteries (68-70), the
abdominal aorta (71), and the arteries of the lower extremities (72). Thus, CIMT has
been widely accepted as a vascular measure of generalized atherosclerosis.
4.4 Methods for CIMT Measurement
CIMT is meausred using high-resolution ultrasound B-mode imaging on
which two echogenic lines representing the lumen-intima interface and the media-
adventitia interface can be identified. This method was first describled by Pignoli et
al in 1986 (73). These investigators studied in vitro and in vivo specimens of human
aortic and common carotid arteries to determine the feasibility of direct measurement
of arterial wall thickness with B-mode imaging. They observed parallel echogenic
lines separated by a relatively hypoechoic space (the double line pattern) and verified
the anatomic interpretation of these ultrasonographic interfaces. The vessels were
grouped into class A, which were macroscopically normal or had fatty streaks, or
class B, which had atherosclerotic lesions. By using high-resolution real-time
scanners with 7-8 MHz probes, they found B-mode measurement of IMT showed
significant correlations with values obtained by gross pathology and by histology in
both class A and class B specimens. They concluded that B-mode imaging
represents a useful tool for the detection and monitoring of changes in IMT, allowing
the evaluation of changes in the arterial wall in areas without localized plaques.
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CIMT was measured by visual assessment of the leading edges in the earlier
studies with manual tracing of the lumen-intima and the media-adventitia interfaces.
In 1994, Selzer et al reported an automated computerized edge tracking method that
reduced measurement variability due to manual tracing (63). The automated edge
tracking using subpixel interpolation determines edge boundaries at a resolution
greater than monitor line resolution. The resulting CIMT measurement consists of
an average of 80-100 independent measurements made along a 1 cm distance in the
far wall of the distal common carotid artery. Using this methodology, the coefficient
of variation for average CIMT (replicate scans on the same day) is 2.5% within
operators. With this methodology, the variability of CIMT measurement is reduced
2-4 times compared to that obtained with manual tracing methods (74).
4.5 IMT Measured in the Common Carotid Artery versus Internal Carotid
Artery and Bifurcation
IMT measurements of the extracranial carotid arteries include the following
segments: the distal straight 1 cm of the common carotid arteries (CCA), the carotid
bifurcations, and the proximal 1 cm of the internal carotid arteries (ICA) (Figure 4.1
(Adapted from Lonn et al. (77)). Some research groups measure the maximum IMT
on both near and far walls of each segment and then compute the mean maximum
CIMT for these 12 measurement sites (CCA, bifurcation, ICA, near and far walls,
and left and right sides); Other research groups measure IMT only on the far walls of
each segment. The majority of research groups restrict IMT measurements to the
distal far wall of the CCA (75).
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There are several reasons why IMT at the ICA and carotid bifurcation are
included less frequently in IMT studies. First of all, CCA is relatively close and
parallel to the skin surface and therefore is the clearest segment to measure with
good quality scans. The ICA and bifurcation are often difficult to visualize. This
disadvantage leads to many missing images and larger intraobserver and
interobserver variability. For example, in the Asymptomatic Carotid Artery
Progression Study (ACAPS), the walls from the CCA, bifurcation and ICA were
visualized 99%, 88% and 67% of the time, respectively (76).
Figure 4.1 Diagrammatic representation o f right carotid artery scan
Intima
Flow Divider
Adventitia ICA Media'
1 cm 1 cm 1 cm
* The tip o f the flow divider is shown representing the anatomic landmark used to define the carotid artery
segments analyzed. A plaque is seen in the far wall of the bifurcation and internal carotid artery (ICA) segments.
CCA = common carotid artery; ECA = external carotid artery.
Secondly, atheroslcrotic lesions appear later in the CCA than in the ICA or at
the bifurcation (78). Focal atherosclerosis plaque occurs commonly in the ICA but is
seldom found in the CCA. Therefore, CCA IMT is more suitable to assess the early
preintrusive stage of atherosclerosis while ICA IMT better represents the later stage
of atherosclerosis when stenosis of the lumen (intrusive lesion) becomes apparent.
29
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Thirdly, some groups use the mean maximum CIMT for 12 measurement
sites as the primary IMT measurement. Image acquisition and reading of 12 points is
a much more time-consuming process than image acquisition and reading of 1
reliable site. In addition, measurement of common carotid IMT is more reproducible
and more complete than the mean maximum CIMT (75; 79). Furthermore, the mean
maximum CIMT measurement often includes plaques in the ICA and bifurcation.
The definition of a plaque is difficult and varies considerably across studies. This
prohibits drawing firm comparisons of trials using the mean maximum CIMT as the
primary outcome.
Several longitudinal studies demonstrated that far wall CIMT of the common
carotid artery not only correlates well with all major risk factors, but also predicts
cardiovascular events better than the mean maximum CIMT (80-82).
4.6 Measurement of IMT in the Near Wall versus Far Wall of the Carotid
Artery
When comparing ultrasound measurements of CIMT with histology,
ultrasonic measurement of the far wall CIMT accurately represents intima-media
thickness, whereas the near-wall CIMT measures approximately 80% of the
histologically-measured thickness (83-85).
Ultrasonographic imaging of interfaces in arteries is based on the difference
in the acoustic impedance between tissues separated by the interface. The anatomic
location of an interface corresponds to the image only when the ultrasound beam
comes from a less dense tissue to a more dense tissue. When scanning the far wall of
30
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carotid arteries, the tissue density increases from blood, the intimal layer, the medial
layer and to the adventitial layer. The first echogenic line (leading edge) adjacent to
the lumen indicates the interface between blood and the intimal layer. The second
echogenic line corresponds to the anatomic interface between the medial and the
adventitial layer. Thus, the distance from the leading edge of the first echogenic line
to the leading edge of the second echogenic line is the combined thickness of the
IMT (62).
Conversely, when the near wall of the carotid artery is imaged, the ultrasound
beam travels first through a more dense tissue (adventitia) to a less dense (media and
intima) and finally to a least dense tissue (blood). In the near wall, any potential
echo from the adventitia-media interface is lost in the echo produced by the lower
parts of the adventitia. For this reason, the imaged thickness of echogenic lines is
dependent on the gain settings rather than the true IMT (62).
In summary, the far wall of the common carotid artery is the clearest segment
to measure and the easiest segment to image in a reproducible fashion. In addition,
there are no systematic differences in CIMT between the left and the right common
carotid artery (86; 87). Therefore, the distal far wall of the right common carotid
artery is the preferable site for measurement of atherosclerosis progression in
longitudinal studies.
4.7 CVD Risk Factors and CIMT
In the nondiabetic population, early cross-sectional ultrasonographic studies
showed that an increased CIMT is associated with elevated levels of established
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cardiovascular risk factors, such as age, systolic blood pressure (SBP), smoking, total
cholesterol, LDL-C and a decrease in HDL-C (66; 67; 88; 89). The Atherosclerosis
Risk in Communities, a population-based cohort study, demonstrated that CIMT
progression is positively associated with current smoking, pulse pressure, white
blood cell count and fibrinogen while negatively associated with HDL-C (90).
As a high-risk population for CVD, individuals with T2DM are known to
have thicker cross-sectional CIMT (1; 91-93) and accelerated CIMT progression
relative to those without T2DM (90; 94). In one of our earlier publications, we
conducted a cross-sectional analysis among 98 males and 193 females with T2DM,
and found age, SBP, pulse pressure and LDL-C were each significantly positively
associated with CIMT (95). Among the same diabetic sample, our data also
indicated that smoking was associated with increased CIMT after controlling for
other cardiovascular risk factors (96). Few data are available evaluating the
associations between CVD risk factors and CIMT progression among diabetes.
4.8 CIMT and CVD
CIMT is positively associated with the prevalence of CVD in observational
studies (97-99). More importantly, several prospective epidemiologic studies
revealed that CIMT is strongly associated with incident CVD in adults without a
history of CVD (62; 81; 82; 100-104). The Kuopio Ischaemic Heart Disease Risk
Factor Study (KJHD) recruited 1257 eastern Finnish men aged 42-60 years without
CVD and followed them up for 1 month to 3 years (62). For each 100 pm increment
of CIMT, the risk of MI increased by 1.11 (95% Cl 1.06-1.16). The Atherosclerosis
32
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Risk in Communities Study (ARIC) investigated the association between CIMT and
incident MI and coronary death risk over a period of 4 to 7 years in 5552 men and
7289 women, aged 45-64 years and without CVD at study entry. For each 190 pm
increment of CIMT, the risk of MI and coronary death increased by 1.36 (95% Cl
1.23-1.51) in men and 1.69 (95% Cl 1.50-1.90) in women (82). The Cardiovascular
Health Study (CHS) recruited 5888 men and women, aged 65 years or older from
four geographical areas of the United States (103). CIMT measurements were
obtained in 4476 subjects without CVD at baseline. For each 200 pm increment of
CIMT, the risk of MI increased by 1.46 (95% Cl 1.33-1.60). A summary of these
studies is shown in Table 4.1. Since only a single CIMT was measured in these
studies (i.e at study entry), it is unknown whether progression of CIMT is associated
with incidence of CVD.
To date, only one study - The Cholesterol Lowering Atherosclerosis Study
(CLAS) has investigated the association between progression of CIMT and incident
CVD by obtaining serial measures of CIMT using the automated computerized edge
tracing methodology (104). CLAS was a 2-year, randomized serial arterial imaging
clinical trial designed to test the efficacy of LDL-cholesterol lowering in reducing
the progression of atherosclerosis. Upon the completion of CLAS, 146 men 40 to 59
years of age with a previous coronary artery bypass graft were followed for an
33
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Table 4.1: Summary o f Prospective Studies Examining the Association between CIMT and Incident CVD Risk
Study CIMT segment Clinical
Outcomes
Mean Follow-
up (years)
# subjects Age (years) Gender CIMT
increment (p m )1
Relative risk
(95% Cl)
CLAS CCA M IJ 8.8 146 40-59 Men 130 1.4(1.2-1.7)
KIHD CCA & Bulb MI 1 month - 3yrs 1257 42-60 Men 100 1.11 (1.06-1.16)
ARIC 12 sites MI and coronary 5.2 5552 45-64 Men 190 1.36 (1.23-1.51)
death 7289 Women 190 1.69 (1.50-1.90)
ARIC 12 sites Stroke 6.2 6349 45-64 Men <600 vs. >1000 3.6(1.5-9.2)
7865 Women <600 vs. >1000 8.5 (3.5-20.7)
CHS CCA & ICA MI & stroke 6.2 5858 >65 Men & Women 200 1.46 (1.33-1.60)
Rotterdam CCA MI 2.7 7983 > 55 Men 163 1.56 (1.12-2.18)
Study
Women 163 1.44(1.00-2.08)
1. Absolute CIMT increment from baseline to end o f study
2. Coronary events include nonfatal acute myocardial infarction, coronary death, and need for coronary artery revisualization due to recurrence or worsening of
angina pectoris
OJ
average of 8.8 years to ascertain mortality and non-fatal cardiovascular events.
Using the CIMT measured at the end of the trial, an increase in absolute CIMT of
130pm was associated with a 1.4 fold increased risk of coronary events during the
8.8-year follow-up. These results were consistent with findings from the previous
studies cited above. Using all IMT measures obtained throughout the trial to
measure rate of change in IMT (in pm/year), an increase of 30 pm CIMT per year
was associated with a 2.2 fold increased risk for subsequent myocardial infarction or
coronary death. Similar results were found for subsequent risk for any coronary
events (which included nonfatal acute myocardial infarction, coronary death, and
need for coronary artery revisualization due to recurrence or worsening of angina
pectoris). The associations between CIMT, progression of CIMT and the risk of
CVD have not been tested in the diabetic population.
4.9 Limitations Of CIMT Measurements
As reviewed, current ultrasound protocols differ in the segments used for
CIMT measurements. Some studies conduct measurements on both near and far
walls of each segment and compute the mean maximum CIMT for 12 measurement
sites (CCA, bifurcation, ICA, near and far walls, and left and right sides). Other
studies perform measurements only on the far walls of each segment, while most
studies conducted IMT measurements on the distal far wall of common carotid artery
(75). Therefore, it is difficult to interpret and directly compare the results from
different ultrasound trials.
35
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CIMT is a direct measure of atherosclerosis. However, it is not a complete
measure of CVD risk. Because cardiovascular events are ultimately based on
thrombosis of the underlying atherosclerosis, CIMT does not directly measure
thrombosis, but only the risk for thrombosis assessed by elevated levels of
atherosclerosis.
36
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Literature Review
Chapter 5: Randomized Clinical Trials of Antihypertensive and Lipid Lowering
Therapy and Cardiovascular Disease among Persons with Type 2 Diabetes
Mellitus
Summary of Key Points
• Major randomized, placebo-controlled trials demonstrate the substantial
benefit of antihypertensive therapy in reduction of cardiovascular
morbidity and mortality in persons with T2DM.
• The Heart Protection Study (HPS) trial demonstrated a significant
beneficial effect of lipid lowering therapy on CVD risk among diabetic
patients in both primary and secondary prevention. The majority of
other secondary prevention trials also support a positive effect of lipid
lowering therapy on recurrent CVD risk.
5.1 Introduction
As major risk factors for CVD, hypertension and dyslipidemia have been
defined targets for intervention in persons at higher risk of CVD (17). Hypertension
is about twice as prevalent among persons with diabetes than among those without
diabetes (38). Although most patients with T2DM do not have marked elevations of
LDL cholesterol, many of these patients nonetheless carry high enough levels of total
cholesterol and low HDL-C to support the development of atherosclerosis (44).
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Therefore, management of hypertension and dyslipidemia has been emphasized in
persons with T2DM. Numerous randomized clinical trials have examined the benefit
of antihypertensive or lipid-lowering therapy on cardiovascular outcomes in persons
with T2DM.
5.2 Trials Testing the Effect of Antihypertensive Therapy on CVD Risk
Major randomized, placebo-controlled trials testing the effect of specific
antihypertensive therapies on CVD risk among patients with T2DM are summarized
in Table 5.1. Due to the obvious ethical concerns, study medications (either active
treatment or placebo) were taken in addition to conventional antihypertensive
treatments. In general, baseline blood pressure was well matched between the active
treatment and placebo groups. Cardiovascular events measured as trial outcomes
included definite nonfatal or fatal MI, sudden cardiac death, coronary artery bypass
graft surgery, angioplasty, nonfatal or fatal stroke, transient ischemic attack,
aneurysm, and endarterectomy.
The Systolic Hypertension in the Elderly Program (SHEP) was the first
randomized trial of antihypertensive therapy in elderly patients (> 60 years of age)
with isolated systolic hypertension (219 > SBP >160 rnmHg and DBP < 90 mmHg)
(105). A subset of 583 patients with T2DM was randomly assigned to
chlorthalidone 12.5 - 25.0 mg/day or the placebo group for 5 years. The cumulative
5-year total cardiovascular event rate was significantly lower in the treatment group
than in the placebo group (21.4 vs. 31.5 per 100 patient-years, RR = 0.66, 95%CI
0.46-0.94).
38
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Table 5.1 Randomized Trials Testing the Effect o f Specific Antihypertensive Therapies on CVD Risk
Study Year No. T2DM 1 Intervention Inclusion criteria CVD
(%)
Mean Age
(years)
Follow-up
(years)2
No. CV
events3
CV event
rate (%)4
RR (95% Cl)5
SHEP 1996 2 8 3 /3 0 0 chlorthalidone SBP 160-220, DBP 60-90 5 70.4 5 5 7 /8 3 2 0 /2 8 0.66(0.46 - 0.94)
HOT 1998 499/501/501 felodipine DBP 100-115 8.7 61.5 3.8 2 2 /4 5 4 / 9 0.49(0.14-0.78)
UKPDS 1998 7 5 8 /3 9 0 captopril/ atenolol SBP >=160, DBP >=90 0 56.5 8.4 8 2 /6 2 11 /1 6 0.68 (p=0.019)
Syst-Eur 1999 2 5 2 /2 4 0 nitrendipine SBP 160-220, DBP < 95 35 70.2 2 1 3 /3 1 5 /1 3 0.38(0.2-0.81)
HOPE 2000 1808/ 1769 ramipril at least one CVD risk factor 69 65.5 4.5 27 7/351 1 5 /2 0 0.75(0.64 - 0.88)
RENAAL 2001 751 /7 6 2 losartan nephropathy 21 60 3.4 247 / 268 3 3 /3 5 0.90 (p=0.26)
IDNT 2001 5 7 9 /5 6 9 irbesartan nephropathy & SBP >=135, 29 58.8 2.6 1 3 8 /1 4 4 2 4 /2 5 0.91(0.72-1.14)
IDNT 2001 5 6 7 /5 6 9 amlodipine DBP >=85 29 58.7 2.6 1 2 8 /1 4 4 2 3 /2 5 0.88(0.69- 1.11)
1. Number o f subjects with T2DM for intervention / placebo group except for HOT (target DBP<80, <85, <90 mmHg) and UKPDS (tight control / less tight
control)
2. Mean or Median follow-up
3. Numbers o f major cardiovascular events for intervention / placebo group except for HOT (target DBP<80 vs. target DBP <90 mmHg) and UKPDS (numbers of
diabetes related death for tight control / less tight control)
4. Major cardiovascular event rates for intervention / placebo group except for HOT (target DBP<80 vs. target DBP <90 mmHg) and UKPDS (numbers o f diabetes
related death for tight control / less tight control)
5. Relative risk o f cardiovascular events for intervention / placebo group except for HOT (target DBP<80 vs. target DBP <90 mmHg) and UKPDS (relative risk of
diabetes related death for tight control / less tight control)
UJ
VO
The Systolic Hypertension in Europe (Syst-Eur) trial enrolled 492 elderly
patients (> 60 years of age) with systolic hypertension (219 > SBP >160 mmHg and
DBP < 95 mmHg) (106). Subjects were randomized to receive nitrendipine (N =
252) or placebo (N = 240) for a median follow-up of 2 years. Active treatment
significantly reduced all cardiovascular events by 62% (RR = 0.38, 95% Cl 0.2-
0.81).
The Heart Outcomes and Prevention Evaluation (HOPE) study randomized
3577 patients with diabetes (98% were T2DM), who had a previous cardiovascular
event or at least one other cardiovascular risk factor, to ramipril or placebo for a
median follow-up of 4.5 years (107). Approximately 56% of subjects had a history
of hypertension (defined as taking antihypertensive medications or BP> 160/90
mmHg). Ramipril lowered the risk of combined cardiovascular events by 25% (RR
= 0.75, p = 0.0004)
Several studies have specifically compared the effects of different blood
pressure targets on cardiovascular outcomes. The United Kingdom Prospective
Diabetes Study - Hypertension in Diabetes Study (UKPDS-HDS) was the landmark
trial in patients with T2DM investigating the role of blood pressure control and
cardiovascular morbidity and mortality (108). This trial is unique in its large sample
size and long follow-up. Subjects newly diagnosed with T2DM and with elevated
blood pressure were randomly assigned to tight (aiming at BP < 150/85 mmHg, n =
758) versus less tight (aiming at BP < 180/105 mmHg, n = 390) blood pressure
control groups for a median follow-up of 8.4 years. Among the 758 patients
40
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assigned to the tight control group, 400 were allocated to captopril and 358 to
atenolol. Captopril and atenolol were equally effective in reducing blood pressure
and the risk of macrovascular complications. Compared to the less tight control
group, the tight control group yielded lower blood pressure (144/82 vs. 154/87
mmHg), a 24% reduction in diabetes-related end points, a 32% reduction in diabetes-
related death, a 44% reduction in stroke and a 21% reduction in myocardial
infarction (108). For each 10 mmHg reduction in SBP, the incidence of myocardial
infarction was reduced by 11% (p<0.0001) (109). Therefore, the results of the
UKPDS-HDS study strongly supported tight blood pressure control in hypertensive
patients with T2DM. They also suggested that blood pressure reduction itself may
be more important than the type of treatment used.
In the Hypertension Optimal Treatment (HOT) study, 1501 patients with
T2DM and DBP between 100 and 115 mmHg were randomized into three groups
with target DBPs of 90, 85 and 80 mmHg (110). After an average 3.8 years of
follow-up, the target 80 mmHg group experienced a 51% greater reduction in major
cardiovascular events relative to the target 90 mmHg group (RR = 0.49, 95% Cl
0.14-0.78).
The other two randomized, double-blind, placebo-controlled trials recruited
patients with T2DM and nephropathy. In the Reduction of Endpoints in NIDDM
with the Angiotensin II Antagonist Losartan Study (RENAAL), a multinational
study, 1513 subjects were assigned to losartan 50-100 mg/day or placebo for a
median follow-up of 3.4 years (111). In the Irbesartan Diabetic Nephropathy Trial
41
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(IDNT), 1715 subjects were randomized into three groups: irbesartan 300 mg/day,
amlodipine lOmg/day, or placebo for a mean follow-up of 2.6 years (112). Both
trials reported nonsignificantly greater reductions of cardiovascular outcomes in the
treatment group relative to the placebo group. However, enrolling subjects with very
high CVD risk (T2DM plus nephropathy) and short follow up in these two trials
might have biased the results towards the null.
In summary, major randomized, placebo-controlled trials demonstrate the
substantial benefit of antihypertensive therapy in reduction of cardiovascular
morbidity and mortality in persons with T2DM.
5.3 Trials Testing the Effect of Lipid Lowering Therapy on CVD Risk
To date, no studies examining the association between lipid-lowering therapy
and cardiovascular outcomes have been conducted solely in patients with T2DM.
Several randomized trials reported subgroup analysis among subjects with diabetes
(often small sample sizes) using combined cardiovascular events (e.g. cardiovascular
mortality, nonfatal myocardial infarction, stroke, and coronary revascularization) as
the outcomes of interest. Four trials were focused on primary prevention, five trials
were focused on secondary prevention, and two presented data on both. The main
results of these trials are summarized in Table 5.2.
5.3.1 Primary Prevention Trials
The Heart Protection Study (HPS) recruited the largest cohort of diabetic
subjects among all randomized trials to date (113). It included data on both primary
and secondary prevention. A total of 20,536 subjects aged 40-80 years with
42
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Table 5.2 Randomized Trials Testing the Effect o f Lipid Lowering Therapy on CVD Risk
Study Year N of
T2DM1
Intervention Inclusion criteria o f lipid
profile (mg/dl) 2
Age
Range
(yrs)3
Folio
w-up
(yrs)4
No. CV
events5
CV event
rate (%)6
RR (95% Cl)7
Primary Prevention
HHS 1987 59/76 Gemfibrozil non-HDL-C >200 40-55 5 2 / 8 3/11 0.32(0.07- 1.46)
AFCAPS
/TexCAPS 1998 84/71 lovastatin
TC 180-264, LDL-C 130-
190, HDL-C <45, TG<400 40-73 5.2 4 / 6 5 / 8 0.56(0.17- 1.92)
ALLHAT-
LLT 2002 1855/1783 pravastatin LDL-C 120-189, TG<350 >=55 4.8 not reported
.
0.89(0.71 - 1.10)
HPS 2002 2006/1976 simvastatin TC>135 40-80 5 276/367 14/19 0.74(0.64 - 0.85)
PROSPER 2002 191/205 pravastatin TC 154-347, TG<530 70-82 3.2 32/28 17/14 1.23(0.77 - 1.95)
ASCOT-LLA 2003 1258/1274 atorvastatin TC<250 40-79 3.3 38/46 3.0/3.6 0.84(0.55 - 1.29)
Secondary Prevention
CARE 1996 282/304 pravastatin TC<240, LDL-C 115-174 21-75 5 81/112 29/37 0.78(0.62 - 0.99)
4S 1997 105/97 simvastatin TC 212-309, TG<220 35-70 5.4 24/44 23/45 0.50(0.33 - 0.76)
LIPID 1998 396/386 pravastatin TC 155-271 31-75 6.1 76/88 19/23 0.84(0.64- 1.11)
Post-CABG 1999 63/53 lovastatin not specified 21-74 4.3 9/14 14/26 0.53(0.18 - 1.60)
HPS 2002 972/1009 simvastatin TC>135 40-80 5 325/381 33/38
0 .8 9 (0 .7 9 -
1.00)
LIPS 2002 120/82 fluvastatin TC 135-270, TG<400 18-80 3.9 26/31 22/38 0.53(0.29 - 0.97)
PROSPER 2002 112/115 pravastatin TC 154-347, TG<530 70-82 3.2 38/31 34/27 1.26(0.85 - 1.87)
1. Number o f subjects with T2DM for intervention / placebo group except for ALLHAT-LLT (pravastatin vs. usual care control group) and Post-CABG
(aggressive vs. moderate cholesterol-lowering treatment group)
2. non HDL-C: non high-density-lipoprotein cholesterol level; TC: total cholesterol level; TG: triglyceride level; HDL-C: high-density-lipoprotein cholesterol
level; LDL-C: low-density-lipoprotein cholesterol level
3. Age range was given instead o f mean age because most trials did not report mean age for diabetic subgroups.
4. Mean or Median follow-up
5. Numbers o f major cardiovascular events for intervention / placebo group except for Post-CABG (aggressive vs. moderate cholesterol-lowering treatment group)
6. Major cardiovascular event rates for intervention / placebo group except for Post-CABG (aggressive vs. moderate cholesterol-lowering treatment group)
7. Relative risk o f cardiovascular events for intervention / placebo group except for Post-CABG (aggressive vs. moderate cholesterol-lowering treatment group)
■L-
U>
coronary disease or other occlusive arterial disease or diabetes were randomized to
receive simvastatin 40 mg/day or placebo for 5 years. Among 3982 patients with
T2DM and without prior CHD, a significant reduction of cardiovascular events was
observed in the simvastatin group (276 events out of 2006 subjects) relative to the
placebo group (367 events out of 1976 subjects, RR = 0.74, 95% Cl 0.64-0.85).
Another large sample size trial was the Antihypertensive and Lipid-Lowering
Treatment to Prevent Heart Attack Trial (ALLHAT-LLT) (114). It was a
randomized, nonblinded trial designed to examine cardiovascular outcomes with
various antihypertensive and lipid-lowering medications in a large cohort of
hypertensive, CVD-free patients with at least one other CVD risk factor who were 55
years or older. Of the 42,448 persons enrolled, 15,297 had diabetes. Ultimately,
3638 of the diabetic subjects were allocated to the lipid intervention arm. Compared
to 1783 subjects who received usual care, there was a nonsignificant 11% reduction
in coronary heart disease death and nonfatal myocardial infarction among 1855
diabetic subjects receiving pravastatin (RR = 0.89, 95% Cl 0.71-1.10) during the
trial. After 6 years, 16.2% of pravastatin-treated subjects had received no lipid-
lowering medication from their primary physician, whereas 26.1% of usual-care
subjects had received a statin. Thus, the contamination of the usual care control
group complicated the interpretation of the results.
The Helsinki Heart Study (HHS) investigated the effect of gemfibrozil versus
placebo on cardiovascular events in patients with non-HDL cholesterol > 200 mg/dl
for a median follow-up of 5 years (115). The Air Force Coronary Atherosclerosis
44
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Prevention Study/Texas Coronary Atherosclerosis Prevention Study
(AFCAPS/TexCAPS) randomized patients with average cholesterol levels to receive
lovastatin 20mg/d or placebo for a mean follow-up of 5.2 years (116). The Anglo-
Scandinavian Cardiac Outcomes Trial -Lipid Lowering Arm (ASCOT-LLA) studied
atorvastatin 10 mg/day versus placebo in diabetic patients with hypertension and 2 or
more other CVD risk factors and total cholesterol < 250 mg/dl for a mean follow-up
of 3.3 years (117). The first two of these trials had relatively few subjects with
diabetes while the third trial included a large number of subjects with diabetes (Table
5.2). All three trials showed nonsignificant reductions of cardiovascular events in
the active treatment relative to placebo groups. Therefore, while most trials support
a benefit of lipid lowering therapy, the HPS is the only primary prevention trial that
demonstrated a significant effect of lipid lowering therapy on reducing CVD risk.
The HPS is also by far the largest of these primary prevention trials.
5.3.2 Secondary Prevention Trials
The Cholesterol and Recurrent Events (CARE) trial enrolled 4159 patients
aged 21-75 years with a history of acute MI (between 3-20 months prior to study
entry), total cholesterol <240, LDL cholesterol 115-174 and triglyceride <350 mg/dl
(118). Subjects were randomized to pravastatin 40 mg/day or placebo for a median
follow-up of 5 years. Among 586 subjects with T2DM, a significant 25% reduction
in cardiovascular events was observed in the pravastatin group (81 events out of 282
subjects) relative to the placebo group (112 events out of 304 subjects, RR = 0.78, p
= 0.05). The baseline mean LDL cholesterol level was 139 mg/dl in both the
45
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pravastatin and placebo group in the overall sample, but was not reported in the
diabetic subgroup. The trial was the first to show that lipid lowering therapy
significantly reduces the risk of recurrent CVD in diabetic subjects with average
LDL cholesterol levels.
The Scandinavian Simvastatin Survival Study (4S) was a randomized trial
of simvastatin in 4,444 subjects, aged 35-70 years with a prior myocardial
infarction and total cholesterol between 212-309 mg/dl and triglycerides no more
than 220 mg/dl (119). Among 202 patients with T2DM and mean LDL
cholesterol equal to 185 mg/dl at baseline, simvastatin reduced CVD risk (RR =
0.50, 95% Cl 0.33-0.76) relative to placebo after a median follow-up of 5.4
years. The 50% magnitude of reduction in CVD risk in the diabetic subgroup
(50%) was greater than that seen in the nondiabetic subjects (32% risk
reduction). This trial was the first to show that lipid lowering therapy
significantly reduces the risk of recurrent CVD in diabetic subjects with high
LDL cholesterol levels.
The Lescol Intervention Prevention Study (LIPS) randomized patients
(aged 18-80 years) who had undergone percutaneous coronary intervention with
total cholesterol between 135-270, triglycerides > 400 mg/dl to fluvastatin 80
mg/day or placebo for a median follow-up of 3.9 years (120). Among 202
patients with T2DM, the risk of cardiovascular events was lower in the
fluvastatin group (26 events out of 120 subjects) than in the placebo group (31
events out of 82 subjects, RR=0.53, p=0.04).
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In the HPS trial, among 1979 patients with T2DM and with prior CHD,
the risk of cardiovascular events was lower in the simvastatin group (325 events
out of 972 subjects) compared to the placebo group (381 events out of 1009
subjects, RR=0.89, 95%CI 0.79-1.0) (113). Thus, the above four trials together
suggested that lipid lowering therapy reduces recurrent CVD risk in diabetic
patients with a broad range of LDL cholesterol levels.
The Long-term Intervention with Pravastatin in Ischemic Disease
(LIPID) trial randomized subjects with known CVD and average LDL
cholesterol 150 mg/dl to pravastatin 40 mg/day or placebo for a mean follow-up
of 6.1 years (121). The Post-Coronary Artery Bypass Graft (Post-CABG) trial
randomized patients who had undergone coronary bypass graft surgery to
aggressive cholesterol-lowering or moderate lowering for a mean follow-up of
4.3 years (122). Both trials observed nonsignificant reductions of cardiovascular
events in the active lipid lowering compared to the placebo groups. The small
sample size is a concern in the Post-CABG trial.
The Prospective Study of Pravastatin in the Elderly at Risk (PROSPER) also
evaluated both primary and secondary prevention (123). Men and women aged 70-
82 years with at least one CVD risk factor (total cholesterol between 154-347 mg/dl,
triglycerides <530 mg/dl) were randomized to pravastatin 40 mg/d or placebo for a
mean follow-up of 3.2 years. Among 396 subjects with T2DM and without previous
CVD and 227 subjects with T2DM and a positive history of CVD, a nonsignificant
increased risk of cardiovascular events was seen in the pravastatin group (RR=1.23,
47
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95% Cl 0.77-1.95; RR=L36, 95% Cl 0.85-1.87, respectively). Thus, relative risks
were elevated in both primary and secondary prevention groups. PROSPER include
a far older group of subjects than do other trials; the older age of subjects might have
influenced the results.
In summary, the HPS trial demonstrated a significant beneficial effect of lipid
lowering therapy on CVD risk in both primary and secondary prevention. The
majority of other secondary prevention trials also support a positive effect of lipid
lowering therapy on recurrent CVD risk. There are several limitations of the
subgroup analysis among these patients with T2DM. First of all, small sample sizes
in single trials are of concern for most of the trials except ALLHAT-LLT, HPS and
ASCOT-LLA. Secondly, detailed baseline characteristics and the trial changes in
the lipid profile are not reported for the diabetic subgroup in most of the trials
(except 4S and Post-CABG). Thirdly, the target lipid-lowering goal was poorly
defined in most of the trials (except AFCAPS/TexCAPS, 4S and Post-CABG).
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Literature Review
Chapter 6: Randomized Clinical Trials of the Effects of Antihypertensive
Agents and Lipid Lowering Medications on CIMT Progression
Summary of Key Points
• In nondiabetic populations, the beneficial effect of lipid lowering therapy
is conclusive while the antiatherogenic effects of antihypertensive agents
remain controversial.
• Few data are available in patients with T2DM.
• Compared to prevention trials using cardiovascular morbidity and
mortality as the primary end point, the CIMT progression trials require
much smaller sample size, shorter duration of follow-up and also allow
evaluation of therapeutic efficacy among relatively young adults or even
adolescents
6.1 Introduction
As major risk factors for CVD, hypertension and dyslipidemia have been
defined targets for intervention in persons at risk for CVD (17). Higher CIMT (81)
and longitudinally-measured CIMT progression are associated with elevated CVD
risk (81; 104). The validity and reliability of quantifying CIMT has been sufficiently
tested to warrant its use as a surrogate endpoint for cardiovascular morbidity in
addition to a direct measure of atherosclerosis. Therefore, in the past decade, the rate
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of change in CIMT has been increasingly used as a primary outcome in
interventional trials to assess the effect of interventions on subclinical atherosclerosis
(60; 124). A number of randomized, double-blinded, placeo-controlled trials have
tested the antiatherosclerosis effect of antihypertensive agents and lipid lowering
therapy on the rate of change in CIMT in nondiabetic populations. Few data are
available in persons with T2DM.
6.2 Antihypertensive Agents and CIMT Progression
In the non-diabetic population, elevated SBP but not elevated DBP, is
associated with increased CIMT cross-sectionally (65; 125; 126), and with
accelerated CIMT progression (127). Antihypertensive therapy is one of the key
intervention efforts to attenuate atherogenesis and modify CVD risk. Previous trials
have enrolled hypertensive patients to compare the effect of different classes of
antihypertensive agents on CIMT progression (128; 129); other trials have enrolled
primarily normotensive persons to test whether antihypertensive agents provide
additional benefit beyond blood pressure lowering (130-133). CIMT was measured
at study entry and every 6 months during the trials except for the INSIGHT trial
(annual CIMT measurements were conducted in INSIGHT). Ultrasonographers and
the technicians measuring IMT in every study were blinded to treatment assignment
to eliminate information bias. Major randomized clinical trials assessing the effect
of antihypetensive agents on CIMT progression are summarized in Table 6.1. The
results are not consistent.
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Table 6.1 Summary o f Randomized Clinical Trials o f the Effect o f Antihypertensive Agents on CIMT Progression
Trial, Subjects CIMT Interval of Intervention N Mean age T2DM CHD Follow Change of BP CIMT rates P-
Year segment CIMT (years) (%) (%) up (yrs) (m m Hg)1 (pm/year)2 value
________________________________________________ measured______________________________________________________________________________________
Trials testing the effects o f different classes o f antihypertensive agents
MIDAS,
1996 DBP 90-115mmHg Mean-max 3 6 months
isradipine vs.
hydrochloroth
iazide 883 58.5 0 4.1 3
-16.0/-13.0
vs. -19.5/-13.0
(at 6 months) 40/50 0.68
INSIGHT,
2001
BP>150/95 or
SBP>160mmHg with Mean fw
one CVD risk factor cca 4 yearly
nifedipine vs. co-
amilozide 324 65.6 22 18 1-4
-20.0/-12.0
v s.-21.0/-12.0 -0.6/5.1 0.09
Trials comparing the effects of an specific antihypertensive agent
MacMahon et al History o f MI, TIA Mean fw
2000 etc.5 cca 2 years
versus placebo
Ramipril 617 61 6 100 4
-6.0/-5.0 vs.
-1.0/-1.0 7.5/5 0.58
PREVENT,
2000
Angiographic
evidence o f CHD Mean-max 6 months Amlodipine 377 56.9 0 100 3
-6.8/-3.8 vs.
0.0/0.1 -4.2 / 11 0.007
SECURE,
2001
At least one CVD risk
factor Mean-max 2 years Ramipril 732 65.6 1 85 4.5
-4.1/-2.8 vs.
0.1/-0.4 13.7/21.7 0.033
Hosomi et al
2001
T2DM without Mean fw
current use o f ACEIs cca yearly Enalapril 98 56.3 100 62.2 2
-4.3/1.0 vs.
-1.9/0.9 10/20 <0.05
1. Changes from baseline in SBP/DBP for treatment / placebo group except for MIDAS (isradipine vs. hydrochlorothiazide) and INSIGHT (nifedipine vs. co-
amilozide)
2. Annual CIMT progression rate for treatment / placebo group except for MIDAS (isradipine vs. hydrochlorothiazide) and INSIGHT (nifedipine vs. co-amilozide)
3. Mean o f the maximum IMT measured at 12 carotid arterial sites
4. Mean o f the distal far wall o f the common carotid artery
5. Subjects had a history o f hospital diagnosis of acute myocardial infarction (MI), angina with coronary disease confirmed by angiography or exercise
electrocardiogram, transient ischemic attack (TIA) or intermitten claudication were eligible
6.2.1 Trials Testing the Effects of Different Classes of Antihypertensive Agents
on CIMT Progression
The Multicenter Isradipine Diuretic Atherosclerosis Study (MIDAS) was the
first randomized clinical trial using CIMT as a primary outcome in a hypertension
trial (128). In this multi-center, double-blind, randomized trial, 883 nondiabetic
subjects (77.8% men) aged 40 years or more with confirmed hypertension (defined
by DBP 90-115 mmHg) were enrolled, and randomly assigned to either isradipine (N
= 442) or hydrochlorothiazide (N = 441). The rate of progression in the mean
maximum CIMT of 12 carotid arterial sites was measured over 3 years and did not
significantly differ between the two treatment groups (p = 0.68). However, a higher
incidence of CVD was observed in the isradipine group than the hydrochlorothiazide
group (5.65 vs. 3.17%, respectively, p = 0.07). High interobserver variability in
CIMT in this multi-center trial might have biased the results towards the null.
Moreover, for obvious ethical reasons, a placebo group could not be included in the
trial. Therefore, the effect of antihypertensive therapy on the carotid artery remained
unclear.
An ancillary trial to the International Nifedipine GITS Study: Intervention as
a Goal in Hypertension Treatment (INSIGHT) trial, randomized 439 hypertensive
patients (blood pressure > 150/95 mmHg or SBP >160 mmHg) with cardiovascular
risk to nifedipine or co-amilozide (129). The IMT in the far wall of the distal right
common carotid artery was used as the primary outcome measure of atherosclerosis.
A total of 324 subjects had > 3 valid serial CIMT measurements. The annual CIMT
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progression rate differed between nifedipine (- 0.6 pm/year) and co-amilozide (5.1
pm/year) groups (p = 0.09). After adjustment for ultrasound centers, the change in
DBP and baseline covariates including age, sex, BMI, ultrasound plaque and DBP,
the differences between the two groups remained insignificant (p = 0.14).
6.2.2 Trials Comparing the Effects of Antihypertensive Agents versus Placebo
on CIMT Progression
A few prospective, multi-center, double-blind, placebo-controlled,
randomized trials have examined the effect of antihypertensive agents and
progression of CIMT. These trials enrolled primarily normotensive patients and the
proportion of patients who had hypertension at baseline ranged from 36.1% (130) to
50% (131). All hypertensive patients (either in the treatment or placebo group)
recruited were prescribed antihypertensive agents other than study medications by
their primary physicians. The proportions of subjects having hypertension were well
matched between the treatment and placebo groups.
The Prospective Randomized Evaluation of the Vascular Effects of Norvasc
Trial (PREVENT) randomly assigned 377 nondiabetic patients, who had
angiographic evidence of CHD, to amlodipine or placebo for 3 years (133). History
of hypertension and baseline blood pressure was well matched between two groups
(the percentage of subjects who had hypertension at study entry was not reported).
The trial tested whether amlodipine would slow the progression of atherosclerosis
measured by the mean maximum CIMT averaged over 12 carotid segments. Over 3
years, subjects randomized to amlodipine had an average regression of the mean
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maximum CIMT o f -12.6 pm, while the placebo group had an average CIMT
progression of +33 pm (p = 0.007). The CIMT was also compared at specific sites.
The average 3-year change of CIMT in the common carotid artery was -46 pm
(regression) for the amlodipine group versus +11 pm (progression) for the placebo
group (95% Cl on difference in change of CIMT -90 to -24 pm). These results
demonstrated that the antiatherogenic effect of amlodipine is detectable by
ultrasound-measured CIMT progression in nondiabetic patients.
The Study to Evaluate Carotid Ultrasound changes in patients treated with
Ramipril and vitamin E (SECURE), also used the mean maximum CIMT as the
primary outcome measure of atherosclerosis (130). The trial tested the effect of
ramipril on CIMT progression over an average 4.5 years of follow-up in 732 patients
who had cardiovascular risk factors (including diabetes). Two doses of ramipril
(2.5mg and lOmg daily) were compared to placebo. At baseline, 36.1% of subjects
had hypertension (the definition of hypertension was not provided), 84% had a
positive history of CHD and only 1% was diagnosed with diabetes. The CIMT
progression rate was 21.7,18.0 and 13.7 pm/year in the placebo, ramipril 2.5 mg/d
and ramipril 10 mg/d groups, respectively (p = 0.033 among groups). After
adjustment for age, sex, baseline IMT, history of high total or low HDL cholesterol,
history of smoking, history of hypertension, baseline total and LDL cholesterol, use
of lipid-lowering drugs and blood pressure changes during the trial, the beneficial
effect of ramipril on atheroscleorisis progression remained statistically significant (p
= 0.046). Because the treatment group differences in CIMT progression remained
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significant after reduction for changes in blood pressure, the investigators suggested
the beneficial effect of ramipril on atherosclerosis is not fully explained by blood
pressure lowering and may be additionally related to a direct vascular protective
effect.
Another randomized trial, conducted in New Zealand, studied the effect of
ramipril on carotid atherosclerosis in 617 patients with a history of CVD within 5
years of enrollment (132). About 6% of subjects had diabetes at baseline. Subjects
who had blood pressure > 160/100 mmHg or SBP <100 mmHg were excluded.
IMT was measured in the far wall of the distal right common carotid artery. After 4
years of follow-up, no statistically significant difference in the average changes from
baseline to follow-up in CIMT was seen between the ramipril (30 pm, N = 308) and
placebo group (20 pm, N = 309, p = 0.58). The investigators concluded the study
provided no evidence of any substantial effect of ACEIs on carotid atherosclerosis
among subjects with CVD who were not hypertensive.
A single-center, double-blind, randomized trial examining the efficacy of
enalapril for preventing CIMT thickening was conducted in 98 Japanese patients
with T2DM (131). The CIMT was averaged over measurements on both the near
and far walls in the right and left common carotid artery. About 50% of subjects had
blood pressure >160/95 mmHg or were taking antihypertensive medication at
baseline. Randomized treatment with enalapril lOmg/d (N = 48) reduced CIMT
progression over 2 years relative to the placebo group (N = 50) (10 vs. 20 pm/year,
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p<0.05). This has been the only trial to date to evaluate the efficacy of
antihypertensive agents on CIMT progression exclusively in persons with T2DM.
In summary, previous trials have yielded inconsistent results in terms of the
effect of antihypertensive agents on CIMT progression. MIDAS and INSIGHT both
enrolled hypertensive patients and tested the effect of different classes of
antihypertensive agents on CIMT progression and both yielded insignificant results.
However, the results from these two trials only indicated no detectable difference of
the antiatherosclerosis effect on CIMT progression between two antihypertensive
agents. The exact effect of antihypertensive agents on CIMT progression remained
unclear.
On the other hand, four trials enrolled both normotensive and hypertensive
persons and compared the effect of an antihypertensive agent versus placebo on
CIMT progression. Three out of four trials yielded significant results.
6.2.3 Critique of Trials Testing Antihypertensive Agents
There are several weaknesses in the trials just described. First of all, the
average annual CIMT progression rates differ considerably across studies due to
different ultrasound protocols, methods of CIMT measurement employed and
different study populations. Thus direct comparison of results is not possible.
Secondly, because plaque occurs more often in the ICA and bulb than in the
common carotid artery, measurement of the mean maximum CIMT over 12 carotid
segments includes thicker lesions in the bifurcation and the ICA (i.e. later stage
atherosclerosis), in addition to the wall thickening characteristic of early
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atherosclerosis. Moreover, IMT measurement on the ICA and bifurcation tends to
have many missing images. The incomplete data might bias the results and the
directions of the biased results are not predictable.
Thirdly, multi-center trials require more than one ultrasonographer to scan
the images. This increases the variability in study measurement, and thus increases
measurement error. In addition, manual tracing of the lumen-intima and the media-
adventitia interfaces (used in some CIMT measurement protocols) increases
variability and measurement error.
Previous trials enrolled mainly nondiabetic subjects. However, it is known
that atherosclerosis is accelerated in persons with T2DM. Small numbers of subjects
with diabetes precluded subgroup analysis in the diabetic sub-sample. The
significant antiatherogenic effect of enalapril observed in a small sample of Japanese
patients with T2DM remains to be confirmed by larger trials.
6.3 Trials Testing the Effects of Lipid Lowering Therapy on CIMT Progression
In the non-diabetic population, elevated total cholesterol, low-density
lipoprotein cholesterol and decreased high-density lipoprotein cholesterol are
associated with increased CIMT cross-sectionally, and with accelerated CIMT
progression(127). Lipid lowering therapy is one of the key intervention efforts to
attenuate atherogenesis and reduce CVD risk (17). Results of numerous trials show
that the effect of lipid-lowering therapy on atherosclerosis progression is detectable
with the progression of CIMT assessed by ultrasonography (134-143). Most of these
trials were conducted in a secondary prevention setting. Table 6.2 summarizes the
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Table 6.2 Summary o f Randomized Trials o f the Effect o f Lipid Lowering Intervention on CIMT Progression
Trial,
Year
Subjects 1 CIMT
segment
Interval of
CIMT
measured
Intervention N Mean T2DM Men
age(yrs) (%) (%)
CHD
(%)
Follow-
up (yrs)
Change o f LDL-
C (%) by
treatment2
CIMT rates P-value
(pm/year)3
Primary Prevention Trials (lipid lowering medications vs. placebo)
AC APS, LDL-C mean
1994 130-189 m ax4 6 months lovastatin 919 61.3 2.3 49.3 0 3 -38.4 (25%) -9 /6 0.001
CAIUS, LDL-C mean fw
1996 150-250 cca5 6 months pravastatin 305 55.3 0 53 0 3 -39.8 (22%) -3.2/7.7 0.0047
KAPS, max fw
1995 LDL-C > 155 cca 6 months pravastatin 447 57.5 1.8 100 6.3 3 -54.0 (27%) 9.6/28.5 0.0019
Secondary Prevention Trials (lipid lowering medications vs. placebo)
CLAS, mean fw colestipol-
1993 Had CABG cca 2 years niacin 78 51 0 100 100 4 XXX -26/18 0.002
PLAC-II, Mean LDL-C mean fw Not Not Not
1995 167.5 6 cca 6 months pravastatin 151 given given given 100 3 -47.2 (28%) 29.5 /45.6 0.03
MARS, mean fw
1996 TC 190-295 cca 6 months lovastatin 188 58 0 92 100 2 -70.6 (45%) -38/19 <0.001
REGRESS, mean fw
1995 TC 155-310 cca 6 months pravastatin 255 56 0.1 100 100 2 -48.1 (29%) -10 / -15 0.19
LIPID, mean fw
1998 TC 155-271 cca 2 years pravastatin 552 61 5 88 75 4 -41.7 (28%) -3.7/16 <0.0001
Comparison of Different Classes o f Statins
ASAP, mean fw atorvastatin vs. -166.8(51%) vs.
2001 LDL-C > 174 cca yearly simvastatin 325 48.5 1.8 40 26 2 -135.5 (41%) -20.5 / -9 0.07
ARBITER, mean fw atorvastatin vs. -72.0 (49%) vs.
2002 T C > 160 cca 6 months pravastatin 161 60 10 71.4 46 1 -75.0 (29%) -34 / 25 0.03
1. LDL-C: low density lipoprotein cholesterol in mg/dL; TC: total cholesterol in mg/dL; CABG: coronary artery bypass graft surgery
2. Changes from the baseline in LDL cholesterol and percentage o f change (changes in LDL-C divided by LDL-C levels at baseline) in groups received lipid
lowering medication except for ASAP (atorvastatin vs. simvastatin) and ARBITER (atorvastatin vs. pravastatin)
3. Annual CIMT progression rate for treatment / placebo group except for ASAP (atorvastatin vs. simvastatin) and ARBITER (atorvastatin vs. pravastatin)
4. Mean o f the maximum IMT measured at 12 carotid arterial sites
5. Mean o f the distal far wall o f the common carotid artery
6. LDL cholesterol with the 60th and 90th percentile for the U.S. population by age and sex
CO
oo
key design features and main results from all relevant trials. In general, CIMT was
measured at study entry and every 6 months during the trial unless otherwise
specified.
6.3.1 Primary Prevention Trials
Two primary prevention trials have demonstrated that reduction in the
progression of CIMT with lipid-lowering therapy is also accompanied by a reduction
of cardiovascular events. In the Asymptomatic Carotid Artery Progression Study
(ACAPS), 919 asymptomatic men and women aged 40-79 years who had at least one
carotid artery lesion of > 1.5mm and with mean LDL cholesterol of 158 mg/dl were
randomized to lovastatin (N = 460) or placebo (N = 459) for 3 years (135). The
annual progression rate of the mean maximum CIMT (of 12 segments) for the
lovastatin group and the placebo groups were significantly different (-9 vs. +6
pm/year, p = 0.001). The cumulative CVD risk differed significantly (p = 0.04),
favoring the lovastatin group. AC APS was the first and the largest multi-center trial
to demonstrate the benefit of lipid lowering therapy on both atherosclerosis
progression and CVD events in asymptomatic subjects with moderately elevated
LDL-C.
In the Kuopio Atherosclerosis Prevention Study (KAPS), 447 subjects aged
44-65 years with elevated LDL-C (> 154 mg/dl) were randomized to pravastatin or
placebo for 3 years (single-center) (136). About 10% of subjects had a previous
myocardial infarction at study entry. The annual progression rate of the common
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carotid artery far wall CIMT differed significantly between the pravastatin and the
placebo group (9.6 vs. 28.5 pm/year, p = 0.019).
A third primary prevention trial was conducted in 305 asymptomatic men and
women with moderate hypercholesterolemia. The Carotid Atherosclerosis Italian
Ultrasound Study (CAIUS) also found a lower rate of change in the mean maximum
CIMT in 151 pravastatin-treated subjects compared to 154 placebo-treated subjects
(-4.3 vs. +9.0 pm/year, p < 0.0007) (139). A similar significant difference was found
in the rate of change in the common CIMT (-3.2 vs. +7.7 pm/year, p = 0.0047).
6.3.2 Secondary Prevention Trials
Several randomized clinical trials have demonstrated that lipid lowering
therapy retards CIMT progression in patients with angiographically confirmed CAD.
In the Cholesterol Lowering Atherosclerosis Study (CLAS), 146 nonsmoking men
with previous coronary artery bypass graft surgery, aged 40-59 years were
randomized to colestipol-niacin or placebo. A total of 78 subjects completed four
treatment years with ultrasound examination at baseline, 2 and 4 years (134). The
annual progression rate of CIMT (assessed on the far wall of the right common
carotid artery) was -26 pm/year for the treatment group and +18 pm/year for the
placebo group (P = 0.002). This study was the first randomized, controlled clinical
trial to demonstrate the significant therapeutic effects of lipid-lowering intervention
on early, preintrusive atherosclerosis by ultrasound assessed CIMT.
In the Monitored Atherosclerosis Regression Study (MARS), 188 patients
aged 37-67 years with total cholesterol of 190-295 mg/dl who had CAD identified on
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coronary angiography received lovastatin 80 mg or placebo for up to 4 years (138).
The annual rate of change in CIMT showed an average reduction in the lovastatin
group and increase in the placebo group (-38 vs. +19 pm/year measured over 2 years,
and -28 vs. +15 pm/year measured over 4 years, respectively) (p < 0.0001).
In the Pravastatin, Lipids, and Atherosclerosis in the Carotids (PLAC-II) trial
(single-center), 151 patients with coronary artery disease (coronary artery disease
was defined as a history of a heart attack or cardiac catheterization demonstrating
>50% stenosis of at least one coronary artery) with at least 1 qualifying carotid artery
lesion of > 1.3 mm were randomized to receive pravastatin or placebo for 3 years
(140). The rate of progression of the mean maximum CIMT, the primary endpoint,
was nonsignificantly reduced with pravastatin (59.3 vs. 67.5 pm/year in the
pravastatin and placebo group, respectively, p = 0.44). However, significant
reduction with pravastatin was seen in the common carotid artery IMT (29.5 vs. 45.6
pm/year in the pravastatin and placebo group, respectively, p = 0.03).
The results of the above three trials demonstrate that progression of
subclinical atherosclerosis can be significantly reduced with lipid-lowering therapy,
and this antiatherogenic therapeutic effect can be detected in small-scale trials by the
change in measurements of CIMT.
In the Regression Growth Evaluation Statin Study (REGRESS), 255 men
aged < 70 years, who had a history of coronary arteriography showing at least one
stenosis > 50% in a major coronary artery and with total cholesterol of 155-310
mg/dl (4-8 mmol/L) were randomized to pravastatin 40 mg/day or placebo for 2
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years (141). The mean femoral IMT decreased 60 pm (absolute change) in the
pravastatin group and increased 130 pm in the placebo group over 2 years (p =
0.004). No significant difference in the change of the mean carotid IMT was
observed between the two groups (p = 0.19).
In the Long-term Intervention with Pravastatin in Ischemic Disease (LIPID)
trial, 552 individuals with a history of CHD but with average or below average total
cholesterol levels (154 to 270 mg/dl) were studied (143). CIMT was measured at the
right distal common carotid artery far wall. After 4 years of follow-up, the mean
CIMT increased 48 pm (absolute change) in the placebo group and decreased 14 pm
in the pravastatin group (p < 0.0001). The beneficial effects of pravastatin on CIMT
progression appeared to be similar among three strata defined by tertiles of
pretreatment total cholesterol.
6.3.3 Trials Comparing Different Classes of Statins
In the Atorvastatin vs. Simvastatin on Atherosclerosis Progression (ASAP)
trial, 325 patients aged 30-70 years who were untreated for familial
hypercholesterolemia or had LDL-C >174 mg/dl received atorvastatin 80 mg/day or
simvastatin 40 mg/day (142). About 31% of subjects had a history of CVD at
baseline. Over 2 years, the mean maximum CIMT was reduced by 31 pm in the
atorvastatin group but increased by 36 pm in the simvastatin group (p = 0.0001). A
borderline significant difference of the absolute change in the common carotid IMT
was seen between the atorvastatin group (-41 pm) and the simvastatin group (-18
pm, p = 0.07). The atorvastatin group also showed greater reduction in LDL-C
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concentration (-4.32 mmol/L) than the simvastatin group (-3.51 mmol/L, p =
0.0001).
The Arterial Biology for the Investigation of the Treatment Effects of
Reducing Cholesterol (ARBITER) trial compared the effects of natural statin (i.e.
pravastatin) and synthetic statin (i.e. atorvastatin) (137). A total of 161 patients
(46% had known CVD) who met the NCEP II criteria for pharmacological lipid
lowering therapy were randomized to open-label treatment with either pravastatin 40
mg/d or atorvastatin 80 mg/d. The primary endpoint was the average CIMT
measured at the far wall of the right and left distal common carotid artery. Over 12
months, a significant difference in the change of CIMT was seen between the
pravastatin and atorvastatin group (+25 vs. -34 pm, p = 0.03).
6.3.4 Summary of Trials of Lipid Lowering Therapy on CIMT Progression
These trials shared several key design features. All trials provided CIMT
measurement at the far wall of the distal right CCA (except ACAPS which measured
the mean maximum CIMT of 12 segments). Despite the variation in study
populations and CIMT progression rates across trials, all placebo-controlled trials
observed a significantly lower CIMT progression rate in the lipid-lowering group
versus the placebo group. Due to the small number of diabetic subjects recruited, no
trial had sufficient power to do a subgroup analysis on the diabetic subgroup.
Nevertheless, consistent results from these trials support the use of ultrasound-
measured CIMT as the primary outcome in the trials investigating the effects of
lipid-lowering therapy on atherosclerosis progression in nondiabetic populations.
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6.4 Conclusions
In summary, in nondiabetic populations, the beneficial effect of lipid
lowering therapy on atherosclerosis progression is conclusive while the
antiatherogenic effects of antihypertensive agents remain controversial. Few data are
available in patients with T2DM. This particular population is known to have
thicker CIMT than nondiabetic subjects of the same age (91-93; 144).
Longitudinally, the progression of CIMT is accelerated in persons with established
T2DM relative to those without diabetes (90; 94).
Based on a large body of evidence and the underlying cardiovascular risk
associated with diabetes, aggressive blood pressure control and lipid lowering is
recommended for patients with T2DM (17). The therapeutic efficacy of different
strategies on CVD and atherosclerosis progression remains to be addressed by
interventional trials in an efficient and cost-effective manner. In order to observe
enough events, prevention trials using cardiovascular morbidity and mortality require
large sample sizes, long duration of follow-up and subjects with older age and higher
risk of CVD at study entry. In contrast, trials using CIMT progression as an
endpoint require much smaller sample sizes, shorter duration of follow-up and also
allow evaluation of efficacy among relatively young adults or even adolescents. The
latter advantage is critical because of the increasing incidence of T2DM and
prevalence of known CVD risk factors in American youth (15).
Therefore, we plan to investigate the association between antihypertensive
and lipid lowering therapy and progression of CIMT among persons with T2DM,
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using data from the Troglitazone Atherosclerosis Regression Trial (TART), a
randomized clinical trial designed to determine whether troglitazone (400mg daily)
reduces the progression of subclinical atherosclerosis in persons with insulin-
requiring type 2 diabetes mellitus.
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Data Analysis and Planned Publications
Chapter 7: Antihypertensive Therapy Reduces Progression of Subclinical
Atherosclerosis In Patients With Type 2 Diabetes Mellitus
Summary of Key Points
• Antihypertenisve therapy reduces the incidence of cardiovascular events
in people with diabetes. However, the effects of treatment on subclinical
atherosclerosis are less well studied.
• To investigate the association between antihypertensive therapy and
progression of CIMT among persons with T2DM, we conducted a post
hoc cohort analysis of data from the Troglitazone Atherosclerosis
Regression Trial
• Our data indicate that regular use of antihypertensive agents reduces the
harmful impact of elevated blood pressure on atherosclerosis
progression in type 2 diabetes. The antiatherogenic effect of
antihypertensive agents, including blood pressure normalization and
possible direct vascular wall protection, can be detected by CIMT
progression obtained with B-mode ultrasound in patients with type 2
diabetes.
6 6
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7.1 Introduction
Diabetes is a major risk factor for cardiovascular disease (CVD).
Hypertension is approximately twice as prevalent among persons with compared to
people without diabetes(38). According to the Third National Health and Nutrition
Examination Survey (1988-1994), 71% of U.S. adults with diabetes have elevated
blood pressure (> 130/85 mmHg) or are taking prescription drugs for hypertension
(39). Individuals with type 2 diabetes mellitus (T2DM) who are hypertensive have
an approximate 2-fold greater risk of cardiovascular events than individuals with
T2DM who are noimotensive (40). The mortality rate for hypertensive diabetic
patients is three times the mortality of normotensive diabetic patients (41).
Many studies have investigated the association between blood pressure
control and CVD events in both nondiabetic and diabetic populations (145).
However, few studies have examined the association between antihypertensive
agents and subclinical atherosclerosis among hypertensive patients with T2DM.
Carotid artery intima-media thickness (CIMT) acquired with high-resolution B-mode
ultrasound is a well-established noninvasive measure of subclinical atherosclerosis.
Progression of CIMT is associated with the progression of coronary artery disease
(69) and risk for CVD morbidity and mortality in non-diabetic populations (81; 82;
104).
To investigate the association between antihypertensive therapy and
progression of CIMT among persons with T2DM, we conducted a post hoc cohort
analysis of data from the Troglitazone Atherosclerosis Regression Trial (TART).
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7.2 Methods
TART was a single-center, randomized, double-blind, placebo-controlled
trial designed to determine whether treatment with a PPARy agonist reduces the
progression of atherosclerosis in subjects with insulin-requiring T2DM. Details of
the study design have been described (95). Briefly, eligible subjects had age at
diagnosis of T2DM > 30 years, fasting serum glucose 7.8-19.4 mmol/L, and were
either using exogenous insulin or were scheduled to be placed on insulin therapy.
After a 8-12 week run-in period during which diabetes care was intensified with
insulin, 299 subjects were randomized to additional treatment with placebo (N=150)
or troglitazone 400 mg daily (N=T49) for two years. The primary trial end point
was the rate of change in CIMT. The Institutional Review Board of the University
of Southern California approved the study and all subjects provided written informed
consent.
CIMT was measured at baseline and every six months using high resolution
B-mode carotid artery ultrasound (ATL ultrasound imager equipped with a linear
array 7.5 MHz probe). Automated edge detection was used to locate the lumen-
intima and media-adventitia echo boundaries at sub-pixel resolution (Prowin, patent
pending) (104). An average of 80-100 measurements were made along a 1-cm
distance of the distal right common carotid artery far wall and averaged to derive the
CIMT measurement. Using this methodology, the coefficient of variation for
average CIMT (replicate scans obtained approximately 2-weeks apart) was less than
1% in this study.
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After randomization, clinic visits occurred at two and four weeks, then
monthly until six months, then bimonthly until 24 months. The target blood pressure
range was 110-140/80-90 mmHg. Subjects with blood pressure greater than 140/90
mmHg were given an angiotensin-converting enzyme inhibitor, followed by a long-
acting calcium channel blocker and then an alpha blocker when necessary. Of note,
antihypertensive agents were prescribed as needed rather than given as a randomized
intervention. Insulin doses were adjusted at each visit to maintain fasting and V z-
hour pre-meal blood glucose levels between 5.5-8.3 mmol/L. A medication record
form was used to record the use of all prescription medications, including
antihypertensive agents (dose, frequency and dates of use). Subjects were requested
to bring all medications to each clinic visit.
Subjects were interviewed with structured questionnaires at baseline to
ascertain age, gender, ethnicity, years of education, age at diagnosis of T2DM, age at
initiation of insulin therapy and current daily insulin dose. Height, weight, waist and
hip circumference were measured at baseline and at each clinic visit during the trial.
Body mass index (BMI) and waist-hip-ratio were derived from these data. BMI was
calculated as weight (kilograms)/height (meters)2.
At each visit fasting blood samples were obtained. Serum glucose levels
were measured by glucose oxidase (Beckman Glucose analyzer II, Beckman
Instruments, Brea CA, USA). Hemoglobin A le levels were measured by high-
pressure liquid chromatography (BioRad, Hercules CA, USA). Plasma total
cholesterol, total triglyceride, and high-density lipoprotein (HDL) cholesterol levels
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were determined by enzymatic assays and standardized to the CDC using the Lipid
Research Clinics protocol. Low-density lipoprotein (LDL) cholesterol levels were
estimated using the Friedewald equation (146).
Systolic blood pressure (SBP) and diastolic blood pressure (DBP) were
obtained using a Dinamap (Critikon, Inc.) system. Subjects were seated for at least 5
minutes prior to blood pressure measurements with the midpoint of the upper arm at
the level of the heart, approximately the fourth intercostal space. The bladder inside
the cuff encircled at least 80% of the arm. At each visit, SBP and DBP were
computed as the average of two measures taken 5 minutes apart.
Multivariate mixed-effects models were used to evaluate the rate of change in
CIMT in relation to blood pressure and the duration of use of antihypertensive
agents. Separate sets of mixed-effect models were fitted for SBP and DBP. CIMT
was regressed on follow-up time (years since randomization) to estimate the average
rate of change in CIMT in each subject. The duration of antihypertensive medication
use was included as an independent variable and SBP (or DBP) was included as
time-varying independent variable. Interaction terms (follow-up time* SBP)
evaluated whether blood pressure influenced the CIMT progression rate. Whether
use of antihypertensive agents modified the influence of blood pressure on CIMT
progression was tested by three-way interaction terms (follow-up time* SBP
*duration of antihypertensive therapy). Age, treatment group, and change from
baseline in fasting glucose were included as potential confounders. Continuous
variables were centered on their means. Analyses were conducted using Statistical
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Analysis System version 8.0 (SAS Institute, Cary, NC) and SAS PROC MIXED for
the mixed-effects models. Data are presented as mean (sd) in tables and text.
Statistical significance was accepted at two-sided p = 0.05.
7.3 Results
A total of 299 subjects were randomized to the TART and 276 subjects (142
troglitazone, 134 placebo) who had at least one follow-up CIMT measurement were
included in the present analysis. The mean age at randomization was 52.5 years
(range 30 to 70 years). Eighty-nine percent of the subjects were of Hispanic origin
and 67% were female. The mean age at diagnosis of T2DM was 42.7 (8.6) years and
the mean duration of diabetes was 9.8 (6.3) years. Sixty-six percent of the sample (n
= 182) had never smoked regularly. Among 94 (34%) subjects who had smoked
daily for at least 6 months, 63 (23%) were former smokers and 31 (11%) were
current smokers. Table 7.1 summarizes the baseline characteristics of the 90 men
and 186 women at the screening visit and the randomization visit.
On average, these subjects were obese and normotensive although almost half
of the sample was on antihypertensive agents, 9% was on lipid modifying therapy at
the screening visit. Elevated fasting plasma glucose, HbAlc and lipid profiles were
improved after approximately 8-12 week run-in phase. At the end of the run-in
phase (i.e. at randomization to treatment), 55%, 6% and 23% of the sample were
taking antihypertensive agents, lipid lowering medications and anti-inflammatory
medications, respectively.
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Table 7.1 Characteristics o f TART Subjects at the Initial Screening Visit and at Randomization
Screening Visit (n=276) Randomization (n=276)
CIMT (pm) 814.4(154.2)1 814.1 (154.4)
Age (years) 52.2 (9.1) 52.5 (9.0)
Hemoglobin A le (%) 9.8 (2.3) 7.5 (1.1)
Fasting glucose (mmol/L) 11.6(4.2) 7.9 (2.1)
Daily insulin dose (U) 53.0 (29.5) 60.2 (33.3)
Systolic blood pressure (mmHg) 135.7(20.0) 130.9(19.0)
Diastolic blood pressure (mmHg) 79.7(10.2) 76.2 (9.4)
Body mass index (kg/m2 ) 31.6 (6.0) 32.0 (6.0)
Weight (kg) 81.3 (17.8) 82.3 (17.6)
Total cholesterol (mmol/ L) 5.2(1.1) 4.4 (0.9)
Triglycerides (mmol/ L) 2.0 (1.2) 1.6 (1.0)
HDL-cholesterol (mmol/ L) 1.3 (0.3) 1.2 (0.3)
LDL- cholesterol (mmol/ L) 2.9 (0.9) 2.6 (0.8)
Taking antihypertensive agents 120 (43) 2 151 (55)
Taking lipid lowering medications 24(9) 16(6)
Receiving insulin therapy 228 (83) 276 (100)
Taking anti-inflammatory medications 52 (19) 64 (23)
1. Mean (SD)
2. N (%)
During the 2-year trial, 184 (67%) of the 276 subjects used antihypertensive
agents. Among the subjects who used antihypertensive agents, 84 (46%) used
angiotensin-converting enzyme inhibitors alone, 85 (46%) used angiotensin-
converting enzyme inhibitors combined with other classes of antihypertensive agents
and 15 (8%) used a calcium channel blocker or a diuretic alone. These proportions
did not differ between troglitazone and placebo groups (p = 0.91). Among subjects
who used antihypertensive agents, the duration of use of antihypertensive agents
ranged from 0.02 to 2.2 years with a median of 1.8 years. Subjects who used
antihypertensive agents had higher baseline CIMT than those who did not (835.1
(159.6) pm vs. 773.0 (134.3) pm, p = 0.002). However, adjustment for age
eliminated the significant difference (adjusted mean (SD): 818.7 (141.1) pm vs.
805.7 (142.9) pm, respectively, p = 0.49). Among 184 subjects receiving
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antihypertensive therapy during the trial, the mean blood pressure was 142/82 mmHg
at the initial screening visit, when 120 subjects were already taking antihypertensive
agents. The mean blood pressure of the 184 subjects during the trial was reduced
significantly from the screening visit, by 7.2 (16.3) mmHg for SBP (p<0.0001) and
by 6.4 (8.8) mmHg for DBP (p<0.0001).
In a single risk factor midxed effects model, higher SBP was associated with
a greater CIMT progression rate (p-value for follow-up time*SBP = 0.03, data not
shown). We present the multivariate mixed effects model in Table 7.2. The
regression coefficient associated with follow-up time represents the average rate of
change in CIMT. During treatment with antihypertensive medications, the
association between SBP and CIMT progression was significantly attenuated in a
duration-dependent manner (p-value for follow-up time*SBP*duration of
antihypertensive therapy = 0.035).
Table 7.2 Parameter Estimates Related to the CIMT Progression Rate from the Multivariable Mixed Effect Model
Effects CIMT progression rate (pm/year)
attributable to each effect
SE P-value
Follow-up time (pm/year) 7.54 1 1.99 0.0002
Follow-up time*SBP (pm/year per mmHg) 0.17 2 0.05 0.002
Follow-up time*Duration o f therapy3
(pm/year per year o f therapy) -2.414 1.49 0.11
Follow-up time*SBP*Duration of therapy
(pm/year per mmHg per year of therapy) -0.08 5 0.04 0.035
1. average increase in CIMT progression rate for SBP centered at 130 mmHg without anithypertensive therapy
2. average increase in CIMT progression rate for each ImmHg increase in SBP without antihypertensive
herapy
3. duration of antihypertensive therapy during the trial (years)
4. average decrease in CIMT progression rate for each 1-year use o f antihypertensive agents
5. at matched SBP, average decrease in CIMT progression rate for each 1-year use o f antihypertensive agents
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The fitted regression model predicted the CIMT progression rate as follows:
the CIMT progression rate was approximately twice as great for SBP = 150mmHg
(7.0 pm/year, SE = 1.5, 95% Cl (4.1, 9.9)) than for SBP = llOmmHg (3.3 pm/year,
SE = 1.5, 95% Cl (0.4, 6.2)) with one year of antihypertensive therapy, while the
rates did not differ significantly between SBP = 150mmHg and SBP = 1 lOmmHg
with two years of antihypertensive therapy (3.0 pm/year, SE = 2.0, 95% Cl (-0.9,
6.9) vs. 2.4 pm/year, SE = 2.4, 95% Cl (-2.3, 7.1)) (Figure 7.1).
Figure 7.1 Predicted annual CIMT progression rates in relation to SBP and the duration o f antihypertensive
therapy from the multivariable mixed-effect model. Shaded bars for SBP = 1 5 0 mmHg, open bars for SBP =
110 mmHg. a. Without taking antihypertensive agents, the predicted CIMT progression rate was 10.9 pm/year
for SBP = 150 mmHg, 4.0 pm/year for SBP = 1 1 0 mmHg. b. Having taken antihypertensive agents for 1 year,
the predicted CIMT progression was 7.0 pm/year for SBP = 150 mmHg, 3.3 pm/year for SBP = 1 1 0 mmHg. c.
Having taken antihypertensive agents for 2 years, the predicted CIMT progression was 3.0 pm/year for SBP =
150 mmHg, 2.4 pm/year for SBP = 110 mmHg.
□ SBP = 110 mmHg ■ SBP = 150 mmHg
0 1 2
Duration o f Antihypertensive Therapy (year)
Further analysis controlling for age, treatment assignment and the change of
fasting glucose during the trial did not attenuate the associations presented in Table
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2. Gender, smoking history, baseline or on-trial change of BMI, baseline or on-trial
change of lipids, and the use of anti-inflammatory medications were not included in
the final model, because they were not significantly associated with CIMT
progression and adjustment for these variables did not alter the parameter estimates
for the 3-way interaction terms. In a separate model, DBP was not significantly
associated with CIMT progression (p-value for DBP*years since randomization =
0.73) (data not shown).
7.4 Discussion
In summary, our post hoc cohort analyses indicate that elevated SBP
accelerates atherosclerosis progression in patients with T2DM and that
antihypertensive therapy reduces the harmful impact of elevated blood pressure on
subclinical atherosclerosis progression. These associations were not attenuated after
adjustment for age, randomized treatment assignment and the change in fasting
glucose during the trial. Major randomized, placebo-controlled trials have
demonstrated the substantial benefit of antihypertensive therapy in reduction of
cardiovascular events in persons with T2DM (145). Our results complement these
data by demonstrating the effects of hypertension and antihypertensive therapy on
the arterial wall changes that precede clinical events by several decades. This high-
risk population is known to have thicker cross-sectional CIMT (91-93; 144) and
accelerated CIMT progression relative to those without T2DM (90; 94). It remains
unclear whether the treatment effects are demonstrable using CIMT progression as a
cost-effective endpoint in the diabetic population.
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Our study is the first to demonstrate the harmful effect of elevated SBP on
CIMT progression among persons with T2DM. In nondiabetic populations, elevated
SBP but not elevated DBP, was associated with increased CIMT cross-
sectionally(65; 125; 126) and longitudinally(127). Several mechanisms by which
hypertension may accelerate atherogenesis, including increased oxidative stress,
leukocyte adhesion and smooth muscle cell growth have been proposed (55; 56).
The findings of our study also indicate that antihypertensive therapy reduces
the deleterious effect of SBP on CIMT progression in a duration-dependent manner
among persons with T2DM. A randomized trial in 98 Japanese patients with T2DM
reported treatment with enalapril lOmg/d (N = 48) reduced CIMT progression over 2
years relative to the placebo group (N = 50) (10 vs. 20 pm/year, p < 0.05)(131). This
is the only other trial to date to evaluate the efficacy of antihypertensive agents on
CIMT progression in persons with T2DM. Our study not only supports these
findings in a larger, primarily Hispanic American sample, but also for the first time
demonstrates the duration-dependent beneficial effect of antihypertensive therapy on
CIMT progression.
Furthermore, we extend these results to show that the beneficial effect of
antihypertensive therapy on subclinical atherosclerosis is not fully explained by
blood pressure normalization. In our regression model, the direct effect of SBP
normalization is evidenced in the significant follow-up time* SBP term. The
significant inverse regression coefficient of the 3-way interaction term (follow-up
time*SBP*duration of antihypertensive therapy) illustrates that even at matched
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SBP, individuals who received antihypertensive therapy had a lower CIMT
progression rate than those who did not. The additional effects may be related to
direct arterial wall protective effects that are not mediated through blood pressure
lowering but by anti-inflammatory mechanisms, such as enhancing nitric oxide and
prostacyclin, decreasing oxidative stress and proliferation of vascular smooth muscle
cells (56). The Study to Evaluate Carotid Ultrasound changes in patients treated
with Ramipril and vitamin E (SECURE) supported these proposed mechanisms by
investigating the effect of Ramipril on CIMT progression in a primarily nondiabetic
population (130). Our results extend previous findings to adults with insulin-
requiring T2DM.
Limitations of our analyses include the fact that the data on the use of
antihypertensive agents were recorded but compliance was not. Resulting
measurement error of the duration of antihypertensive therapy may have led to
biased results. However, since the measurement error is likely to have been random
across different CIMT levels, the biased associations would be towards the null, so
that the true association between duration of antihypertensive therapy and
progression of CIMT could have been stronger than the significant associations we
found. Secondly, our study did not have sufficient variability in different classes of
antihypertensive agents to address whether certain classes may be more or less
effective than others in reducing atherosclerosis progression. Thirdly, use of
antihypertensive agents was not randomized in our study. However, a standard trial
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protocol for administering medication use removed many selection biases that may
otherwise have been evident.
7.5 Conclusion
In summary, our results indicate that elevated SBP accelerates atherosclerosis
progression in patients with T2DM and antihypertensive therapy reduces this
harmful impact of elevated blood pressure in a duration-dependent manner. The
effect of antihypertensive treatment was partially, but not completely explained by
differences in blood pressure, suggesting an additional and perhaps direct effect of
treatment on the arterial wall. Our study is one of the first to demonstrate that the
antiatherogenic effect of antihypertensive agents, including blood pressure
normalization and possible direct vascular wall protection, can be detected by CIMT
progression obtained with B-mode ultrasound in patients with T2DM. Given the
epidemic of T2DM and the consequent health care burden in the U.S., ultrasound
measurement of CIMT is a suitable end point for cost-effective intervention trials in
patients with T2DM.
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Data Analysis and Planned Publications
Chapter 8: Lipid Modifying Therapy Reduces Progression of Subclinical
Atherosclerosis among Patients with Type 2 Diabetes Mellitus
Summary of Key Points
• The beneficial effects of lipid modifying therapy on subclinical
atherosclerosis are conclusive in the non-diabetic population but less
well-studied among the diabetic population.
• To investigate the association between lipid modifying therapy and
progression of CIMT among persons with T2DM, we conducted a post
hoc cohort analysis of longitudinal data from the Troglitazone
Atherosclerosis Regression Trial
• Our data indicate that lipid modifying therapy with statins is associated
with a reduction in the progression of subclinical atherosclerosis in
diabetic patients in a duration-dependent manner. This beneficial effect
is stronger with high LDL-C levels. Our data support aggressive and
sustain lipid modifying therapy in patients with T2DM.
8.1 Introduction
Diabetes is a major risk factor for cardiovascular disease (CVD). Prospective
studies indicate that diabetic dyslipidemia is an independent contributor to CVD and
thereby a defined target for intervention in persons with type 2 diabetes mellitus
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(T2DM) (9). As an underlying etiology of CVD, the development of
atheroscleorosis is acclerated among diabetic patients (37). The beneficial effects
of lipid modifying therapy (LMT) on subclinical atherosclerosis are conclusive in the
non-diabetic population but less well-studied among the diabetic population (134-
140; 147).
Carotid artery intima-media thickness (CIMT) acquired with high-resolution
B-mode ultrasound is a well-established noninvasive measure of subclinical
atherosclerosis (69). Few studies have examined the association of LMT and
atherosclerosis progression among diabetic patients although this particular
population is known to have thicker cross-sectional CIMT (91-93; 144) and
accelerated CIMT progression relative to those without diabetes (90; 94).
Although several randomized trials have demonstrated a significant
beneficial effect of LMT on CVD risk in both primary and secondary prevention
among patients with T2DM (113; 118-120), whether LMT in general and statins in
particular have antiatherogenic effects on CIMT progression is not clear in the
diabetic population. In particular, if the antiatherogenic effects of LMT is detectable
using CIMT progression in the diabetic population, the interventional trials in this
population can be conducted in an efficient and cost-effective manner as smaller
sample sizes and shorter duration of follow-up are needed relative to studies using
CVD morbidity and mortality as endpoints. To investigate the association between
LMT and progression of CIMT among persons with T2DM, we conducted a post hoc
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cohort analysis of longitudinal data from the Troglitazone Atherosclerosis
Regression Trial (TART).
8.2 Methods
TART was a single-center, randomized, double-blind, placebo-controlled,
carotid arterial wall imaging trial designed to determine whether improvement in
glycemic control and reduction in insulin requirements with a PPARy agonist reduces
the progression of subclinical atherosclerosis in subjects with insulin-requiring
T2DM. The study design details have been described (95).
Briefly, eligible subjects were men and women >30 years old with age at
diagnosis of T2DM > 30 years old, fasting serum glucose between 140-350 mg/dl on
> 2 occasions and were receiving or scheduled to receive < 150 U insulin per day
without other anti-diabetic medications. After a 8-12 week run-in period during
which diabetes care was intensified with insulin, 101 male and 198 female subjects
with T2DM were randomized to either placebo (N=150) or troglitazone 400 mg daily
(N=149) in double-blind fashion for a two-year treatment period. After
randomization, clinic visits occurred at two and four weeks, then monthly until six
months, then bimonthly until 24 months. The primary trial end point was the rate of
change in CIMT. The Institutional Review Board of the University of Southern
California approved the study and all subjects provided written informed consent.
Measurement of CIMT was conducted at baseline and every six months
during the trial by a single ultrasonographer using high resolution B-mode carotid
artery ultrasound (ATL ultrasound imager equipped with a linear array 7.5 MHz
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probe). Automated edge detection to locate the lumen-intima and media-adventitia
echo boundaries at sub-pixel resolution was used to measure CIMT (Prowin, patent
pending) (104). The CIMT measurement consisted of an average of 80-100
independent measurements made along a 1 cm distance of the distal right common
carotid artery far wall. Using this methodology, the coefficient of variation for
average CIMT (replicate scans obtained approximately 2-weeks apart) was less than
1% in this study.
During the trial, lipid modifying medications, antihypertensive agents and
insulin were provided as part of the trial protocol. All other non-study medications
were prescribed by the primary physicians of the patients. To maintain LDL-C
between 130-160 mg/dl, subjects with fasting plasma LDL-C >160 mg/dl were
given a 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitor
(primarily atorvastatin). The target range for sitting blood pressure was 110-140/80-
90 mmHg for all subjects. Subjects with blood pressure greater than 140/90 mmHg
and no greater than 170/110 mmHg (subjects with blood pressure greater than
170/110 mmHg at an initial screening visit were not eligible for the study) were
given primarily an angiotensin-converting enzyme inhibitor, followed by a long-
acting calcium channel blocker and then an alpha blocker when necessary.
Glucocorticoids (except inhaled and topical), oral anti-diabetic medications and
nicotinic acid were prohibited during the trial. At each follow-up visit, insulin doses
were adjusted to maintain fasting and Zi hour pre-meal blood glucose levels between
100-150 mg/dl. Throughout the trial, a standard medication record form was used to
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record the use of all prescription medications, including lipid modifying medications
(dose, frequency and dates of use). Subjects were requested to bring all medications
to each clinic visit.
Subjects were interviewed with standardized structured questionnaires at
baseline to ascertain their age, gender, ethnicity, years of education, age at diagnosis
of T2DM, age at initiation of insulin therapy and current daily insulin dose. Height,
weight, waist and hip circumference were measured and body mass index (BMI) and
waist-hip-ratio (WHR) were derived from these data. BMI was calculated as weight
(kilograms)/height (meters)2.
Blood samples were obtained at each clinic visit after an 8-12 hour overnight
fast. Serum glucose levels were measured by glucose oxidase (Beckman Glucose
analyzer n, Beckman Instruments, Brea CA, USA). Hemoglobin A le levels were
measured by high-pressure liquid chromatography (BioRad, Hercules CA, USA).
Plasma total cholesterol, total triglyceride, and high-density lipoprotein cholesterol
(HDL-C) levels were determined by enzymatic assays and standardized to the CDC
using the Lipid Research Clinics protocol. HDL-C levels were measured after
apolipoprotein B-containing lipoproteins were precipitated in whole plasma with
heparin manganese chloride. LDL-C levels were estimated using the Friedewald
equation (146). At each visit, blood pressure was computed as the average of two
measures after subjects sat with legs dangling for at least 5 minutes.
Data are presented as mean (SD) unless specified otherwise. The annual rate
of change in CIMT was the primary endpoint. A total of 276 subjects (142 in the
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troglitazone and 134 in the placebo group) had at least one CIMT measurement after
randomization and were included in the analysis. Multivariable mixed-effect models
were used to evaluate the rate of change in CIMT in relation to measures of LDL-C
and duration of LMT. During the trial, CIMT at a given follow-up time was
regressed on the follow-up time (years since randomization). The regression
coefficient associated with follow-up time represents the average rate of change in
CIMT. Duration of LMT was included as an independent variable and measures of
LDL-C at each time point were included as time-varying independent variables. The
association between LMT and CIMT progression were evaluated by 2-way
interaction terms (follow-up time*duration of LMT). Whether LDL-C levels
modified this association was tested by a 3-way interaction term (follow-up
time*LDL-C*duration of LMT). All main effects and lower-order interaction terms
were included in these models. Age, randomized treatment assignment, and LDL-C
levels at randomization were included as covariates to control for potential
confounding effects. Age and LDL-C levels were centered by their means. Gender,
smoking history, baseline or on-trial change of BMI, and other lipids measures were
not included in the final model, because they were not significantly associated with
IMT progression and adjustment for these variables did not alter the parameter
estimates for the 3-way interaction terms. All statistical testing was performed at a
5% level of significance using Statistical Analysis System version 8.0 (SAS Institute,
Cary, NC) and SAS PROC MIXED for the mixed effects models.
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8.3 Results
Overall, 299 subjects were randomized to TART. A total of 276 subjects (90
men and 186 women) had at least one follow-up CIMT and were included in the
present analysis. The mean age at randomization was 52.5 years (range 30 to 70
years). Eighty-nine percent of the subjects were of Hispanic origin and 67% were
female. The mean age at diagnosis of T2DM was 42.7 (8.6) years and the mean
duration of diabetes was 9.8 (6.3) years. Sixty-six percent of the sample had never
smoked regularly (n = 182). Among 94 (34%) subjects who had smoked daily for at
least 6 months, 63 (23%) were former smokers and 31 (11%) were current smokers.
Table 8.1 summarizes the baseline characteristics of the analysis sample at the
screening visit and the randomization visit.
Table 8.1 Characteristics o f TART Subjects at the Initial Screening Visit and at Randomization
Screening Visit (n=276) Randomization (n=276)
CIMT (pm) 8 1 4 .4 (1 5 4 .2 )1 814.1 (154.4)
Age (years) 52.2 (9.1) 52.5 (9.0)
Hemoglobin A le (%) 9.8 (2.3) 7.5 (1.1)
Fasting glucose (mmol/L) 11.6(4.2) 7.9 (2.1)
Daily insulin dose (U) 53.0 (29.5) 60.2 (33.3)
Systolic blood pressure (mmHg) 135.7 (20.0) 130.9(19.0)
Diastolic blood pressure (mmHg) 79.7(10.2) 76.2 (9.4)
Body mass index (kg/m2) 31.6 (6.0) 32.0 (6.0)
Weight (kg) 81.3 (17.8) 82.3 (17.6)
Total cholesterol (mmol/ L) 5.2(1.1) 4.4 (0.9)
Triglycerides (mmol/ L) 2.0 (1.2) 1.6 (1.0)
HDL-cholesterol (mmol/ L) 1.3 (0.3) 1.2 (0.3)
LDL- cholesterol (mmol/ L) 2.9 (0.9) 2.6 (0.8)
Taking antihypertensive agents 120 (43) 2 151 (55)
Taking lipid lowering medications 24(9) 16(6)
Receiving insulin therapy 228 (83) 276(100)
Taking anti-inflammatory medications 52(19) 64 (23)
1. Mean (SD)
2. N (%)
On average, these insulin-dependent type 2 diabetic subjects were obese and
normotensive. Almost half of the samples were using antihypertensive agents,
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and 9% were receiving LMT at the screening visit. Elevated fasting plasma glucose,
HbAlc and lipid profiles were improved during the 8-12 week run-in phase. At the
end of run-in, (i.e, at randomization to treatment), 55%, 6% and 23% of the sample
were taking antihypertensive agents, lipid lowering medications and anti
inflammatory medications, respectively.
During the 2-year trial, 80 (29%) subjects received LMT. Among them, 72
(90%) subjects used statins alone, 2 (3%) used a fibric acid compound alone due to
high triglycerides greater than 200 mg/dL and 6 (7%) used a statin and a fibric acid
compound combined. These proportions did not differ between troglitazone and
placebo groups (p = 0.27). The duration of LMT ranged from 0.03 to 2.1 years with
a median of 0.7 years. Subjects who received LMT had similar baseline CIMT as
those who did not (819.4 (144.6) pm vs. 812.4 (158.3) pm, p = 0.73). Among the 80
subjects receiving LMT during the trial, the mean LDL-C was 126.1(41.2) mg/dL at
the initial screening visit when 24 subjects were already receiving LMT. The mean
LDL-C level of these 80 subjects during LMT was significantly reduced by
23.7(42.9) mg/dL (p < 0.0001).
In a single risk factor mixed effects model, duration of LMT was not
significantly associated with CIMT progression rate (p-value for follow-up
time*duration of LMT = 0.46). A multivariate mixed effects model was fitted to
evaluate the annual rate of change in CIMT in relation to LDL-C levels and duration
of LMT, after controlling for LDL-C levels at randomization (Table 8.2). In Table
8.2, the regression coefficient associated with follow-up time represents the average
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annual rate of change in CIMT. A significant interaction between duration of LMT
and LDL-C on the CIMT progression rate (p = 0.03) illustrates that LMT reduces the
CIMT progression in a duration-dependent manner, and the association is stronger in
subjects with higher initial LDL-C. Concrete examples of the application of these
coefficients are as follows: for LDL-C = 160 mg/dl at randomization and a reduction
of lOmg/dl during the trial, the CIMT rates were 6.33, 4.48 and 2.62 pm/year by 0, 6
and 12 months of LMT, respectively. For LDL-C = 190 mg/dl at randomization and
a reduction of lOmg/dl during the trial, the CIMT rate was 7.07, 4.54 and 2.02
pm/year by 0, 6 and 12 months of LMT, respectively (Figure 8.1). Adjustment for
on-trial change of HDL-C and triglycerides did not alter the parameter estimates for
the 3-way interaction terms (data not shown).
Table 8.2 Parameter Estimates Related to the CIMT Progression Rate from the Multivariable Mixed Effect Model
for LDL-C levels, Duration of LMT and CIMT Progression_________________________________________________
Effects
CIMT progression rate (pm/year)
attributable to each effect SE P-value
Follow-up time (pm/year) 5.14 1 1.48 0.0006
Follow-up time *LDL-C at randomization
(pm/year per mg/dL) 0.005 2 0.05 0.92
Follow-up time *LDL-C at follow-up
(pm/year per mg/dL) 0.02 3 0.02 0.26
Follow-up time *Duration o f LMT
(pm/year per year o f therapy) -1.69 4 2.87 0.56
Follow-up time *LDL-C*Duration o f LMT
(pm/year per mg/dL per year o f therapy) -0.04 5 0.02 0.023
1. average CIMT progression rate for LDL-C at randomization = 97 mg/dL and on-trial LDL-C = 105 mg/dL,
without LMT
2. average increase in CIMT progression rate for each lmg/dL increase in LDL-C at randomization centered at
97 mg/dL
3. average increase in CIMT progression rate for each lmg/dL increase in LDL-C during follow-up centered
at 105 mg/dL
4. average decrease in CIMT progression rate for one year increase in duration o f LMT
5. the effect o f LMT on CIMT progression is stronger with higher LDL-C
Further analysis controlling for age, gender and randomized treatment
assignment (troglitazone vs. placebo) did not alter the associations presented in
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Table 8.2. Gender, smoking history, baseline or on-trial change of BMI, and the use
of anti-inflammatory medications were not included in the final model, because they
were not significantly associated with CIMT progression and adjustment for these
variables did not alter the parameter estimates for the 3-way interaction terms (data
not shown).
Figure 8.1 Predicted annual CIMT rates (using Table 8.2 parameter estimates) in relation to LDL-C levels and the
duration o f LMT from the multivariate mixed-effects model (assuming a lOmg/dL reduction o f LDL-C during
the trial)
□ LDL-C=160 at randomization
■ LDL-C= 190 at randomization
0 0.5 1
Duration of Lipid Modifying Therapy (year)
8.4 Discussion
In summary, our post hoc cohort analyses using TART data indicate that
LMT is associated with a reduction in CIMT progression in a duration-dependent
manner among patients with T2DM. The magnitude of change in CIMT progression
is greater among subjects with higher baseline LDL-C levels. The beneficial effect
of LMT is not accounted for by on-trial changes in HDL-C and triglycerides.
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Over the past decade, a large body of published work has demonstrated a
lowering of CIMT progression rates by LMT, despite the variation in study
populations and CIMT progression rates across trials (134-140; 147). Due to the
small number of diabetic subjects recruited, no trial had sufficient power to perform
a subgroup analysis on any diabetic subgroups. Our findings expand previous
research to a purely type 2 diabetes population that is predominantly Hispanic.
Several clinical practice guidelines have suggested aggressive LMT in type 2
diabetes (18; 45). Our study is the first to indicate that LMT with statins is
associated with a reduction in the progression of subclinical atherosclerosis in
patients with T2DM in a duration-dependent manner, thereby supports aggressive
and sustain LMT in diabetic patients. Several potential biological mechanisms of
statins on atherosclerosis regression have been proposed. Statins antagonize HMG-
CoA reductase, which leads to a reduction in the synthesis and secretion of
lipoprotein by the liver, an upregulation of LDL receptors on hepatocytes, and
increasing clearance of circulating apolipoprotin E- and B-containing lipoproteins.
Consequently, increasing LDL clearance and decreasing VLDL secretion reverse
atherosclerosis progression (148). The potent lipid lowering effects of statins on
CVD or recurrent CVD risk has been evidenced among diabetics in several clinical
trials (113; 118-120). However, a benefit of LMT on atherosclerosis progression has
not been demonstrated in a diabetic population.
Our study also demonstrated that the beneficial effect of LMT on
atherosclerosis progression is not fully explained by lipid lowering. In our
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regression model, the significant inverse regression coefficient of the 3-way
interaction term (follow-up time*LDL-C*duration of LMT) illustrated that even at
matched LDL-C, individuals who received LMT had a lower CIMT progression rate
than those who did not. In vitro and in vivo studies suggest statins also have anti
inflammatory effects mediated by increased expression of endothelial nitric oxide
aynthase and reduced expression of proinflammatory factors (149). Moreover, a
prospective study on 21 patients with T2DM observed significant reduction of
circulating immune complexes containing modified lipoproteins during treatment
with simavastain. At the end of 3 to 6 month washout period, modified lipoproteins
returned to baseline levels, which suggested the role of simvastatin as an
immunomodulator in type 2 diabetes (150). Therefore, the antiatherogenic effect of
LMT in persons with T2DM may be additionally related to anti-inflammatory and
immunomodulatory effects of statins. These findings are also supported by the
recognition that atherosclerosis is a chronic inflammatory process rather than a
degenerative process that inevitably progresses with age and results simply from the
accumulation of lipids (48).
Limitations of our analyses include the fact that the duration of LMT was
recorded but compliance was not. Resulting misclassification error related to the
duration of LMT may have led to biased results. However, since misclassification of
receiving LMT should not be related to the CIMT level, the biased associations
would be towards the null, and the true association between LMT and progression of
CIMT would have been stronger than the significant associations we found.
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Secondly, our study did not have sufficient variability in different classes of lipid
modifying medications to address whether certain classes may be more or less
effective than others in reducing atherosclerosis progression. Thirdly, use of lipid
modifying medications was not randomized in our study. However, a standard trial
protocol for administering these medications removed many selection biases that
may otherwise have been evident.
8.5 Conclusion
In conclusion, our results indicate LMT is associated with a reduction in the
progression of subclinical atherosclerosis in a duration-dependent manner among
type 2 diabetes. The beneficial effect of LMT is partially related to a reduction in
LDL-C levels and not explained by the changes in other lipid levels. Our study is
one of the first clinical data to suggest that statins reduce subclinical atherosclerosis
progression by mechanisms beyond mere lipid lowering in type 2 diabetes, thereby
supports current guidelines for aggressive LMT in patients with T2DM. Given the
epidemic of T2DM and the consequent health care burden in the U.S. (46),
randomized trials in type 2 diabetes should be designed to confirm these data and
identify the different antiatherogenic effects of statins in this high-risk population.
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Grant Proposal
Chapter 9: Pooled Analysis of Metabolic Factors and Atherosclerosis
Progression
Summary of Key Points
• Current NCEP ATP III definition of metabolic syndrome has recently
been questioned in terms of the completeness of the diagnostic criteria,
equal weight of each component, and arbitrary dichotomy of each
component.
• Accumulating evidence links obesity and insulin resistance to
inflammation, and thereby supports inflammatory mechanisms as
pathological pathways of both insulin resistance and atherosclerosis
progression. In addition, obesity and obesity-induced insulin resistance
may lead to enhanced synthesis of hemosatic factors, which predispose
an individual to atherothrombosis.
• It remains to be determined whether markers of inflammation and
hemostasis will provide additional predictive information on
atherosclerosis progression beyond traditional risk factors.
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9.1 Specific Aims
This proposal is submitted in response to PA-05-094 “Secondary Analyses in
Obesity, Diabetes, Digestive and Kidney Disease”. We propose to pool data from
five completed clinical trials to evaluate the contributions of metabolic abnormalities
to progression of subclinical atherosclerosis.
Obesity and concurrent metabolic abnormalities, including insulin resistance,
hypertension, and dyslipidemia, in an individual confer an increased risk of
cardiovascular morbidity and mortality. These traditional cardiovascular risk factors
were proposed as components of the metabolic syndrome (MetS) by the Third
Report of the National Cholesterol Education Program’s Adult Treatment Panel III
(NCEP ATP HI) (45). However, the current NCEP ATP III definition has
recently been questioned in terms of the completeness of the diagnostic criteria,
equal weight of each component, and arbitrary dichotomy of each component
(151). The joint statement from the American Diabetes Association and the
European Association for the Study of Diabetes called for “an evidence-based
analysis assessing the rationale and value of adding other CVD risk factors to
the definition” of the MetS (151).
The underlying etiologies of the MetS and its relation to progression of
atherosclerosis are not completely understood. Accumulating evidence links
obesity and insulin resistance to inflammation, and thereby supports
inflammatory mechanisms as pathological pathways of both insulin resistance
and atherosclerosis progression. In addition, obesity and obesity-induced insulin
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resistance may lead to enhanced synthesis of hemosatic factors, which predispose an
individual to atherothrombosis. It remains to be determined whether markers of
inflammation and hemostasis will provide additional predictive information on
atherosclerosis progression beyond traditional MetS risk factors.
We therefore propose to use data from five clinical trials — the Estrogen in
the Prevention of Atherosclerosis Trial (EPAT), the Women’s Estrogen-Progestin
Lipid-Lowering Hormone Atherosclerosis Regression Trial (WELL-HART), the
Vitamin E Atherosclerosis Prevention Study (VEAPS), the Troglitazone
Atherosclerosis Regression Trial (TART) and the B-Vitamin Atherosclerosis
Intervention Trail (BVAIT), in which the annual rate of change in carotid artery
intima-media thickness (CIMT) was the primary measure of subclinical
atherosclerosis progression, to achieve two specific aims:
1. To evaluate whether inflammatory and hemostatic factors will provide
additional predictive information on subclinical atherosclerosis progression
beyond the five metabolic variables used in the current definition of MetS.
i. By principal component analysis, we will identify core factors that are
statistically independent of one another using two sets of original
variables: (1) the standard 5 metabolic variables used in the NCEP ATP
III definition of MetS, and (2) the standard 5 metabolic variables plus
inflammatory and hemostatic variables.
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ii. To investigate the associations between the two sets of principal
components and progression of CIMT in the pooled sample, and
determine the best fitting model.
2. To examine whether the associations between the principal components and
CIMT progression vary by CVD and diabetes status.
We hypothesize that (1) addition of inflammatory and hemostatic variables
will alter the principal components identified by the current definition of the MetS,
and therefore contribute additional information on the pathogenesis of
atherosclerosis progression. (2) The magnitude of the associations between principal
components of the MetS and CIMT progression will be greater in persons with CVD,
and with type 2 diabetes.
9.2 Background and Significance
9.2.1 NCEP ATP III Definition of the MetS
In 2001, the Third Report of the National Cholesterol Education Program’s
Adult Treatment Panel (NCEP ATP III) guidelines proposed a clinical definition for
identifying individuals with the MetS (45). The five components delineated for the
MetS include abdominal obesity (waist circumference >35 inches in women and 40
inches in men), hypertriglyceridemia (> 150 mg/dL), low high-density lipoprotein
cholesterol (HDL-C) (< 40mg/dL in men and < 50mg/dL in women), elevated blood
pressure (> 130/85 mmHg) and hyperglycemia (fasting plasma glucose >110
mg/dL). A clinical diagnosis of the MetS can be made by satisfying three of the five
criteria (45). Using this definition, the age-adjusted prevalence of the MetS was 237
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per 1,000 in US adults over 20 years of age (152) and 435 per 1,000 among adults
over 50 years of age (153).
Although NCEP ATP III also identified six components of the MetS as
abdominal obesity, atherogenic dyslipidemia, raised blood pressure, insulin
resistance with or without glucose intolerance, and prothrombotic and
proinflammatory states, the current clinical definition of MetS does not incorporate
measures of inflammatory and hemostatic activity because of the challenge of
routine assessment. As chronic subclinical inflammation is known to be part of the
MetS (154), and hemostatic factors have been shown to predict cardiovascular events
(155; 156) and to interact with other components of the MetS (157; 158), the
integration of inflammatory and hemostatic markers with other traditional markers of
the MetS may provide important insights to mechanistic hypotheses, risk prediction
and preventive interventions.
9,2.2 Pathogenesis of the MetS
The pathogenesis of the MetS is complex and not completely understood. As
an underlying risk factor for CVD, it has been suggested that obesity, specifically
visceral obesity plays a key role in the development of the MetS (159). In persons
with visceral obesity, excessive circulating free fatty acids (FFA), derived from
visceral adipocytes, worsen the insulin resistance in the liver and skeletal muscle.
Visceral adipocytes also release cytokines (interleukin-6 (IL-6) and tumor necrosis
factor a (TNF-a)) that promote inflammation, dyslipidemia, and hypertension, and
impair thrombolysis (160). On the other hand, insulin resistance has been suggested
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as the underlying mechanism in the development of the MetS. As quantification of
insulin action in vivo is not always strongly related to the presence of the MetS
(161), it raises the possibility of other unknown mechanisms beyond insulin
resistance.
Of note, accumulating evidence links inflammation to obesity and insulin
resistance, and thereby supports inflammatory mechanisms as pathological pathways
of both insulin resistance and atherosclerosis (162). In obese mice, TNF-a was
shown to be hyperexpressed and to mediate insulin resistance (163). In obese
humans, adipose tissue has been shown to cause elevated plasma concentrations of
TNF-a and IL-6. These inflammatory mediators induce insulin resistance, which
promotes further inflammation, suppresses the anti-inflammatory effect of insulin,
and reduces the beneficial effect of insulin in thrombin formation and fibrinolysis in
atherothrombosis (162). In particular, decreased insulin sensitivity may lead to
enhanced synthesis of C-reactive protein, intercellular adhesion molecule-1, tissue
factor and plasminogen activator inhibitor-1 (PAI-1) during the acute-phase
response.
Therefore, it is critical to incorporate inflammatory and hemostatic factors in
the definition of MetS to explore whether they contribute additional information on
the pathogenesis of the MetS and atherosclerosis progression.
9.2.3 MetS and CVD
Among adults age 50 years and older, the prevalence of CVD was 8.7% in
those without either the MetS or diabetes, 7.5% in those with diabetes but without
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the MetS, 13.9% in those with the MetS but without diabetes, and 19.2% among
those with both the MetS and diabetes (153). Persons with the MetS have a 2- to 3-
fold greater risk of developing CVD and almost 6- fold greater risk of cardiovascular
death than those without (153; 163-172).
9.2.4 MetS and CIMT
Subjects with the MetS are known to have thicker CIMT in cross-sectional
analyses (173-175) and accelerated progression of carotid atherosclerosis (assessed
by carotid stenosis) relative to those without the MetS (176). The data assessing the
association between the MetS and CIMT progression are scant. It is known that
obesity (177), dislipidemia (177), hypertension (90; 127) and hyperglycemia (90; 94)
are each independently associated with accelerated CIMT progression. In addition,
elevated concentrations of high sensitivity C-reactive protein (hs-CRP) and soluble
intracellular adhesion molecules-1 (sICAM-1) were each independently associated
with CIMT progression in Japanese population while similar data are scant in the US
(178; 179). Increased fibrinogen has been shown to be associated with thicker CIMT
in a cross-sectional analysis in the Atherosclerosis Risk in Communities study (180;
181).
9.2.5 Inflammatory and Hemostatic Variables and the MetS
So far, two cross-sectional studies have utilized NCEP ATP in definition to
examine the associations between the MetS and inflammatory and hemostatic
markers (182; 183). In 8570 men and women aged 20 years or older from the Third
National Health and Nutrition Examination Survey (1988-1994), higher prevalence
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of elevated hsCRP and fibrinogen concentrations and white blood cell counts were
observed among subjects with the MetS compared to those without the syndrome
(182). In 2722 nondiabetic older men aged 60-79 years and free of CVD, men with
the MetS had a significantly higher prevalence of elevated tissue plasminogen
activator (tPA) antigen, factor VII antigen concentrations and white blood cell counts
relative to those without the syndrome (183). Our preliminary data from the EPAT
study are consistent with these studies (see Preliminary Studies section 3.3).
Increased hs-CRP levels are associated with higher body mass index (BMI)
(184-188), waist circumference (154), triglyceride levels (184; 187; 188), blood
pressure (186; 188), fasting glucose (184; 187), fasting insulin (154), and reduced
HDL-C levels (184; 187; 188) and insulin sensitivity (154). Increase in PAI-1
antigen, tPA antigen, fibrinogen and factor VII antigen, are commonly accompanied
by obesity, dyslipedemia and hypertension (157; 189; 190), hyperglycemia and
hyperinsulinemia in nondiabetic (154; 191-193) and diabetic populations (194; 195).
In particular, among more than 1,000 nondiabetic subjects from the Insulin
Resistance Atherosclerosis Study, elevated CRP and fibrinogen concentrations have
been associated with increased BMI and waist circumference, increased fasting
glucose and insulin concentration, and insulin resistance as determined by the
frequently sampled intravenous glucose tolerance test (154). The interrelations
between known components of current definition of the MetS and inflammatory and
hemostatic markers indicated that these markers might be part of the MetS.
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9.2.6 Inflammatory Variables and CVD
Atherosclerosis is viewed as a chronic inflammatory process involving
monocyte-derived macrophages and specific subtypes of T lymphocytes at every
stage of the disease process (48). Hs-CRP and sICAM-1 are two crucial
inflammatory markers, deriving from the liver and the vascular smooth muscle cells,
respectively. These inflammatory markers are associated with cardiovascular event
risk and have been shown to provide additional predictive information beyond
traditional CVD risk factors (196; 197). As an indicator of cellular responses to
inflammation, elevated leukocyte count has also been suggested as a predictor of
cardiovascular risk (198). A meta-analysis of 19 prospective studies indicated that
individuals whose leukocyte count was in the upper tertile had a 1.4 fold greater risk
of developing CVD relative to those whose leukocyte count was in the lower tertile
(199).
Twenty-two prospective studies in various populations have demonstrated
that elevated hs-CRP levels are associated independently with a greater risk of CVD
(199). Moreover, hs-CRP has been found to be an independent marker of insulin
resistance. Subjects with high CRP levels plus the MetS had 2-fold risk of CVD
relative to those with either high CRP levels or the MetS (200). Therefore, hs-CRP
may be a valuable addition to the definition of the MetS.
9.2.7 Hemostatic Variables and CVD
A prothrombotic state refers to defects in the coagulation and fibrinolytic
systems that predispose an individual to atherothrombosis. PAI-1 antigens, tPA
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antigens, fibrinogen and factor VII antigens are known prothrombotic factors. In a
meta-analysis of 18 prospective studies, elevated fibrinogen significantly increased
the risk of CVD (199). Comparison of individuals with baseline fibrinogen levels in
the upper tertile with those in the lower tertile yielded a summary risk ratio for CHD
of 1.8 (95% Cl, 1.6-2.0). In the Physician’s Health Study, tPA antigen concentration
in the highest quintile was associated with a 2.8-fold risk of myocardial infarction
relative to the lowest quintile, after adjustment for age and smoking (95% Cl, 1.5-
5.4) (201).
9.2.8 Factor Analysis in the MetS
The MetS can present in various ways due to different components that
constitute the syndrome; thus it is critical to identify and quantify these
constellations of highly interrelated metabolic abnormalities. As a useful tool for
data reduction and summarization, factor analysis has been widely used to assess the
correlation between a large set of observed variables by extracting latent factors.
Principal component analysis is one of the most popular approaches to extract
factors. The underlying assumption is that there exist a number of unobserved latent
factors that account for the correlations among observed variables that are highly
correlated. In this approach, the observed variables are transformed into principal
components (latent factors) that account for decreasing proportions of the variance of
the data. Each principal component is associated with an eigenvalue that reflects the
variance explained by the component. A common practice is to extract factors with
an eigenvalue of at least 1.0. Based on the factor loadings, which represents the
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association between each variable and each latent factor, we can name and interpret
each factor. Thus the underlying patterns among many highly intercorrelated risk
variables are revealed.
During the past decade, a few studies have employed factor analyses to
explore the principal components of conventional cardiovascular risk factors,
including insulin sensitivity, hyperglycemia, obesity, dyslipidemia and hypertension,
in diverse populations (202). Overall, insulin sensitivity consistently loads on the
same factor with hyperglycemia, obesity, and in many cases dyslipidemia. Blood
pressure usually loads on a unique, separate factor. Only one study performed factor
analysis on variables including multiple components of the MetS plus inflammatory
and hemostatic factors among 322 nondiabetic elderly individuals aged 65-100 years
(203). Total seven uncorrelated factors were yielded including body mass,
insulin/glucose, lipids, blood pressure, inflammation, vitamin K-dependent proteins
and progoagulant activity. The results of the study suggested that inflammatory and
hemostatic factors might contribute additional information on CVD risk besides
other components of the MetS. Our study proposes to extend their findings by
investigating the associations between the core factors of the MetS and progression
of subclinical atherosclerosis.
9.2.9 Rationale for Specific Aims
Despite the known interrelations of proinflammatory and prothrombotic
states with other metabolic factors and their potential role in predicting increasing
CVD risk, few data are available assessing the roles of a proinflammatory state
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and/or a prothrombotic state on the progression of subclinical atherosclerosis.
Although it is known that obesity (177), dislipidemia (177), hypertension (90; 127)
and hyperglycemia (90; 94) are independently associated with accelerated CIMT
progression, these conventional risk factors do not fully account for the excess
cardiovascular risk observed among subjects with the MetS (204). We will therefore
incorporate all these known risk factors with inflammatory markers (hsCRP,
SICAM-1), and hemostatic factors (fibrinogen, tPA antigen, PAI-1 antigen and
factor VII antigen) in the principal component analysis to further understand the
interrelations of atherogenic factors of the MetS and investigate their integrated
effects on the progression of subclincial atherosclerosis.
9.2.10 Significance of the Proposed Work
Approximately one fourth of adults in the US have the MetS and a large body
of evidence has shown that patients with the MetS have more prevalent CVD or are
at greater risk of developing it (153; 163-172). Current NCEP ATP III definition
has recently been questioned in terms of the completeness of the diagnostic
criteria, equal weight of each component, and arbitrary dichotomy of each
component (151). The joint statement from the American Diabetes Association
and the European Association for the Study of Diabetes called for “an evidence-
based analysis assessing the rationale and value of adding other CVD risk
factors to the definition” of the MetS (151).
The results of this study will provide important insights to further
understand the underlying structures of the MetS by incorporating
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proinflammatory and prothrombotic factors in the principal component
analysis and subsequently assess the predicting value of core factors on the
progression of subclinical atherosclerosis. We may also gain a better
understanding of the impact of core factors of the MetS on subclinical
atherosclerosis progression in various populations, which may improve
cardiovascular risk prediction and further evaluation of antiatherogenic interventions.
9.3 Preliminary Studies
9.3.1 Previous Work using CIMT progression at Atherosclerosis Research Unit
We propose to pool data from five atherosclerosis progression trials
conducted in the past decade at the Atherosclerosis Research Unit (ARU), University
of Southern California, using similar trial and data collection protocols. The wide
variety of populations sampled, including persons with and without CVD, and with
and without diabetes, will allow us to evaluate the impact of MetS on atherosclerosis
progression among these subgroups. The annual rate of change in CIMT was a trial
endpoint of subclinical atherosclerosis progression in each trial.
Since 1982, we have conducted several single-center, randomized, non-
invasive ultrasonographic carotid arterial wall imaging trials at the ARU. The
Cholesterol Lowering Atherosclerosis Study (CLAS), completed in 1987, is the only
study to date that has reported an association between progression of CIMT and
incident CVD using repeated measures of CIMT with a mean follow-up of 8.8 years
post trial. In this study, we found an increase of 30 pm CIMT per year was
associated with a 2.2 fold increased risk for subsequent myocardial infarction or
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coronary death (104). Our findings supported the use of CIMT progression as a cost-
effective marker for cardiovascular event risk. We also used CLAS data to show
CIMT progression was significantly correlated with QCA-measured change in
coronary artery lesions (69).
Subsequently, we completed MARS in 1993 (205), EPAT in 1998 (206),
WELL-HART (207), VEAPS (208) and TART in 2000. By using CIMT progression
as an endpoint to address the efficacy of anti-atherogenic intervention, these trials
demonstrated that CIMT is a reliable, valid and noninvasive measure of sublinical
atheroscleorsis and smaller sample sizes and shorter duration of follow-up are
needed in ultrasonographic carotid arterial wall imaging trial compared to large trials
with cardiovascular morbidity and mortality endpoints. Currently we have BVAIT,
WISH and ELITE in progress.
In 1994, we reported o our methodologic developments in relation to an
automated computerized edge tracking method that reduced measurement variability
due to manual tracing of the lumen-initima and the media-adventitia interfaces
(63).Our automated edge tracking method uses subpixel interpolation to determine
edge boundaries at a resolution greater than monitor liner resolution. The resulting
CIMT measurement consists of an average of 80-100 independent measurements
made along a 1cm distance in the far wall of the distal common carotid artery. Using
this methodology, the coefficient of variation for average CIMT (replicates scans on
the same day) is 2.5% within operators. Variability of CIMT measurement is
reduced 2-4 times compared to that obtained with manual tracking methods (74).
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9.3.2 Atherosclerosis Regression Trials to be Included in the Proposed Work
All five trials were randomized, single-center, double-blind, placebo-
controlled, clinical trials designed to test the effect of hormone therapy (EPAT,
WELL-HART), vitamin E (VEAPS), troglitazone (TART) or B-vitamins (BVAIT)
on the progression of subclinical atherosclerosis using annual rate of change in
CIMT. The BVAIT trial is in progress and will be completed by July 2006.
The designs of the trials (except BVAIT) were described previously (95; 206-
208). We summarize the key design characteristics of each trial in Table 9.1. All
baseline characteristics presented in Table 1 and the results reported in the following
paragraphs are based on evaluable subjects in each trial who had at least one
measurement of CIMT during the trial follow-up (thereby contributing to assessment
of CIMT progression). Overall, 564 of the total of 1539 evaluable subjects met
criteria for the diagnosis of MetS at baseline, according to NCEP ATP III guidelines.
9.3.2.1 Estrogen in the Prevention of Atherosclerosis Trial (EPAT)
Briefly, EPAT was designed to evaluate the effect of unopposed micronized 17(3-
estradiol on the progression of subclinical atherosclerosis in 222 healthy
postmenopausal women without preexisting CVD. Among 199 subjects with CIMT
progression data, a significantly lower CIMT progression rate was observed among
women taking unopposed estradiol compared to those taking placebo (-1.7 vs.
3.6pm/year, p = 0.046). When stratified by lipid lowering therapy during the trial, a
significant difference between the estradiol and placebo groups remained among
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Table 9.1 Design and Baseline Characteristics o f Five Randomized Atherosclerosis Regression Trials
EPAT WELL-HART VEAPS TART BVAIT
Intervention unopposed oral 17(3-estrodiol
1 mg/day
Micronized 17P-estrodiol
alone or 17P-estrodiol plus
medroxyprogesterone
acetate
DL-a-tocopherol 400
IU/day
troglitazone 400mg/day folic acid 5 mg, B 1 2
0.4 mg and B6 50 mg
Key inclusion
criteria
> 45 years old;
Postmenopausal;
No existing CVD;
LDL-C > 130mg/dL
< 75 years old;
Postmenopausal;
LDL-C 100-250 mg/dL;
TG < 400 mg/dL;
> 1 coronary artery lesion
> 40 years old;
No existing CVD;
LDL-C > 130mg/dL
30 - 70 years old;
Age at diabetes
diagnosis > 30 yrs; FG
<350 mg/dL;
receiving insulin<150
U/day without other
antidiabetic medication
> 40 years old;
postmenopausal (for
women);
No existing diabetes,
CVD or cancer;
Study period Apr. 1994-N o v . 1998 Jun. 1995-O ct. 2000 Jul. 1996-S e p . 2000 Jan. 1997-A p r. 2000 Nov. 2000 -onging
Follow-up (years) 2 3 3 2 2.5 4.5
Randomized 222 226 353 299 506
Evaluable subjects
(Tx / PI) 2
1 9 9 (1 0 2 /9 7 ) 226 (76/74/76)1 3 3 2 (1 6 2 / 170) 276 (1 4 2 / 134) Trial in progress
CIMT in pm, mean
(sd )3
764(132) 845 (219) 754(132) 814(154) 753 (150)
Mean age (range),
yrs
62.2 (46 - 80) 63.5 (4 8 - 7 5 ) 56.2 (40 - 82) 52.5 (3 0 - 7 0 ) 6 1 .4 (4 0 -8 8 )
Female, n(%) 199(100) 226(100) 172 (52) 186 (67) 197 (39)
Postmenopausal,
n(%of female)
199(100) 226 (100) 120 (70) 136 (73) 197(100)
Diabetes, n(%) 6 (3 ) 115 (51) 0 (0 ) 276(100) 0 (0 )
CVD, n(%) 0 (0 ) 226 (100) 0 (0 ) 21(8) 0(0 )
Ethnicity, n(%)
White 118(59) 69 (31) 248 (75) 13(5) 328 (65)
Black 22(11) 38(17) 31(9) 16(6) 75(15)
Hispanic 40 (20) 100 (44) 31(9) 246 (89) 55 (11)
Asian 18(9) 19(8) 15(5) 0 (0 ) 45 (9)
Other
K D
0 (0 ) 7 (2 ) 1(0) 3 (1 )
o
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Table 9.1 continued
EPAT WELL-HART VEAPS TART BVAIT
Education, n(%)
<12 y 30(15) 91 (40) 7 (2 ) 198 (72) 4 (1 )
>12 y 169 (85) 135 (60) 325 (98) 78 (28) 502 (99)
Smoking History,
n(% )
Current
smoker4
0(0 ) 26 (12) 11(3) 31(11) 18(4)
Former smoker 103 (52) 89 (39) 110(33) 63 (23) 171 (34)
Never smoked 96 (48) 111 (49) 211 (64) 182 (66) 316(62)
Subjects with
MetS5, n(%) 84 (42) 160 (71) 60(18) 181 (66) 79 (16)
1. Estrogen vs. Estrogen-Progestin vs. placebo group
2. Active treatment / Placebo
3. Mean CIMT at baseline
4. Current smoker is defined as having smoked daily for at least 6 months upon study entry
5. The metabolic syndrome is defined using the metabolic variables at randomization, according to NCEP ATP III guidelines. Subjects who had normal blood
pressure but were receiving antihypertensive therapy at baseline are counted as satisfying one o f the five criteria.
o
00
women who did not receive LLT (-1.3 vs. 13.4 pm/year, p = 0.002) but was not
apparent among those who received LLT (-1.9 vs. -1.6 pm/year, p > 0.2) (206).
9.3.2.2. Women’s Estrogen-Progestin Lipid-Lowering Hormone Atherosclerosis
Regression Trial (WELL-HART)
In the WELL-HART study, we enrolled 226 postmenopausal women with at
least one coronary artery lesion evident on the baseline coronary angiogram. This
trial tested whether 17p-estradiol alone (estrogen group) or administered sequentially
with medroxyprogesterone acetate (estrogen-progestin group) slowed the progression
of atherosclerosis relative to usual care (placebo group). Among 226 subjects with
CIMT progression data, 81% of subjects received lipid lowering therapy during the
trial. After a median of 3.3 years of follow-up, there was no significant difference in
coronary artery disease progression measured by sequential quantitative coronary
angiography between the placebo, estrogen and estrogen-progestin groups (207).
Consistently, in the total sample, no significant difference in the CIMT progression
rate was observed among the three groups. However, in nondiabetic subjects, a
significantly lower CIMT progression rate was seen in the estrogen group (-7.8
pm/year, N = 32) than in the placebo group (-3.5 pm/year, N = 33, p = 0.049) (209).
9.3.2.3 Vitamin E Atherosclerosis Prevention Study (VEAPS)
VEAPS was designed to determine the effects of DL-a-tocopherol
supplementation on subclinical atherosclerosis progression in 353 individuals aged
40 years or older, with an LDL-C level of 130 mg/dL or above and no clinical signs
or symptoms of CVD. Among 332 subjects with CIMT progression data, we
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found no significant difference in the CIMT progression rate between the Vitamin E
and placebo group (4.0 vs. 2.3 pm/year, p = 0.08) despite significantly increased
plasma Vitamin E levels, reduced oxidized LDL and reduced LDL oxidative
susceptibility in the Vitamin E group compared to the placebo group (208).
9.3.2.4 Troglitazone Atherosclerosis Regression Trial (TART)
TART was designed to investigate whether improvement in glycemic control
and reduction in insulin requirements with a PPARy agonist (troglitazone) reduces
the progression of sublinical atherosclerosis in 299 insulin-requiring type 2 diabetic
subjects. A total of 276 subjects had at least one measurement of CIMT during the
trial follow-up. The CIMT progression rate was significantly reduced by PPARy
agonist therapy (1.3 pm/year, N = 97) relative to placebo (8.4 pm/year, N = 91, p =
0.03) in the participants with a relatively high baseline CIMT (> 800 pm). No such
effect was observed in participants with a lower baseline CIMT (<800 pm) (the
manuscript of the primary results is currently under review).
9.3.2.5 B-Vitamin Atherosclerosis Intervention Trail (BVAIT)
BVAIT was designed to test whether B vitamin supplementation (folic acid 5
mg, B12 0.4 mg and B6 50 mg) will reduce the progression of early carotid artery
atherosclerosis among subjects with elevated fasting homocysteine levels (> 8.5
umol/L), who were at least 40 years of age, postmenopausal (for women), and
reported no evidence of diabetes, heart disease, stroke, or cancer. A total of 506
subjects were randomized into the trial and it is nearing completion. So far, 177
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subjects have completed the trial, and 273 subjects are scheduled to be closed out in
one year.
9.3.3 Proinflammatory and Prothrombotic Data from EPAT
Of these five trials, EPAT is the only trial that currently has measurements on
proinflammatory and prothrombotic factors at baseline and during the trial follow-up
(R01-AG-18798, Howard N. Hodis, PI). We summarized the preliminary results
from EPAT in Tables 92-9.4. Among 199 subjects with CIMT progression data, 84
(42%) subjects met the diagnosis of MetS according to the NCEP ATP m criteria,
while 115 (58%) did not (Table 9.2).
Table 9.2 Baseline Characteristics of 199 Evaluable Subjects in EPAT
Variable MS Present MS Absent p- value1
(N=84) (N=I 15)
CIMT, pm 804.9 (149.9) 734.2 (107.8) 0.00032
AGE (year) 63.7 (6.9) 59.8 (6.5) <0.0001
Ethnicity, n(%)
White 54 (64) 64 (56) 0.007
Black 5(6 ) 17(15)
Hispanic 22 (26) 18(16)
Asian and other 3 (4 ) 16(14)
Education, n(%)
< 12 y 2(2 ) 2 (2 ) 0.75
>12 y 82 (98) 113 (98)
Diabetes mellitus, n(%) 5 (6 )
1(1)
0.04
Smoking history, n(%)
Former smoker 46 (55) 57 (50) 0.47
Never smoked 38 (45) 58 (50)
Blood pressure, mmHg
Systolic 133.5 (12.7) 123.6(15.7) <0.0001
Diastolic 78.6 (7.2) 76.7 (9.5) 0. 12
Waist circumference (inch) 37.2 (4.4) 33.4 (4.5) <0.0001
Weight (kg) 80.9(15.0) 70.1 (14.7) <0.0001
BMI (kg/m2 ) 30.9 (5.2) 27.2 (5) <0.0001
Fasting glucose (mg/dL) 95.3 (14) 85.7 (8.1) <0.0001
Hemoglobin A le (%) 5.2 (0.7) 4.9 (0.4) 0.002f
Fasting insulin (uU/ml) 12.4 (7.6) 7.4 (3.2) <0.0001
HOMA index 3.0 (2.2) 1.6 (0.7) <0.00012
Total cholesterol (mg/dL) 255.6 (34.1) 244.3 (30.6) 0.015
Triglycerides (mg/dL) 209.4 (75.4) 121.9 (45) <0.0001
HDL-cholesterol (mg/dL) 45.9 (8) 59.5 (11) <0.0001
LDL- cholesterol (mg/dL) 168.7(29.5) 160.4 (26.9) 0.04
1. P-value from independent t-test or chi-square test
2. P-value from t-test for unequal variances
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Compared to subjects without the MetS, subjects with the MetS were older,
more likely to be of Hispanic origin, and were more likely to have diabetes. The
MetS group also had significantly higher levels of hs-CRP, sICAM-1, fibrinogen,
PAI-1 antigen and tPA antigen after adjustment for age and ethnicity (Table 9.3).
PAI-1 antigen and fibrinogen were only measured at the end of study on 138
subjects. All analyses on these two variables were therefore adjusted for randomized
treatment assignment to control for the potential confounding effect of unopposed
estradiol.
Table 9.3 Adjusted Comparisons of Inflammatory and Hemostatic Factors by MetS group: EPAT
MetS Present MetS Absent P-value1
hsCRP (pg/ml) 2.7(0.2)2 1.9(0.2) 0.008
SICAM-1 (ng/ml) 305.2(6.4) 277.2(5.3) 0.001
Fibrinogen (mg/dl) 401.6(9.1) 373.6(8.1) 0.03 3
PAI-1 antigen (ng/ml) 12.8(1.2) 7.1(1.1) 0.00073
tPA antigen (ng/ml) 13.9(0.5) 12(0.4) 0.004
Ddimer (ng/ml) 649.2(39.8) 609(32.9) 0.45
Factor VII antigen (%) 140.3(3.5) 132(2.9) 0.08
1. ANCOVA, adjusting for age and ethnicity
2. Adjusted mean (SE)
3. ANCOVA, adjusting for age, ethnicity and treatment assignment
Moreover, increased hsCRP concentration was significantly associated with
increased overall and centralized adiposity, elevated SBP, increased triglycerides,
decreased HDL-C, increased fasting glucose and insulin concentration, and insulin
resistance as determined by homeostasis model assessment (HOMA) (Table 9.4).
Similar associations were also observed for slCAM-1, PAI-1 and fibrinogen except
some of the associations were not statistically significant. These results support
markers of inflammation and hemostasis as components of the MetS.
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The correlations between hsCRP and hemostatic factors, except for D-dimer
and factor VII, were stronger in subjects with the MetS than in those without the
syndrome (Table 9.5). The stratum-specific correlations were not significantly
different from one another due to limited power of these preliminary data (data not
shown). Further analyses excluding 6 diabetic subjects yielded similar results (data
not shown).
Table 9.4 Partial Spearman Correlation Coefficients o f Inflammatory and Hemostatic Factors and Other
Components o f the MS (N=199)
hsCRP 1 sCIAM-1 1 PAI-1 2 Fibrinogen 2
BMI 0.4(<0.0001) i 0.23(0.001) 0.34(<0.0001) 0.26(0.002)
Waist 0.41(<0.0001) 0.22(0.003) 0 .3 8 (0 .0 0 0 1 ) 0.3(0.0005)
SBP 0.22(0.004) -0.01(0.88) 0.09(0.29) 0.08(0.34)
Fasting glucose 0.2(0.012) 0.07(0.36) 0.32(0.0002) 0.07(0.45)
HOMA 0.27(0.0004) 0.26(0.0005) 0 .4 2 (0 .0 0 0 1 ) 0.20(0.02)
Fasting insulin 0.29(0.0002) 0.3(<0.0001) 0 .4 2 (0 .0 0 0 1 ) 0.22(0.011)
Triglyceride 0.17(0.022) 0.28(0.0001) 0.3 4 (0 .0 0 0 1 ) 0.1(0.26)
HDL cholesterol -0.2(0.007) -0.33(<0.0001) -0.12(0.15) -0.18(0.04)
1. Adjusted for age and ethnicity
2. Adjusted for age, ethnicity and treatment assignment
3. r (p-value)
Table 9.5 Pearson Partial Correlation Coefficients o f hsCRP and sCIAMl with hemostatic factors: EPAT
Partial Correlations with hsCRP ( ug/m l)1
MS Present MS Absent
Fibrinogen (mg/dl) 2 0.28(0.041) 0.22(0.07)
PAI-1 antigen (ng/ml)2 0.29(0.036) 0.07(0.57)
tPA antigen (ng/ml) 0.35(0.004) 0.06(0.53)
Ddimer (ng/ml) 0.06(0.66) 0.39(<0.0001)
Factor VII antigen (%) 0.07(0.58) 0.02(0.82)
Partial Correlations with sICAMl fna/ml')1
MS Present MS Absent
Fibrinogen (mg/dl) 2 0.12(0.37) 0.05(0.65)
PAI-1 antigen (ng/ml)2 -0.003(0.98) -0.11(0.36)
tPA antigen (ng/ml) 0.24(0.04) 0.18(0.06)
Ddimer (ng/ml) -0.03(0.82) 0.01(0.89)
Factor VII antigen (%) 0.05(0.69) 0.21(0.031)
1. Adjusted for age and ethnicity
2. Adjusted for age, ethnicity and treatment assignment
3. r (p-value)
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9.4 Research Design and Methods
9.4.1 Pooled Study Design
In the pool study design, all five trials were randomized, single-center,
double-blind, placebo-controlled, clinical trials conducted at the Atherosclerosis
Research Unit at USC in the past decade. The annual rate of change in CIMT was
the primary measure of subclinical atherosclerosis progression. Baseline and on-trial
clinical data from five trials will be pooled to conduct a factor analysis on all
components of the MetS to generate latent factor scores. The underlying assumption
of factor analysis is that there exist a number of unobserved latent factors that
account for the correlations among observed variables. These latent factor scores for
each subject at each clinic visit will be used as independent variables in the mixed
effects models, with the rate of change in CIMT as the primary outcome.
In particular, to achieve our primary aim, we will compare model fitting
between two mixed effects models. In the first model, we will include the first set of
metabolic variables, defined by the current definition of the MetS, in the factor
analysis to generate the first set of factor scores. The association between these
factor scores and CIMT progression will then be examined using the mixed effects
models. In the second model, the markers of inflammation and hemostasis will be
added in the factor analysis, which we hypothesize may generate greater factor
scores and thereby provide better prediction of CIMT progression.
Two major strengths of our factor analysis approaches are (1)
continuous variables rather than arbitrary dichotomy are sustained throughout
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the analyses (2) each component of the MetS is given different weight depending
on its correlation coefficients with latent factors. Eventually, weighted
summary scores are generated rather than a binary variable of the MetS.
Each trial intervention, usage of antihypertensive, lipid lowering and anti
inflammatory medications during the trial, and age at randomization will be included
as potential confounders. The pooled study cohort has a wide range of age at
randomization (30-88 years old) and various cardiovascular risk profiles, including
the presence or absence of CVD, and the presence or absence of diabetes. The
associations between factor scores and CIMT progression will be investigated in the
entire sample and various subgroups, such as subjects with CVD, or subjects with
diabetes.
9.4.2 Ultrasonography and CIMT Measurement
Carotid artery ultrasonography for measurement of CIMT was conducted at
baseline and every six months during each trial by a single ultrasonographer using
high resolution B-mode carotid artery ultrasound (ATL ultrasound imager equipped
with a linear array 7.5 MHz probe). The ECG signal and ultrasound images were
simultaneously recorded on videotape. Blood pressure was recorded throughout the
procedure using a Dinamap XL vital signs monitor (Johnson & Johnson Medical
Inc., Arlington, TX). Subjects were placed supine and the neck positioned to present
the optimal angle for ultrasound examination using a 45 head block apparatus. The
common carotid artery was first imaged in cross section with the jugular vein
juxtaposed above the carotid artery. The scanhead was then rotated 90 degrees
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around the central image line, maintaining the jugular vein stacked above the CCA
while obtaining a longitudinal view of both vessels. The proximal portion of the
carotid bulb was included in all images as an anatomical reference point for
standardization of IMT measurements. Minimum gain necessary for clear
visualization of structures was used. Once the transducer was positioned, the image
was recorded for a minimum of 60 seconds. Automated edge detection to locate the
lumen-intima and media-adventitia echo boundaries at sub-pixel resolution was used
to measure CIMT (Prowin, patent pending). The CIMT measurement consisted of
an average of 80-100 independent measurements made along a 1 cm distance of the
distal right common carotid artery on the far wall. Using this methodology, the
coefficient of variation for average CIMT (replicate scans on the same day) is 2.5%
within operators. Variability of CIMT measurement is reduced 2-4 times compared
to that obtained with manual tracking methods (74).
9.4.3 Clinical Data Collection
The methods for clinical data collection were consistent across trials as all
trials were conducted in the same research clinic with a stable research staff during
the past decade. At study entry for each trial, subjects were interviewed with
standardized structured questionnaires to ascertain their age, gender, ethnicity, years
of education, history of diabetes and CVD, family history of CVD, smoking history,
alcohol use and menopausal status. Height, weight, waist and hip circumference
were measured at baseline and at scheduled visits throughout the trial and body mass
index (BMI) and waist-hip-ratio (WHR) were derived from these data. BMI was
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calculated as weight (kilograms)/height (meters)2. Blood pressure was measured
twice after subjects sat with legs dangling for at least 5 minutes and the average was
computed.
9.4.4 Laboratory Measurements
The laboratory measurements were consistent across trials as all the
measurements were performed by the same laboratory. Blood samples were
obtained at baseline and at scheduled visits throughout the trial after an 8-12 hour
overnight fast. Leukocyte count was measured by standard methods (Coulter
counters). Serum glucose levels were measured by glucose oxidase (Beckman
Glucose analyzer II, Beckman Instruments, Brea CA, USA). Hemoglobin Ale levels
were measured by high-pressure liquid chromatography (BioRad, Hercules CA,
USA). Fasting plasma insulin levels were measured by radioimmunoassay that
provides < 0.2% cross-reactivity with proinsulin. Although there was no direct
measure of insulin resistance (IR), a surrogate measure of IR will be assessed using
the homeostatis model assessment (HOMA). IR will be calculated as fasting insulin
(pU/mL) X fasting glucose (mmol/L) / 22.5 (210).
Plasma total cholesterol, total triglyceride, and HDL-C levels were
determined by enzymatic assays and standardized to the CDC using the Lipid
Research Clinic protocol. HDL-C levels were measured after apolipoprotein B-
containing lipoproteins were precipitated in whole plasma with heparin manganese
chloride. Low-density lipoprotein cholesterol (LDL-C) levels were estimated using
the Friedewald equation (146).
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9.4.5 Non-study Medication Use
Throughout each trial, a standard medication record form was used to record
the use of all prescription medications, including antihypertensive agents, lipid-
lowering medications and anti-inflammatory medications (dose, frequency and dates
of use). Subjects were requested to bring all medications to each clinic visit for
accurate ascertainment of non-study medications. For each medication, the date of
first use, date of last use, dosage and frequency, and reasons for use were recorded.
A common medication-coding scheme was used for all trials.
9.4.6 Lifestyle Assessment
Nutritional information was collected at baseline and every 6 months during the
trial with the Nutrient Analysis System 3-Day Food Reporter that utilized the USDA
database to analyze the quantity and types of food consumed for 3 consecutive days
including 2 weekdays and 1 weekend day prior to the clinic visit. This system allows
food not depicted in the booklet to be added to the daily intake. The resulting dietary
database comprises intake on 64 macro- and micronutrients including total calories,
proteins by major source, fats by major type as well as individual fatty acids,
carbohydrates by major type, vitamins, minerals, and fiber (solube and insoluble).
Physical activity information was collected at baseline and every 6 months
during the trial with Stanford 7-Day Physical Activity Recall. The form records
specific activity (at least moderate intensity) and duration of all activities for 7
consecutive days including 5 weekdays and 2 weekend days prior to the clinic visit.
A common activity-coding scheme (Compendium of Physical Activites) was used
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for all trials. The resulting physical activity dataset will allow us to evaluate physical
activity by estimated energy expenditures and by time spent in specific types of
activities.
9.4.7 New Data Collection
9.4.7.1 Variables to be Measured
In each trial, a minimum of 30ml of extra plasma and serum were collected at
baseline and scheduled visits throughout the trial, stored at -70°C for back-up and
future studies. The USC Core Lipid Laboratory conducted sample storage. We list
all variables to be collected by trials in Table 9.6. All laboratory assays will be
performed concurrently on previously frozen samples.
Hemoglobin A le levels will be measured by high-pressure liquid
chromatography (BioRad, Hercules, California). Fasting plasma insulin levels will
be measured by radioimmunoassay that provides < 0.2% cross-reactivity with
proinsulin (Diagnostic Products Corporation, Malvern, Pennsylvania). Fibrinogen
will be measured using an immunoephelometric method (211). PAI-1 (212), tPA
antigen (213) and factor VII antigen (214) will be analyzed by enzyme-linked
immunosorbent assays (ELISA) as previously described. Plasma hsCRP and
sIC AMI levels will be measured by ELISA assays with a coefficients of variance of
5.2 and 5.0%, respectively (215).
9.4.7.2 Data Transmission
Laboratory data will be obtained on hard copies and sent to the Data
Coordinating Center (DCC) with data entry at the DCC. Otherwise, if data from the
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laboratory are transmitted to DCC using electronic media, text containing identifying
information will not be included in the data files.
9.4.8 Data Quality Control
For the existing data, data quality control had been implemented through
each trial period. As for CIMT data, we did not face the quality control challenges
of a multicenter study since all five trials were single-center trials with a stable team
of ultrasonographers. In terms of clinical data, clinic coordinators reviewed all study
forms for completeness and accuracy prior to sending to data clerk for data entry.
As for laboratory data, we monitored timeliness of receipt of laboratory data and end
point data and identify outstanding laboratory samples. In terms of lifestyle data, the
nutrient analysis system we used had a minimum of 90% compliance with study pill
count in five trials.
Prior to data entry, all forms are inventoried and undergo a 2-step review
(study coordinator and data monitor) for completeness and apparent accuracy;
incomplete/inaccurate forms are returned to the study coordinator for rectification.
Key features of the data entry system at the time of data entry include: data entry
screens that mimic the data forms, must-enter fields for key variables, and range
checks. Key laboratory data are double entered by 2 individuals and program-
compared before addition to a master database.
For newly entered data, error check routines were run to identify logical
errors in the data, missing data, out-of-range or implausible values, and data outliers
(both within a given visit and in comparison to a subject’s earlier visits). A
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structured database was set up and a logic check program was developed to avoid
double entry for the same record, and to ensure sensible ranges for continuous
variables. Every discrepancy was checked against hard copies or clinical charts for
corrections. The data quality report was generated once a month and for the Data
Safety Monitoring Board. The report included data on the number of errors in
completion of study forms, summarization of missed visits and visits outside of the
ideal visit window. Inaccuracies as well as incompleteness in data forms were
considered errors and were computed by study form. For new data to be collected,
the same scheme will be carried out to ensure laboratory data quality and data entry
quality.
9.4.9 Statistical Analyses
All analyses will be restricted to subjects with known measurements of all
variables of interest listed in Tables 9.6 and 9.7. The distributions of each variable
will be evaluated for normality. Log transformations will be performed when the
distributions are skewed. Descriptive analysis will be performed with mean (SD,
range) for continuous variables and cross tabulation for categorical variables. The
planned analyses described below are detailed by the major specific aims.
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Table 9.6 Baseline and ontrial Measurements o f Five Atherosclerosis Progression Trials
EPAT VEAPS WELL-HART TART BVAIT
Trial Endpoint
CIMT (pm) x 1 X X X X
Clinical Data
Age at randomization (yr) X X X X X
Gender (M/F) X X X X X
Ethnicity X X X X X
BMI (kg/m2 ) X X X X X
Waist (cm) X X X X X
Flip (cm) X X X X X
History of CVD (Y/N) X X X X X
Smoking (packyear) X X X X X
Alcohol use X X X X X
Diabetes (Y/N) X X X X X
Menopause (Y/N) X X X X X
Family history o f CVD (Y/N) X X X X X
SBP (mmHg) X X X X X
DBP (mmHg) X X X X X
Laboratory Data
Total cholesterol (mg/dl) X X X X X
Triglyceride (mg/dl) X X X X X
LDL cholesterol (mg/dl) X X X X X
HDL cholesterol (mg/dl) X X X X X
Glucose (mg/dl) X X X X X
WBC (109 /L) X X X X X
1. measured in the original study
Table 9.7 New Laboratory Data to be Collected in the Proposed Work
EPAT VEAPS WELL-HART TART BVAIT
Insulin (jiU/ml) x 1 + 2 X X +
HbAlC (%) X + X X +
Fibrinogen (mg/dl) X + + X +
tPA antigen (ng/ml) X + X +
PAI-1 antigen (ng/ml) X + + X +
factor VII antigen (%) X + + + +
hsCRP (pg/ml) X + + + +
SICAM-1 (ng/ml) X + + + +
1. measured in the original study
2. to be measured for the pooled analysis
9.4.9.1 Specific Aim 1, Step 1: By principal component analysis, we will identify
principal components of the MetS that are statistically independent of one
another using two sets of measured variables: (1) the standard 5 metabolic
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variables used in the NCEP ATP III definitions of MetS (2) the standard 5
metabolic variables plus inflammatory and hemostatic variables.
The MetS can present in various ways due to different components that
constitute the syndrome; thus it is critical to identify and quantify these
constellations of highly interrelated metabolic abnormalities. As a useful tool for
data reduction and summarization, factor analysis has been widely used to assess the
correlation between a large set of observed variables by extracting latent factors.
Principal component analysis is one of the most popular approaches to extract
factors. By new data collection, we will have both baseline and on-trial
measurements of two sets of metabolic components of the MetS, we propose to
perform statistical analyses in a time dependent manner.
First of all, we will include each set of metabolic variables at baseline in the
principal component analysis with Varimax orthogonal rotation (using the PROC
FACTOR procedure in SAS). Latent factors will be selected when the eigenvalues
are > 1. An eigenvalue of 1.0, known as the Kaiser-Guttman rule, is a commonly
used threshold recommended on factor analysis (ref factor analysis book). After
retaining factors, a Varimax orthogonal rotation will be implemented so that
measured variables will have high loadings on one factor but not others. Factor
rotation facilitates interpretation of the latent factors. The rotated factor loading of a
variable on a latent factor can be interpreted as the Pearson correlation coefficient
between the original variable and its corresponding latent factor. Higher loadings
indicate stronger associations between measured variables and associated latent
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factors. Only variables with factor loading value > 0.3 will be identified as
significant loadings (this criterion may need to be adjusted when necessary). If a
variable fails to load significantly on any factor, the variable will be eliminated and
the analysis will be repeated. Once all the significant loadings for each factor are
identified, we can interpret what the underlying factor may present.
Factor scores will then be computed for each factor and each subject (using
the PROC SCORE procedure in SAS). For each subject, a factor score of each factor
is a weighted sum of the standardized values of the variables multiplied by the factor
loadings in that particular factor. Then we will apply factor solutions from baseline
to get factor scores for on-trial metabolic variables. Consequently, for a given
clinical visit, the principal component analysis will yield two sets of factor scores
representing two sets of principle components of the MetS.
Two major strengths of our factor analysis approaches are (1) continuous
variables rather than arbitrary dichotomy are sustained throughout the analyses (2)
each component of the MetS is given different weight depending on its correlation
coefficients with latent factors. Eventually, weighted summary scores are generated
rather than a binary variable of the MetS.
9.4.9.2. Specific Aim 1, Step 2: To investigate the associations between the two
sets of principal components of the MetS and progression of CIMT in the
pooled sample, and determine the best fitting model.
As the principal component analyses will yield two sets of factor scores at
each clinical visit, we will perform the following analyses on baseline factor scores
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and ontrial factor scores, respectively, with a systemic modeling strategy (see Table
9.8 for details).
Table 9.8 Proposed statistical modeling using multivariate linear mixed effects models
Model Outcome Independent variables Potential Covariates
1 CIMT rate 1st set o f baseline factor 1) age at baseline
scores 1 2) each trial intervention
2 CIMT rate 1st set o f baseline factor 1) age at baseline
scores
2)
each trial intervention
3)
ontrial use o f antihypretensive agent
4)
ontrial use o f lipid modifying meds
5) ontrial use o f anti-inflammatory meds
3 CIMT rate 1st set o f baseline factor 1) age at baseline
scores
2)
each trial intervention
3)
ontrial use of antihypretensive agent
4) ontrial use of lipid modifying meds
5) ontrial use of anti-inflammatory meds
6)
ontrial change in weight, waist, blood pressure,
HDL-C, triglycerides and glucose 2
4 CIMT rate 2n d set o f baseline factor 1) age at baseline
scores 3
2)
each trial intervention
5 CIMT rate 2n d set o f baseline factor 1) age at baseline
scores 2)
each trial intervention
3)
ontrial use o f antihypretensive agent
4)
ontrial use o f lipid modifying meds
5) ontrial use o f anti-inflammatory meds
6 CIMT rate 2n d set o f baseline factor 1) age at baseline
scores
2)
each trial intervention
3)
ontrial use o f antihypretensive agent
4)
ontrial use o f lipid modifying meds
5) ontrial use o f anti-inflammatory meds
6)
ontrial change in weight, waist, blood pressure,
HDL-C, triglycerides and glucose 2
7 CIMT rate 1st set o f time-dependent 1) age at baseline
factor scores 4
2)
each trial intervention
8 CIMT rate 1st set o f time-dependent 1) age at baseline
factor scores
2)
each trial intervention
3)
ontrial use o f antihypretensive agent
4)
ontrial use o f lipid modifying meds
5) ontrial use o f anti-inflammatory meds
9 CIMT rate 2n d set o f time-dependent 1) age at baseline
factor scores5
2)
each trial intervention
10 CIMT rate 2n d set of time-dependent 1) age at baseline
factor scores
2)
each trial intervention
3)
ontrial use o f antihypretensive agent
4)
ontrial use o f lipid modifying meds
5) ontrial use of anti-inflammatory meds
1. 1s t set o f baseline factor scores will be generated from the baseline data o f the standard 5 metabolic factors
2. calculated as ontrial average minus baseline
3. 2n d set o f baseline factor scores will be generated from the baseline data o f the standard 5 metabolic factors
plus inflammatory and hemostatic variables
4. 1st set o f time-dependent factor scores will be generated from the longitudinal data o f the standard 5
metabolic factors 2n d set o f time-dependent factor scores will be generated from thelongitudinal data o f the
standard 5 metaboli factors plus inflammatory and hemostatic variables
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For baseline factor scores in the pooled sample, a multivariate mixed effects
model will be fitted to evaluate the rate of change in CIMT in relation to each set of
baseline factor scores (using the PROC MIXED procedure in SAS). The annual rate
of change in CIMT was a trial endpoint in all trials. CIMT will be regressed on
follow-up time (years since randomization) to estimate the average rate of change in
CIMT in each subject. The baseline factor scores will be included as independent
variables. Whether the principal components of the MetS at baseline influence the
CIMT progression rate will be tested by two-way interaction terms (follow-up
time*factor scores).
In terms of covariates in the models, we will first include age at
randomization and dummy variables of study intervention in each trial. Then,
dummies variables for use of nonstudy medications (continuously or intermittently
receiving hormone, antihypertensive, lipid modifying and anti-inflammatory therapy
no less than three months during each trial) will be created and used as covariates to
control for their potential beneficial effects on CIMT progression. Furthermore, on
trial changes in weight, waist, blood pressure, HDL-C, triglycerides and glucose (on
trial average - baseline) will be added selectively in the models to control for the
influence of change in metabolic variables over time on CIMT progression during
the follow-up. Of note, adjustment for on-trial changes in the metabolic variables
will be selective based on whether they are associated with the factor scores.
Akaike’s Information Criterion (AIC) will be computed to compare the fit of the
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models. Whichever model has the largest AIC will be used as the best model for
further subgroup analyses.
For longitudinal Factor Scores in the pooled sample, a multivariate mixed
effects model will be fitted to evaluate the rate of change in CIMT in relation to each
set of factor scores (using the PROC MIXED procedure in SAS). CIMT will be
regressed on follow-up time (years since randomization) to estimate the average rate
of change in CIMT in each subject. The factor scores will be included as time-
dependent independent variables. Whether the principal components of the MetS
influence the CIMT progression rate will be tested by two-way interaction terms
(follow-up time*factor scores).
In terms of covariates in the models, we will first include age at
randomization and dummy variables of study intervention in each trial. Then,
dummies variables for use of nonstudy medications (continuously or intermittently
receiving hormone, antihypertensive, lipid modifying and anti-inflammatory therapy
no less than three months during each trial) will be created and used as covariates to
control for their potential beneficial effects on CIMT progression. Akaike’s
Information Criterion (AIC) will be computed to compare the fit of models.
Whichever model has the largest AIC will be used as the best model for further
subgroup analyses.
9.4.9.3 Specific Aim 2: To examine whether the associations between principal
components of MetS and CIMT progression vary by CVD status and diabetes
status.
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Once a significant two-way interaction term (follow-up time*factor score) is
found, three-way interaction terms (follow-up time*factor score*CVD status, follow-
up time*factor score*diabetes status) will be performed to examine whether the
associations between metabolic factor scores and CIMT progression differ by CVD
status and diabetes status. If a significant effect modification on a multiplicative
scale is observed, a stratified analysis will be conducted accordingly to further
illustrate the different associations of the metabolic factor scores and CIMT
progression rates in each stratum. Because few data are available to examine these
associations between gender and Hispanics had the highest age-adjusted prevalence
of the MetS among all US ethnic groups (152), we will also explore whether the
associations between metabolic factor scores and CIMT progression vary by gender
and Hispanics vs. non-Hispanics.
To rule out the effect of trial interventions on CIMT progression, we also
plan to repeat all the above analyses using the pooled data with placebo group only.
All analyses will be conducted using Statistical Analysis System Version 9.0 (SAS
Institute, Cary, NC). All analyses will be two-sided at a significance level of 0.05
unless otherwise indicated.
We will not include dietary nutrients and physical activity variables in the
linear mixed effects models because (1) these variables directly influence key
components of the MetS, such as visceral obesity, hyperglycemia, and (2) the
adjustment for these variables would likely be adjusting out any effects of the MetS
on the progression of subclinical atheroscleoris.
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9.4.9.4 Power Estimation
•y
Assuming a 10% lost on frozen blood samples and R for known 15
covariates = 0.1, a sample size of 1539 subjects has 90% power at the two-sided 0.05
significance level to detect a correlation coefficient of 0.08 between baseline factor
scores and annual change of CIMT. Table 9 summarized the power calculations for
the total sample and each subgroup with various assumption of R for known 15
covariates.
Table 9.9 Power Estimations (two-sided significance level = 0.05)
Sample Subjects Correlation Correlation Correlation Correlation
size Available 1 coefficient coefficient coefficient coefficient
(pooled)
2 3 4 5
Power = 80%
Total sample 1539 1385 0.07 0.06 0.0 5 0.04
CVD present 247 222 0.18 0.16 0.13 0.10
Diabetes Present 397 357 0.14 0.12 0.10 0.08
Hispanics 472 424 0.13 0.11 0.10 0.07
Women 980 882 0.09 0.08 0.07 0.05
Men 559 503 0.12 0.10 0.09 0.07
Power = 90%
Total sample 1539 1385 0.08 0.07 0.06 0.05
CVD present 247 222 0.2 0.18 0.15 0.12
Diabetes Present 397 357 0.16 0.14 0.12 0.09
Hispanics 472 424 0.15 0.13 0.11 0.09
Women 980 882 0.1 0.09 0.08 0.06
Men 559 503 0.14 0.12 0.1 0.08
1. Sample size after accounting for a 10% lost o f stored blood sample.
2. Correlation coefficients between one factor score and annual change o f CIMT assuming R2 for known 15
covariates = 0.1
3. Correlation coefficients between one factor score and annual change o f CIMT assuming R2 for known 15
covariates = 0.3
4. Correlation coefficients between one factor score and annual change o f CIMT assuming R2 for known 15
covariates = 0.5
5. Correlation coefficients between one factor score and annual change of CIMT assuming R2 for known 15
covariates = 0.7
9.5 Human Subjects Research
9.5.1 Protection of Human Subjects
9.5.1.1 Risks to the Subjects
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a. Human Subjects Involvement and Characteristics
Subjects were recruited by study investigators and staff via direct contact
with the individuals in the original trials. A total of 1539 subjects, aged 30-88 years,
will be analyzed. Among them, 980 (64%) were women, 397 (26%) had diabetes,
247 (16%) had a positive history of CVD, 878 (90% of female) were
postmenopausal women, 776 (50%) were White, 182 (12%) were Black, 472 (31%)
were Hispanic, 97 (6%) were Asian and 12(1%) were of other ethnicity. A more
detailed description of the subject population iss provided in Table 9.1.
b. Sources of Materials
Data for the proposed analyses will primarily come from the existing
databases of five randomized trials conducted at USC during the period of 1994-
2005. Information in the database was obtained from subjects during each trial
period. Data were collected on paper data collection forms and then entered into a
central computerized database. The original data were collected from three sources:
(1) interview administering standardized questionnaires, including age, age, gender,
ethnicity, years of education, history of diabetes and CVD, family history of CVD,
smoking history, alcohol use and menopausal status (2) clinical measurements,
including height, weight, waist, hip circumference and blood pressure (3) laboratory
measurements, including fasting glucose, insulin, HbAlc, lipid leves, and total
homocysteine levels (4) carotid artery ultrasonography for measurement of CIMT
during the trial.
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c. Potential Risks
The potential risk to study subjects is loss of confidentiality of medical
information.
9.5.1.2 Adequacy of Protection Against Risks
a. Recruitment and Informed Consent
No new recruitment will be undertaken. All subjects had consented for using
their data and further analyzing their stored samples for future studies when they
were enrolled in each of the five trials. Waivers of individual informed consent and
HEPAA (for data verification via chart review) will be sought from the USC IRB for
the current proposal.
b. Protection Against Risk
Original data forms and clinical charts of each trial are stored at the
Atherosclerosis Research Unit in locked cabinets. Electronic versions of the
databases of each trial are kept on a secured, password-protected network that
requires special administrative approval for access. In the database, each subject has
a unique study ID number and there are no subject identifiers that can link individual
subjects to their data. Results to be presented or published will not identify
individual study subjects.
9.5.1.3 Potential Benefits to the Subjects and Others
Individual subjects and others will benefit from the proposed analysis by
gaining knowledge of the impact of various clustering patterns of the MetS on
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subclincial atherosclerosis progression and their possible differential impact on
persons with CVD or diabetes.
9.5.1.4 Importance of the Knowledge to be Gained
The pooled database from five atherosclerosis regression trials is a unique
database that allows us to investigate the underlying structures of the MetS before
and after incorporating proinflammatory and prothrombotic factors in the analysis.
The proposed analysis will thereby provide an evidence-based analysis to
evaluate the value of adding markers of inflammation and hemostasis to the
definition of MetS. Moreover, we will gain unique observational information
about potential risks of core factors of the MetS on atherosclerosis progression
among various high-risk populations, such as subjects with CVD, or subjects
with diabetes. Evaluating the predictive value of inflammatory and hemostatic
marker on atherosclerosis progression may improve cardiovascular risk
prediction and further evaluation of antiatherogenic interventions.
9.5.2 Inclusion of Women
The subjects of the EPAT and WELL-HART studies were all
postmenopausal women due to their study designs. There were 172 (52%), 186
(67%) and 197 (39%) women in the VEAPS, TART and BVAIT trials, respectively.
Overall, the pooled data will include 980 (64%) women for the analysis. The
relatively balanced gender mix will enable us to explore the hypotheses of interest in
each gender subgroup.
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9.5.3 Inclusion of Minorities
All ethnic groups were eligible to participate in the five clinical trials. In the
pooled data, 776 (50%) were White, 182 (12%) were Black, 472 (31%) were
Hispanic, 97 (6%) were Asian and 12 (1%) were other. Given the sufficient sample
size of Hispanic subjects, a subgroup analysis is warranted.
9.5.4 Inclusion of Children
Children were excluded from all five trials because the research topics
studied in all five trials were not relevant to children. Moreover, a standard
definition of the MetS in children and adolescents is not available. The minimum
age in the pooled sample is 30 years old.
9.6 Study Timeline
Table 9.10 Proposed Study Timeline
Months after study Initiation Progress
1-4 Samples to the labs
1-6 Set up structured database for pooled data
4-12 Data quality control (data cleaning)
13-18 Data analysis
19-24 Final report, manuscript writing
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Progression of subclinical atherosclerosis in type 2 diabetes: Therapeutic and metabolic influences
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