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Human histological analysis of autogenous block grafts

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Human histological analysis of autogenous block grafts

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HUMAN HISTOLOGICAL ANALYSIS OF AUTOGENOUS BLOCK GRAFTS

by

Andrew Kim

____________________________________________________

A Thesis Presented to the
FACULTY OF THE USC GRADUATE SCHOOL
UNIVERSITY OF SOUTHERN CALIFORNIA
In Partial Fulfillment of the
Requirements for the Degree
MASTER OF SCIENCE
(CRANIOFACIAL BIOLOGY)

December 2012

Copyright 2012 Andrew Kim

ii

Dedication

This thesis is dedicated to my friends and family who have always supported me
through the good times and the bad. Thank you for encouraging and motivating me to
always move forward. Special thanks to my mother, Eun-Sook Kim, who has sacrificed
very much for me. Your unconditional love will be always remembered.

iii

Acknowledgements

I would like to thank my program director, Dr. Hessam Nowzari, for your
mentorship and guidance was greatly appreciated. Thank you Dr. Sandra Rich for your
support in the administrative components and evaluation of the project. Thank you Dr.
Kar for your help in designing and executing the clinical components of the project.
Thank you Dr. Kang Min Ahn and Dr. Seonghong Min for your crucial thoughts and
feedback in interpreting the results. I would also like to thank Dr. Mahvash Navazesh for
providing me invaluable advice on my research.
Thank you Dr. James Adams and Dr. Lina He for assisting me in the laboratory
components of the research. Thank you Dr. Yang Chai and Pablo Bringas for helping me
with the histological photography.

iv

Table of Contents
Dedication ii
Acknowledgements iii
List of Figures v
Abstract vi
Chapter 1: Introduction and Background 1
Chapter 2: Materials and Methods 21
Chapter 3: Results 27
Table 1. Clinical synopsis 28
Chapter 4: Discussion 45
Chapter 5: Conclusion 51
References 52

v

List of Figures

Figure 1. Recipient site defect 22
Figure 2. Donor bone harvest 22
Figure 3. Graft fixated in place 23
Figure 4. Healed block graft 24
Figure 5. Bone core harvesting 24
Figure 6. Bone core in 10% NBF 25
Figure 7. Case 1(a): histologic overview 30
Figure 8. Case 1(b): 20x magnification 30
Figure 9. Case 1(c): 20x magnification 31
Figure 10. Case 1(d): 20x magnification 31
Figure 11. Case 1(e): 40x magnification 32
Figure 12. Case 1(f): 40x magnification 32
Figure 13. Case 2(a): histologic overview 34
Figure 14. Case 2(b): 10x magnification 34
Figure 15. Case 2(c): 10x magnification 35
Figure 16. Case 2(d): 20x magnification 35
Figure 17. Case 2(e): 20x magnification 36
Figure 18. Case 2(f): 20x magnification 36
Figure 19. Case 2(g): 40x magnification 37
Figure 20. Case 2(h): 40x magnification 37

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Figure 21. Case 3(a): histologic overview 38
Figure 22. Case 3(b): 20x magnification 39
Figure 23. Case 3(c): 20x magnification 39
Figure 24. Case 3(d): 40x magnification 40
Figure 25. Case 3(e): 20x magnification 40
Figure 26. Case 3(f): 40x magnification 41
Figure 27. Case 3(g): 40x magnification 41
Figure 28. Case 4(a): 40x magnification 43
Figure 29. Case 4(b): 40x magnification 43
Figure 30. Case 4(c): 40x magnification 44

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Abstract

Background: Autogenous block bone grafting is a treatment option for ridge deformities.
Currently, the healing process of autogenous bone grafting in humans has not been
histologically reported and the vitality of bone after graft placement has been challenged.
Objective: The purpose of this study is to histologically evaluate vitality of autogenous
bone grafts in patients who have been treated for ridge deformities in preparation for
implant placement.
Materials and Methods: At 4, 5, and 7 months after bone augmentation and at the time
of implant placement, a core of 2x3 mm bone graft was trephined for histological
analysis (n=4; mean age 37.3 years; 1:1 male/female). Samples were fixated with 10%
neutral buffered formalin (NBF), decalcified, embedded in paraffin, and stained with
Hemotoxylin & Eosin. Angiogenesis, bone vitality (presence of osteocytes within
lacunae), and the Haversian systems were examined.
Results: Bleeding was observed at the time of sample collection. All surgeries healed
uneventfully and all implants were stabilized at 50 Ncm. Diffuse angiogenesis and
cellular vitality were identified in all sections. Osteogenesis resulted in vital bone
formation, which was demonstrated by the presence of osteocytes within lacunae, lining
osteoblasts, different staining color, immature organization, and demarcation lines. Vital
bone was adjacent to blood supply and was amalgated with graft bone. Symphysis grafts
exhibited more angiogenesis and osteogenesis than ramus grafts.
Conclusion: This human histologic study demonstrated that autogenous bone grafting

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leads to vital bone, confirmed by bleeding at the time of sample collection, histologic
detection of angiogenesis, osteogenesis, and diffuse cellular activity. Remodeling was
ongoing at 4, 5, 7 months and was more pronounced in symphysis grafts than in ramus
grafts. Further studies are needed to investigate the mechanism of graft healing in
humans and factors that influence bone graft remodeling.

1

Chapter 1: Introduction and Background

Implant dentistry is a predictable treatment modality that has been used to replace
missing teeth. After the discovery of “osseointegration” with titanium in rabbit bone by
Per-Ingvar Branemark, the technique was rapidly adopted by the dental profession as the
standard of care for the replacement of missing teeth. Osseointegration has been defined
as, “direct structural and functional contact between ordered, living bone and the surface
of a load-carrying implant” (Branemark et al., 1977). Because this phenomenon is
dependent on bone-to-implant contact, successful implant therapy requires sufficient
bone volume to house and provide stability for the implant that is placed. The long-term
prognosis can be negatively affected by inadequate bone volume (Lekholm et al., 1986).
Clinicians often encounter cases where the patient does not have adequate bone
for implant therapy. Bone can be lost due to natural healing processes following tooth
extraction, periodontitis, endodontic lesions, trauma, oral pathology, surgical removal of
bone, etc (Carranza et al., 2006).
Implant therapy may require the removal of an existing tooth to create room for
implant placement. Several investigators have studied the influences of tooth extraction
on alveolar ridge resorption (Araujo & Lindhe, 2005; Atwood, 2001; Atwood & Coy,
1971; Carlsson, Bergman, & Hedegard, 1967; Johnson, 1969; Pietrokovski & Massler,
1967; Schropp et al., 2003; Tallgren, 1972). Most dimensional ridge alterations occur in
the first three months following exodontia (Johnson, 1969; Schropp et al., 2003). Most
ridge resorption occurs in a horizontal direction rather than a vertical direction, and

2

appears to be more pronounced in the facial aspect rather than the lingual aspect
(Pietrokovski & Massler, 1967; Schropp et al., 2003). Using a canine model, Araujo &
Lindhe (2005) explained that a thin layer of “bundle bone” is lost following tooth
extraction and is responsible for the collapse of the facial ridge, which is a significantly
thinner than the lingual.
Clinicians can choose from a variety of treatment modalities to augment hard
tissue in ridges with insufficient volume to house implants. These treatment options
include: autogenous onlay block graft, non-rigid guided bone regeneration (GBR), ridge
split, and titanium mesh (Block, Chang, & Crawford, 1996; Dahlin et al., 1988;
Kostopoulos & Karring, 1994; Maiorana et al., 2001; Nyman, 1991). In the literature,
autogenous bone is regarded as the “gold standard” for bone grafting due to its
osteoinductive and osteoconductive properties (Springfield, 1992). Osteoinduction has
been described as the process in which primitive, undifferentiated pluripotent cells are
stimulated to develop into bone-forming cell lineage (Albrektsson & Johansson, 2001;
Urist & Mc, 1952). Osteoconduction is the process by which grafting material serves as
a scaffold for new bone growth (Albrektsson & Johansson, 2001; Lindhe, Lang, &
Karring, 2008).

Clinical Studies Associated with Autogenous Bone Grafts: Literature Review
Implant success
Intra-oral mandibular autogenous onlay block grafting has become a predictable
treatment option to treat ridge defects (Aalam & Nowzari, 2007; Donos, Mardas, &

3

Chadha, 2008; Garg et al., 1998; Misch et al., 1992; Schwartz-Arad & Dori, 2002;
Tonetti & Hammerle, 2008). The mandibular ramus and symphysis are commonly
harvested intra-oral sites for block bone. Other extra-oral sources of autogenous bone
are: iliac crest, calvarium, and fibula. The donor site selection is dependent on several
factors including: available donor bone, proximity to adjacent teeth, anatomic
considerations, recipient defect size, and morphology.
Studies have supported that implants placed in ridges augmented with autogenous
bone have comparable success rates (90.9-100%) with implants placed in pristine bone
(Buser et al., 2002; Chiapasco et al., 1999; Cordaro, Amade, & Cordaro, 2002; Ozkan et
al., 2007; Sethi & Kaus, 2001; Verdugo et al., 2011). One study compared implant
success in block grafted sites with pristine sites. In a cohort study, Ozkan et al. (2007)
evaluated 15 patients where eight patients received 17 implants in symphysis block
grafted sites and seven patients received 18 implants in pristine sites. Success was
determined in a twelve-month follow-up by resonance frequency analysis (RFA) and
radiographic peri-implant bone levels. All implants in both groups were determined to be
clinically successful without a statistically significant difference in RFA values and
crestal bone loss.
Additional studies have evaluated implant success rates in autogenous grafted
sites without a control group (without graft). These studies have supported that
comparable success rates for block-grafted sites are achieved when compared with
studies that have independently evaluated implant success in pristine sites (Adell et al.,
1981, 1990; Albrektsson et al., 1988; Henry et al., 1996; Nevins & Langer, 1993).

4

There is a significant variation in grafting technique, sample size, implant success
criteria, and duration of follow-up in these studies. Buser et al. (1996, 2002) treated 40
patients (66 implants) using retromolar or symphysis block graft with autologous
particulate and expanded-polytetrafluroethylene (e-PTFE) membrane (Buser et al., 1996).
In a 60-month follow-up study, three patients (five implants, 7.6% of implants) dropped
out of the study. Criteria for implant success were: 1) absence of persistent subjective
complaints, 2) absence of peri-implant infection, 3) absence of mobility, and 4) absence
of continuous radiolucency around the implant. In the remaining 61 implants, one
implant failed due to peri-implant infection yielding a success rate of 98.3% (Buser et al.,
2002).
In a study by Chiapasco et al. (1999), 15 patients (44 implants) were grafted with
symphysis, iliac, or calvaria bone to treat ridge defects. In a mean follow-up time of 22.4
months, the success rate of the subsequent implants was 90.9% following Albrektsson’s
success criteria: 1) absence of mobility, 2) no signs of pain or suppuration, 3) direct
implant-to-bone contact was visible on radiographs, 4) >1 mm vertical bone resorption in
the first year after prosthetic loading (Albrektsson et al., 1986). Cordaro et al. (2002)
treated osseous defects with autogenous block grafts from the ramus or symphysis. All
implants were successful in a 12 month follow-up period based on Albrektsson’s success
critiera.
Sethi et al. (2001) evaluated 118 implants placed in 60 patients who had been
treated with autogenous block augmentation (ramus or symphysis) in a follow-up period
of 77 months (mean 21.7 months). Nine patients (15%) dropped out of the study, which

5

amounted to 29 implants (25%). During the follow-up period, two implant failures
occurred for a success rate (implant survival) of 98.3%. Verdugo et al. (2011) augmented
ridge defects with ramus or symphysis grafts on 15 consecutive patients and found a
100% implant success rate in a 40 month follow-up based on functional and asethetic
stability.

Bone gain
Clinical studies have demonstrated that autogenous onlay ridge augmentation is a
safe and predictable treatment option for ridge augmentation. These experiments indicate
that block grafts can significantly augment (3.2 - 4.6 mm) the ridge to provide adequate
dimensions for implant placement (Buser et al., 1996; Chiapasco et al., 1999; Cordaro et
al., 2002; Ozkan et al., 2007; Rabelo et al., 2010; Schwartz-Arad & Levin, 2005; Sethi &
Kaus, 2001; Verdugo et al., 2011; von Arx & Buser, 2006). There was a considerable
variation in bone graft source, consideration of graft resorption, addition of particulate
grafts, application of membranes, and follow-up period.
Ozkan et al (2007) treated horizontal ridge defects using symphysis grafts. Ridge
dimensions were augmented from 3.2 +/- 0.3 mm to 6.4 +/- 0.3 mm. Initial graft
thickness was measured at 3.8 +/- 0.3 mm, and calculated resorption values were 0.6 +/-
0.5 mm (15.8%) for a total gain of 3.2 mm. In a study by Chiapasco et al. (1999) using
bone from the symphysis, iliac crest, and calvaria, there was a bone gain of 4.0 +/- 0.82
mm (25.1% resorption). Cordaro et al. (2002) treated osseous defects with horizontal and
vertical ridge augmentation with autogenous block grafts from the ramus or symphysis.

6

Horizontal bone gain was 5.0 +/- 0.23 mm (23.5% resorption), and vertical bone gain was
2.2 +/- 0.66 mm (42% resorption).
A study by Verdugo et al. (2011) revealed significantly less bone resorption from
ramus and symphysis block grafted sites in a long-term follow up study. Pre- and post-
operative bone measurements were 3.3 +/- 0.7 mm and 7.4 +/- 1.2 mm (4.1 mm gain). A
40 month post-operative evaluation with cone beam computed tomography (CBCT)
revealed that 97% volume maintenance was maintained.
Some studies have combined autogenous block grafts with principles of guided
bone regeneration by introducing a barrier membrane. Buser et al. (1996, 2002)
combined the use of e-PTFE membrane with symphysis or retromolar block grafts. Mean
pre-operative ridges measured at 3.5 mm and mean bone gain was found to be 3.6 mm
(post-operative 7.1 mm). Another study by von Arx et al. (2006) combined symphysis or
retromolar block bone with anorganic bovine bone matrix particulate graft (Bio-Oss
®
,
Geistlich AG, Wolhusen, Switzerland) and collagen membrane (Bio-Gide
®
, Geistlich
AG). The alveolar ridge was augmented from mean widths of 3.06 mm to 7.66 mm (4.6
mm augmentation) with a mean 4.96 initial graft thickness (7.2% resorption).
The studies mentioned above were able to augment sufficient bone for implant
placement in all sites. On the other hand, Schwartz-Arad et al. (2005) harvested onlay
block grafts from the symphysis, ramus, iliac crest, and retromolar area for extensive
rehabilitation in 10 patients (87 implants). Two patients required additional grafting for
facilitation of implant placement. Rabelo et al. (2010) used block and/or particulated
autogenous bone from the symphysis, ramus, iliac crest, calvaria, and implant site

7

with/without PRP and evaluated whether sufficient bone for implant placement was
obtained (Rabelo et al., 2010). The study reported that 93.4% of 136 grafted sites had
sufficient bone for implant placement.

Review of Bone Healing Part I: Bone Fracture

It is essential to understand the healing mechanism associated with autogenous
block grafting when considering ridge augmentation procedures. The principles that
govern bone graft healing are largely similar to those that are responsible for bone
healing from trauma or fracture.
Osseous healing processes have been initially investigated in orthopedic literature
studying bone fractures (Axhausen, 1908; Barth, 1893). Studies have investigated the
time dependent healing mechanism in experimental fractures in animals and are
summarized below (Brighton & Hunt, 1986, 1991, 1997; Ham & Harris, 1971;
Rhinelander & Baragry, 1962; Urist & McLean, 1941). The healing process for bone
fracture wound healing can be separated into three phases: 1) Reactive phase, 2)
Reparative phase, and 3) Remodeling phase.

Phase I: Reactive phase
The reactive phase of bone fracture healing is characterized by a pronounced
inflammatory response and granulation tissue formation. Immediately following fracture,
an inflammatory response triggered by trauma and hemorrhage begins. Within the first

8

day, extravasated blood, hemorrhagic debris, inflammatory exudate and leukocytes
surround the injury site. By the third day, angiogenesis occurs and the blood clot begins
to be replaced by reparative granulation tissue to become the fibrous callus.

Phase II: Reparative phase
Cartilaginous components begin to appear within the callus between the fourth
day to third week to slowly develop into the fibrocartilaginous callus, which is made of
hyaline cartilage, fibrocartilage, and fibrous tissue. At this time, there is evidence of
intramembranous bone apposition from the periosteum and endosteum at some distance
away from the necrotic fracture zone. The ossification zone comes in contact with the
callus, which becomes the framework for future bone apposition to begin endochondral
ossification. This transition to a hard tissue callus occurs between the second to sixth
week. Simultaneous to the ossification of the callus, necrotic segments of the bone begin
to undergo resorption through macrophages, foreign-body giant cells, and osteoclastic
activity. Previously existing compact bone adjacent with the callus is also targeted by
osteoclastic activity to allow penetration of blood vessels and expansion of bone marrow.
Compact bone becomes as porous as cancellous bone. Bony union of the fracture is
accomplished subsequent to ossification of the fibrocartilage into woven bone.

Phase III: Remodeling phase
The hard tissue callus continues to remodel until it resembles the original bone
physiology with distinct cortical bone, cancellous bone, and marrow space. Osteoclastic

9

and osteoblastic activity is present until complete maturation is accomplished. This
process can take up to three to five years.

Review of Bone Healing Part II: Extraction Sockets

Authors who have investigated extraction socket healing have described many
similar observations with fracture healing. Many of these observations can be used to
draw paralleling similarities with the healing processes involved in autogenous block
grafts. The process of healing of extraction sockets can be summarized from the
experimental extraction histology studies (Amler, 1969; Araujo & Lindhe, 2005;
Cardaropoli, Araujo, & Lindhe, 2003; Evian et al., 1982).

Blood clotting and wound cleansing
During the first twenty-four hours of healing, a blood clot is formed within the
socket. A combination of platelets, fibrin network, and erythrocytes is developed to
inhibit bleeding and provide a matrix for subsequent healing processes. The blood clot
enhances inflammatory cell activity (neutrophils and macrophages) and provides growth
factors to stimulate mesenchymal cells.

Tissue formation
Two to three days following extraction, the clot undergoes fibrinolysis to be
replaced with granulation tissue, characterized by angiogenesis and new extracellular

10

matrix. Mesenchymal, fibroblast-like cells proliferate, release growth factors, and begin
secreting a provisional connective tissue. After one week of healing, the extraction
socket shows evidence of young connective tissue and primary osteoid formation in the
apical portion of the socket. The opening of the socket is covered by newly formed
epithelium, which migrates at approximately 0.5 - 1 mm/day (Engler, Ramfjord, &
Hiniker, 1966).
Multinucleated osteoclasts attach to the hard tissue borders of the extraction
socket to resorb bundle bone. Osteoprogenitor cells migrate along the newly formed
blood vessels, differentiate into osteoblasts, and secrete osteoid (woven bone). Such cells
will eventually become trapped by the osteoid and become osteocytes. Three weeks into
healing, connective tissue continues to mature meanwhile further mineralization of the
osteoid is apparent. After six weeks, a trabecular network of osseous apposition fills the
socket.

Tissue modeling and remodeling
Woven bone inside the extraction socket is gradually replaced by lamellar bone
and bone marrow. In the coronal aspect of the socket, a hard tissue cap forms, which
transitions from woven bone to lamellar bone and eventually to cortical bone by the sixth
month. The primary osteons are gradually replaced by secondary osteons with continued
osteoclastic and osteoblastic activity until remodeling is complete.

11

Review of Bone Healing Part III: Autogenous osseous transplantation

Several studies have investigated the histological healing process of osseous
transplantations or onlay grafts in animals (Adeyemo et al., 2008; Albrektsson, 1980b;
Andrade et al., 2010; Chen et al., 1994; de Carvalho, Vasconcellos, & Pi, 2000; Fonseca
et al., 1980, 1983; Maletta et al., 1983). There are considerable variations in animal
models, donor sources, recipient sites, and surgical techniques in these studies. General
trends regarding the complex healing process can be summarized from the combined
contributions of these studies. The healing process of osseous transplantation or onlay
block grafts is summarized below.

Phase I. Reactive phase
Autogenous block grafted sites are subject to similar reactive phases that occur in
bone fracture and extraction socket healing. Blood clot and inflammatory infiltrate that is
rich in erythrocytes, platelets, and leukocytes (neutrophils, lymphocytes, and
macrophages) are introduced in the grafted site. Albrektsson et al. (1980b) observed
considerable necrotic debris and edema within transplanted bone segments until the fifth
day of healing. Fonseca et al. (1980) found significant inflammatory infiltrate and edema
limited to the area immediately surrounding the graft.
At this point, there is little evidence of angiogenesis within the graft material from
the soft tissues and underlying host bone (Albrektsson, 1980b; Chen et al., 1994; Kusiak,
Zins, & Whitaker, 1985). However, there is evidence of hyper-vascular response in the

12

soft tissue surrounding the avascular graft. The soft tissue surrounding the periphery of
the blood clot begins to transition into granulation tissue by the third day.

Phase II. Reparative phase
Considerable amounts of inflammatory infiltrate with significant edema are still
present by the first week of healing. There is a significant decrease in number of viable
osteocytes within the graft lacunae. Occasional osteocytes within the lacunae of the graft
may still be present at this point of the healing. During the early reparative phase, a rim
of osteoblasts and osteoclasts can be seen lining the grafts and cortex of the mandible.
Osteoclastic activity is most active where the graft approximates with the cortex of the
recipient site. Although minimal, most initial revascularization within the grafts appears
to originate from the surrounding soft tissues (Chen et al., 1994; Fonseca et al., 1980;
Kusiak et al., 1985). The most significant angiogenesis has only occurred along the
periphery of the graft facing the mandibular cortex. Minimal revascularization can be
seen within the buccal cortex of the recipient site.
The process of revascularization, osteoclastic, and osteoblastic activity continues
as inflammation subsides during the following weeks. During these reparative processes,
bone apposition generally tends to follow osteoclastic resorptive activities (Enneking et
al., 1975). By the second to fourth week, the mandibular cortex demonstrates evidence of
transitioning to trabecular bone along its borders. Pre-existing lacunae become void of
viable osteocytes. The intertrabecular spaces are widening, and trabecular bone is thin in
appearance and lined by osteoblasts. Vascular proliferation may be seen arising from the

13

medullary portion of the alveolar process and penetrating through the buccal cortex and
into the block bone. Considerable more revascularization has established within the
graft, although avascular regions may remain.
Tissues surrounding the graft begin to develop into osteoid-like tissues or osseous
callus to incorporate the block into the recipient site. Osteogenesis surrounding the
autogenous onlay bone grafts is observed in the 1) lateral, 2) inferior, and 3) superior
regions to the graft (Andrade et al., 2010). The majority of new bone is formed in the
lateral region and between the cortex of the graft and host mandible, which allows graft
consolidation. Bony trabeculation develops superior to the graft due to osteoinductive
properties from the periosteum (Adeyemo et al., 2008; Andrade et al., 2010; Fonseca et
al., 1980). The graft matrix gradually resorbs to become replaced by new bone turnover.
Graft turnover is slower in dense cortical regions than cancellous regions of the block
bone graft (Albrektsson, 1980b; Deleu & Trueta, 1965; Phillips & Rahn, 1990; Sullivan
& Szwajkun, 1991). There are conflicting views on whether the bone graft becomes
completely revascularized and replaced with new vital bone.
During the second to fourth month of healing, a new buccal cortical plate with
immature lamellar pattern and irregular trabeculae begins to establish. If complete
angiogenesis and graft turnover has occurred, the recipient site buccal cortex and graft
remodel to lamellar bone to the extent that it may become difficult to distinguish their
original borders. Osteoblastic and osteoclastic remodeling is still present with reduced
activity. If incomplete revascularization has occurred, avascular components of the block
graft may be present with lacunae void of cellular activity (Albrektsson, 1980b; Andrade

14

et al., 2010; Enneking et al., 1975; Fonseca et al., 1980; Kusiak et al., 1985; Proussaefs,
Lozada, & Rohrer, 2002). Significant reduction in graft volume and rounding of the
peripheral edges may be observed.

Phase III. Remodeling phase
A second phase of osteogenesis occurs in the healing process of a transplanted
osseous segment (Albrektsson et al., 1980b). During this phase, bone continues to
remodel by developing hard tissue with concentric laminae. Lamellar bone is replaced by
cortical bone, cancellous bone, and marrow spaces that consist of loose connective tissue
and possibly adipose cells.
By the sixth month, overlying soft tissue completely matures without evidence of
inflammatory activity. However, the new bone still consists of poorly organized and
immature lamellar bone, indicating that remodeling is still incomplete. Howship’s
lacunae and osteoclasts may still be present along the buccal surfaces of the graft.
Occasional osteoblasts and osteoclasts may be seen indicating continued remodeling of
the bone. Remodeling continues until bone maturation is complete with distinct cortical
and cancellous layers are present.
If necrotic regions of bone are present at this point, it is unclear whether the
avascular components will remain or eventually remodel long term. Additionally, it is
unknown whether the presence of avascular portions of bone results in any negative
clinical affects.

15

Devitalization, cellular death, apoptosis
Several authors have shown that autogenous blocks lose their vascular supply
after surgical transplantation into the recipient site (Chen et al., 1994; Kusiak et al., 1985;
Phillips & Rahn, 1990; Sullivan & Szwajkun, 1991). Kusiak et al. (1985) used silicone
rubber techniques in New Zealand rabbits and observed that grafts were avascular or
minimally vascularized after 1, 3, and 7 days of healing and highly revascularized after
14 or 21 days in rabbits. Chen et al. (1994) and Sullivan & Szwajkun (1991) found
similar findings in rabbits and rats respectively. In a sheep model study, Phillips & Rahn
(1990) observed poor vascularization after 2 weeks and significant vascular infiltration
after 20 weeks.
During the healing process, osseous grafts undergo necrosis and osteocytes within
lacunae degenerate. Cellular viability is mostly lost during the first 24 hours and may be
completely lost by the third day (Kenzora et al., 1978). On the other hand, osteocyte lysis
occurs to a greater extent after several weeks, which can be seen histologically as empty
lacunae. Studies indicate that osteocytes associated with the autogenous block bone are
lost by the second to fourth week of healing (Fonseca et al., 1980; Maletta et al., 1983;
Thompson & Casson, 1970). Therefore, graft turnover through revascularization, repair,
and remodeling is crucial to replace necrotic bone following autogenous block grafting.

Graft turnover
Angiogenesis is essential in the repairing and remodeling of autogenous osseous
transplants (Albrektsson, 1980a, 1980b; Albrektsson & Albrektsson, 1978). Initially, the

16

block graft receives most of its blood supply from the surrounding soft tissues, especially
along the periphery of the graft facing the mandibular cortex (Fonseca et al., 1980).
Kusiak et al. (1985) and Fonseca et al. (1980) demonstrated that significant graft
revascularization has occurred by the 2nd to 4th week post-operative healing, whereas
Chen et al. (1994) demonstrated significant revascularization at only 10 days. At this
time, vascular proliferation can be seen penetrating from the medullary portion of the
recipient site through the cortex and into the graft bone.
Albrektsson et al. (1980b) estimated that vascular ingrowth rate within the graft
was 0.2-0.4 mm/day for cancellous bone and 0.15-0.3 mm for cortical bone. The degree
of bone turnover was directly related with the amount of vascularization around vessel
sizes of 25-60 microns. Deleu & Trueta (1965) assessed revascularization of bone grafts
in the anterior chamber of the eye in albino guinea pigs and found that revascularization
began/completed in cancellous bone and cortical bone at 2 days/2 weeks and 6 days/1-2
months respectively. Cancellous bone was associated with rapid vascularization, whereas
cortical bone was associated with slower and sometimes incomplete revascularization. In
cortical bone, vessel penetration occurs through enlargement of pre-existing cavities and
Haversian canals or through the introduction of new resorption cavities. Once significant
revascularization has occurred, bone resorption and apposition occur with the adjacent
blood supply.
Several authors described the possibility that osseous transplants may not become
completely revascularized and leave remnants of necrotic bone (Albrektsson, 1980b;
Enneking et al., 1975; Maletta et al., 1983; Shirota et al., 1996; Urbani et al., 1998).

17

Albrektsson et al. (1980b) demonstrated that on occasion, osseous transplants in rabbit
tibia were not fully revascularized, which was accompanied by minimal bone
remodeling. This may leave segments of necrotic bone without adequate blood supply
and with lacunae void of osteocytes.
Several studies have demonstrated that block grafts are completely revascularized
and turned over into new bone with osteocytes within lacunae (Adeyemo et al., 2008; de
Carvalho et al., 2000; Fonseca et al., 1980; Phillips & Rahn, 1990; Proussaefs et al.,
2002). Fonseca et al. (1980) studied the healing of onlay cortico-cancellous iliac bone
chips (2x2x2mm and 2x2x5mm) on the mandible in the monkey model and demonstrated
that complete graft turnover was accomplished. The bone grafts became
indistinguishable from the alveolar ridge at 3 months of healing. Adeyemo et al. (2008)
found similar findings in cortico-cancellous iliac onlay grafts (10x20x7.5mm) on sheep
mandible. Mature bone completely replaced the bone graft in 16 weeks. Phillips & Rahn
(1990) studied the revascularization of parietal and rib onlay grafts (30x10x4mm) on
sheep mandible. After 20 weeks of healing, the onlay grafts demonstrated complete
revascularization regardless of graft type.
In a study by de Carvalho et al. (2000), the healing of posterior mandibular
cortico-cancellous bone grafts was investigated using the mandible in a dog model. The
grafts were placed on three different types of recipient beds: cortical, perforated, and
decorticated. The study showed that the graft was completely incorporated,
revascularized, and turned over in the perforated and decorticated groups after 90 days.

18

In the cortical recipient bed, the graft demonstrated deficient revascularization,
insufficient remodeling, and remnants of non-vital graft bone.
Using a point-counting technique, Zins et al. (1983) evaluated the percentage of
total bone and necrotic graft bone remaining in 15x5x3 mm iliac crest and calvaria grafts
on the cranium of monkeys after 12 weeks. Both graft types exhibited similar total bone
values for endochondral and intramembranous bone: 54.6 +/- 4.2% and 54.6 +/- 4.8%
respectively. Both graft types demonstrated significant regions of non-vital bone within
the grafts: 26.5 +/- 3.6% and 14.1 +/- 2.2% respectively.
In a human histology study of autogenous onlay bone graft, Proussaefs et al.
(2002) demonstrated the remodeling capabilities of a ramus onlay graft in a case report.
A biopsy was sampled with a 2 mm internal-diameter trephine bur at one year after bone
grafting. Histologic results demonstrated that the autogenous ramus onlay block graft
could re-establish its vitality and undergo remodeling. This study supports that complete
remodeling and revascularization is possible in autogenous block grafting in humans.
In another human histology case series, Urbani et al. (1998) collected biopsy
samples in a bucco-lingual direction after 6 months following mandibular symphysis
block grafting. Recipient beds were perforated at the time of grafting surgery. The
histological results of the study showed distinct cellular and acellular regions of bone that
were fused together. The acellular portion consisted of empty lacunae surrounded by
cortical bone that was normal in appearance. This study suggests the possibility of
leaving un-remodeled necrotic remnants of bone after autogenous block grafting in
humans.

19

Shirota et al. (1996) reconstructed mandibular defects with non-vascularized iliac
bone grafts following tumor removal. After 6-12 months of healing, histologic samples
were collected with a 3.0 mm trephine bur from the graft bone during implant surgery.
Sections revealed remaining areas of devitalized graft bone, which was characterized by
empty lacunae, adjacent with newly formed trabecular bone.
Additionally, various orthognathic surgical procedures such as segmental
osteotomy involve complete separation of dento-osseous segments and fixation to a new
location. Similar to autogenous block grafts, vascular is supply is interrupted and must
re-establish itself for proper incorporation. Several studies have indicated the osseous
portion of the transplant becomes well incorporated and revascularized within the host
site, despite the risk of repositioned teeth being subject to pulp necrosis, root resorption,
ankylosis, and/or exfoliation (Bell, 1969; Bell & Levy, 1971; Bell, Schendel, & Finn,
1978; Holland & Robinson, 1986; Luke & Boyne, 1968). Bonding reactive new bone
formation occurs between the transplant and host site through an osseous callus by the
sixth to eighth week. Remodeling and bone apposition of the osseous component of the
segment can be seen in the marrow vascular spaces at this time.
Enneking et al (1975) evaluated bone healing using an inverted segmental fibula
transplant and sham control in the dog model throughout 48 weeks after surgery.
Additionally, the transplanted segments were subject to physical loading to determine its
fracture torque and maximum stress values. In some cases, various regions did not
exhibit remodeling and had persistent regions necrotic bone. There was an interlocking
jigsaw fit between necrotic and viable cortical bone. Factors for premature bone

20

remodeling cessation could not be determined. However, no inherent weakness could be
found associated with non-vital cortical bone inload-bearing capacities. The presence of
necrotic regions of bone may not have deleterious effects in mechanical function, unless
it subject to future resorption or development of porosities.
Another study by Maletta et al. (1983) showed that necrotic remnants of bone
may be subject to resorption and subsequent volume loss. In this experiment, autogenous
rib onlay grafts were placed on the edentulous maxilla of monkeys and were followed for
8 months of healing. Significant amounts of necrotic bone with acellular elements in the
lacunae were present at six months. By the eighth month, significant resorption of
necrotic bone had occurred to reduce the volume of the augmented alveolar ridge.

Purpose of the study
There are considerable variations in the previously mentioned study designs
including: source of bone graft, follow-up period, recipient site location, recipient site
preparation, animal species, etc. The resulting histologic vitality of autogenous onlay
bone grafts is inconclusive based on these studies. Currently, the histological outcome
and vitality of mandibular onlay grafts in humans is not well documented. The purpose of
this case series is to assess the histology and vitality of the bone graft in patients who
have been treated with autogenous mandibular onlay ridge augmentation.

21

Chapter 2: Materials and Methods

Four patients were consecutively selected in the Advanced Periodontology Clinic
at Ostrow school of Dentistry of USC for dental implant placement following significant
ridge reconstruction with autogenous mandibular onlay grafts. This research project was
approved by the University of Southern California Institutional Review Board (IRB #:
Study - HS-11-00194). All patients reviewed and signed informed consent and HIPAA
documents prior to enrolling in the study. The patients received comprehensive dental
evaluation and medical histories were reviewed. Patients were excluded if the bone graft
did not represent a significant portion of the reconstructed ridge. The implant recipient
site was evaluated with cone beam computed tomography (CBCT).
The autogenous cortico-cancellous block grafts were harvested from either the
mandibular ramus or symphysis based on available donor bone. A full-thickness flap was
elevated to gain access to the donor bone, and the block graft was harvested using
surgical round burs and piezotomes. The host cortex was perforated with multiple
bleeding points. The recipient sites and autogenous blocks were shaped to allow
adaptation between host and graft bone (Figures 1-3). The block graft was fixated with
titanium screws (truFIX
TM
Complete Fixation System).

22

Figure 1. Recipient site defect

Figure 2. Donor bone harvest

23

Figure 3. Graft fixated in place

At least four months following ridge augmentation, patients were treated with
implant surgery. At the time of implant surgery, a trephine bur (2 mm inner diameter, 3
mm outer diameter; 3i Biomet) was used to harvest a bone core that was approximately
3-4 mm long (Figures 4, 5). A non-invasive approach to harvesting block bone cores
were used by taking the sample from implant osteotomy sites with trephine burs as
described in other studies in humans (Artzi, Tal, & Dayan, 2000; Cammack et al., 2005;
Lee et al., 2009; Minichetti et al., 2004). Discarded bone cores taken from the site were
immediately fixated in 10% neutral buffered formalin (NBF, Figure 6). The implant
osteotomy was prepared with twist drills and countersink drills as necessary to facilitate
implant placement.

24

Figure 4. Healed block graft

Figure 5. Bone core harvesting

25

Figure 6. Bone core in 10% NBF

The bone core samples were prepared for histological analysis by the Histology
Core Laboratory, University of Southern California. The bone cores were allowed to
become fixated in 10% NBF for one day and were decalcified in 10% formic acid for 5
days. Afterwards, the samples were dehydrated in 70% (1 hour), 80% (1 hour), 95% (1
hour), 100% (1 hour x 2) ethanol and 100% (1 hour x 2) xylene, and embedded in 100%
paraffin (1 hour x 2) using the Microm Spin Tissue Processor STP 120 (Fisher
Scientific). The paraffin infiltrated tissues were embedded in wax and mounted on
cassettes using the Microm Tissue Embedding Center EC 350 (Fisher Scientific). The
samples were cut into thin 5 μm sections using the Microm Rotary Microtome (Fisher
Scientific) and were stained with hematoxylin and eosin stain (H&E stain) using the
Thermo Scientific Varistain® Gemini ES (Fisher Scientific). The histological samples

26

were evaluated for grafted bone vitality by assessing the presence of osteocytes inside
lacunae, re-organization of haversian systems, angiogenesis, and osteoblastic/osteoclastic
activity. Digital images of the histological slides were obtained using the Olympus
SZX12 and Zeiss AX10 microscopes.

27

Chapter 3: Results

Clinical Observations
The clinical synopsis for the four treated defects is summarized in Table 1 (mean
age 37.3 years; 2:2 males and females). One patient (case #3) was a smoker (<5
cigarettes/ day) who abstained from smoking for one month after surgery. The grafted
sites included two maxillary central incisors, one maxillary lateral incisor, and one
maxillary first premolar. Two defects were treated with bone from the mandibular
symphysis, whereas two defects were treated with bone from the mandibular ramus. All
surgeries healed uneventfully for bone grafting (healing time of 4, 5, and 7 months) and
implant placement. All bone grafts were clinically integrated with the host bone at the
time of trephining and implant placement. All trephined sites exhibited bleeding and all
implants were stabilized at 50 NCm.

28

Table 1. Clinical synopsis
Implant Primary
Stability
Yes
Yes
Yes
Yes
Bleeding Upon
Trephination
Yes
Yes
Yes
Yes
Graft
Healing Site
4 months
4 months
5 months
7 months
Donor
Symphysis
Ramus
Symphysis
Ramus
Recipient
Site
#8
#10
#12
#9
Medical
History
Non-
contributory
Non-
contributory
Smoker (<5
cigs/day)
Non-
contributory
Gender
Female
Female
Male
Male
Age
(Years)
48
30
42
29
Patient
1
2
3
4

29

Histologic Observations
The histological samples from this case series demonstrated varying degrees of
diffused angiogenesis and cellular activity within lacunae. Angiogenesis and marrow
spaces were more pronounced in mandibular symphysis grafts than ramus grafts. A dense
cortical bone was the predominant feature of the ramus grafts. Symphysis grafts
demonstrated more osseous turnover than ramus grafts which was confirmed by the
presence of osteocytes within lacunae and location of the demarcation lines. Cellular
activity within lacunae was found adjacent to blood vessels and marrow space.

Case 1
The presence of vital new bone were differentiated from unremodeled graft bone by
the presence of demarcation lines and osteocytes within the lacunae (Figures 7-12).
Histologic assessment revealed vital bone (characterized with osteocytes within lacunae)
amalgated with unremodeled graft bone (characterized by empty lacunae). Demarcation
lines were visualized which marked the borders between vital and unremodeled bone.
The vital bone consisted of lamellar bone with distinct layers and disorganized woven
bone. Vital bone was found adjacent with the blood supply from Haversian canals and
connective tissue. Endothelial cells, erythrocytes, fibroblasts, lymphocytes, and woven-
bone like calcified structures were visible embedded within the connective tissue. Lining
osteoblasts were found between the segments of vital bone and connective tissue. In few
regions, demarcation lines could not be distinguished bordering vital bone and necrotic
bone near smaller vessels (Figure 8).

30

Figure 7. Case 1(a): histologic overview

Figure 8. Case 1(b): 20x magnification

31

Figure 9. Case 1(c): 20x magnification

Figure 10. Case 1(d): 20x magnification

32

Figure 11. Case 1(e): 40x magnification

Figure 12. Case 1(f): 40x magnification

33

Case 2
Histologic assessment revealed dense cortical bone as the predominant feature of
the sample. Most of the bone appeared to be non-vital without cellular activity within the
lacunae. Islands of vital bone adjacent with the blood supply were observed throughout
the necrotic graft bone (Figure 13-15). The vital bone was clearly distinct from the
necrotic bone with demarcation lines, presence of osteocytes within lacunae, and the
disorganized histological appearance (Figures 16-18). The vital bone demonstrated
different stages of bone maturation including disorganized woven bone and maturing
woven bone with developing layers. Regions of connective tissue with lining osteoblasts,
fibroblasts, erythrocytes, and woven-bone like calcified tissues were found surrounded by
the new bone. One area of vital bone did not surround blood vessels or connective tissue
in this section (Figure 19). Additionally, one region of connective tissue was not
surrounded by newly formed vital bone (Figure 20).

34

Figure 13. Case 2(a): histologic overview

Figure 14. Case 2(b): 10x magnification

35

Figure 15. Case 2(c): 10x magnification

Figure 16. Case 2(d): 20x magnification

36

Figure 17. Case 2(e): 20x magnification

Figure 18. Case 2(f): 20x magnification

37

Figure 19. Case 2(g): 40x magnification

Figure 20. Case 2(h): 40x magnification

38

Case 3
The histological sample had less total bone area and was dominated by regions of
connective tissue and adipose tissue (Figure 21). The section consisted of unremodeled
graft bone and new vital bone, which were differentiated by the presence of osteocytes
within lacunae, organization of bone, and demarcation lines (Figures 22-24). Cancellous
bone-like mineralized substances were found embedded within the connective tissues of
the sample. Osteoblasts were detected lining the newly formed bone and cancellous
bone-like tissues. One region of the core demonstrated a great number of osteoblast-like
cells with large nuclei along surfaces of newly formed disorganized woven bone (Figures
25-27). A large number of fibroblasts was detected in the connective tissue adjacent with
this area.

Figure 21. Case 3(a): histologic overview

39

Figure 22. Case 3(b): 20x magnification

Figure 23. Case 3(c): 20x magnification

40

Figure 24. Case 3(d): 40x magnification

Figure 25. Case 3(e): 20x magnification

41

Figure 26. Case 3(f): 40x magnification

Figure 27. Case 3(g): 40x magnification

42

Case 4
Localized tears from the histologic samples were detected after sectioning with
the microtome. Ultimately, the sample was lost during the process of obtaining
additional sections. The available histologic slides were assessed at 40x magnification.
Avascular cortical bone with empty lacunae was the predominant feature of this
histologic specimen. The outlines of osteons around Haversian canals were detected
throughout the sample. There was evidence of vital bone with cellular activity within
lacunae surrounding the Haversian canals and islands of connective tissue (Figures 28-
29). Occasional lining osteoblasts were detected along the border between connective
tissue and bone. Fibroblasts and cancellous bone-like mineralized tissue were observed
embedded in the connective tissue. Few of the Haversian canals demonstrated limited
surrounding vital bone (Figures 30).

43

Figure 28. Case 4(a): 40x magnification

Figure 29. Case 4(b): 40x magnification

44

Figure 30. Case 4(c): 40x magnification

45

Chapter 4: Discussion

This human histologic study demonstrates the potential for angiogenesis and graft
turnover in autogenous bone grafting. This was confirmed by bleeding at the time of
sample collection, detection of angiogenesis, and diffuse cellular activity. These results
support the findings from animal models that have shown the capability of autogenous
bone grafts to become vital bone (Adeyemo et al., 2008; Albrektsson, 1980b; Andrade et
al., 2010; Chen et al., 1994; de Carvalho et al., 2000; Fonseca et al., 1980; Maletta et al.,
1983).
The histologic vitality of autogenous bone grafts in humans is not well
documented. In the first human histology study, Shirota et al. (1996) evaluated non-
vascularized autogenous iliac grafts that were used to reconstruct the segmental defects in
the mandible (Shirota et al., 1996). Histologic specimens collected at 6 and 12 months
revealed minimal vital bone in the grafted bone. These results reveal the histologic
outcome of endochondral grafts and not intramembranous grafts. Kusiak et al. (1985)
and Zins et al. (1983) demonstrated in the rabbit model that endochondral bone grafts
exhibit less revascularization, graft turnover, and volume maintenance than
intramembranous bone grafts (Kusiak et al., 1985; Zins & Whitaker, 1983).
The current study was the first to assess histology of mandibular autogenous bone
(ramus and symphysis) alone in humans. Cases published by Proussaefs et al. (2002) and
Urbani et al. (1998) evaluated ramus bone in conjunction with inorganic bovine mineral
(Bio-Oss
®
) and symphysis grafts with absorbable pins respectively. The histologic

46

specimen showed areas of vital bone in contact with non-vital bone, which were
differentiated by the presence of cellular activity within the lacunae and shade of the
bone.

Angiogenesis and osteogenesis
Autogenous grafts lose their vascular supply after transplantation into the
recipient site, and depend on revascularization for revitalization of the grafts (Chen et al.,
1994; Kusiak et al., 1985; Phillips & Rahn, 1990; Sullivan & Szwajkun, 1991).
Albrektsson et al. (1980b) showed through vital microscopy in rabbits that vascular
ingrowth rates within the osseous transplants were 0.15-0.4 mm/day. Bone turnover was
observed around new vessels that were 25-60 microns wide.
This study highlights the importance of angiogenesis in bone graft turnover,
because vital bone was observed adjacent to blood supply. A “halo” of newly formed
vital bone often surrounded sources of blood supply. Only one region in the third sample
appeared to be independent of adjacent blood supply, which is likely because the two-
dimensional section did not properly capture nearby vessels from the three dimensional
core.
Osteogenesis and cellular activity within lacunae may be the result of: 1) cells that
survive the surgery, 2) osteoconduction, and 3) osteoinduction (Khoury, Antoun, &
Missika, 2007). On 1863, Wolff suggested that osteoblasts survive osseous
transplantation and contribute to osteogenesis. However on 1893, Barth observed in a
microscope study that surviving graft cellular activity is insufficient to explain

47

osteogenesis and graft turnover. Moreover, osseous transplants devitalize and serve as a
mineralized matrix that is used as a structural scaffold for osteogenesis. In addition,
osteoinduction through stimulation of undifferentiated pluripotent cells into bone-forming
cell lineage can enhance osteogenesis. Autogenous grafts are considered the “gold
standard” of bone grafts due to their osteoconductive and osteoinductive properties.
In this study, the presence of demarcation lines, organization of bone, and
presence of osteocytes within the lacunae differentiated vital bone and graft bone. The
immature structural organization of vital bone from the samples suggests that cellular
activity within lacunae of block graft sites is the result of newly formed bone and new
osteocytes, rather than graft cellular survival. In animal studies, osteocyte viability is
mostly lost after transplantation by the first 1-3 days, while most cellular apoptosis has
occurred by the 2nd-4th week (Fonseca et al., 1980; Kenzora et al., 1978; Maletta et al.,
1983; Thompson & Casson, 1970).
Few regions of vital bone from the first and fourth samples did not have clear
borders between the necrotic bone and vital bone. In addition with the absence of an
obvious demarcation line, the structural organization and staining color of this vital
lamellar bone could not be easily differentiated from the necrotic graft bone. Therefore,
it is difficult to determine whether these regions with osteocytes within lacunae are the
result of cellular survival or osteogenesis. Application of additional histological
techniques such as Masson’s Trichrome stain may help differentiate between new and old
bone.

48

Several authors have described the possibility that osseous transplants may
become completely revascularized and turned-over with new bone (Adeyemo et al.,
2008; de Carvalho et al., 2000; Fonseca et al., 1980; Phillips & Rahn, 1990; Proussaefs et
al., 2002). Other authors have demonstrated that osseous transplants may not become
completely revascularized and leave remnants of necrotic bone (Albrektsson, 1980b;
Enneking et al., 1975; Maletta et al., 1983; Shirota et al., 1996; Urbani et al., 1998). In
this study, all specimens demonstrated the potential for angiogenesis and graft turnover.
However, varying amounts of necrotic graft bone and vital osseous tissue were present.
Newly formed vital bone was well-amalgated into the necrotic graft bone.
The presence of lining osteoblasts in all samples confirms that bone remodeling is
an ongoing process at sample collection. This finding is in agreement with published
animal studies (Adeyemo et al., 2008; Andrade et al., 2010; de Carvalho et al., 2000;
Fonseca et al., 1980). In the third sample, significant osseous remodeling was observed
with a great number of osteoblasts with large nuclei. Osteoclasts could not be
histologically differentiated from osteoblast-like-cells, which may encourage future
investigations to include additional histologic stains such as tartrate resistant acid
phosphatase (TRAP) staining in the histologic protocol to better identify osteoclastic
activity.
The healing of bone grafting is a complex and dynamic process that is subject to
different phases of healing: reparative, modeling, and remodeling phase. In the present
study, histologic samples were collected at 4, 5, and 7 months which may represent
healing between the modeling and remodeling phase. All histologic samples showed

49

varying amounts of osteogenesis and remaining necrotic bone. At this point, osseous
healing is incomplete and the ultimate histologic outcome of the bone graft is unknown.
Enneking et al. (1975) suggested that remaining necrotic bone persists and does not have
deleterious effects on physical load of inverted fibula studies in dogs. Maletta et al.
(1983) demonstrated the possibility for necrotic bone to resorb after eight months of
healing in monkeys. Based on currently available literature, it is inconclusive whether
the necrotic bone at sample collection remains or remodels into vital osseous tissue.
In this case series, ramus bone grafts exhibited less revascularization and osseous
turnover than symphysis bone. This may be explained by the morphological quality of
the donor bone. Ramus bone is predominantly cortical bone, whereas symphysis bone is
characterized as cortico-cancellous bone. The present study is in agreement with animal
studies that have shown that cortical bone grafts exhibit slower revascularization than
cancellous grafts (Albrektsson, 1980b; Deleu & Trueta, 1965; Phillips & Rahn, 1990;
Sullivan & Szwajkun, 1991). Phillips & Rahn (1990) observed that grafts with more
cancellous components revascularized faster than cortical grafts in Sprague Dawley rats.
Additionally, Sullivan & Szwajkun (1991) observed similar findings in the sheep model.

Limitations
The present examination is a pilot for future studies that will investigate the
mechanism of autogenous bone graft healing in humans. Larger sample sizes and cores
collected in longer healing periods could reveal additional information regarding the graft
remodeling process. Additional laboratory techniques such as micro-computed

50

tomography (micro CT) and staining techniques such as Masson’s Trichrome and TRAP
staining could provide information on bone density, differentiation of new and old bone,
and identification of osteoclasts.
The interface between the bone graft and recipient site is not currently
documented in humans. In the methodology of the present study, bone cores were
harvested from implant osteotomy sites, which cannot guarantee capturing the graft-host
interface. Bone grafts must consolidate with the recipient site during the healing process
with osteogenesis located: superior to the graft, lateral to the graft, between the graft and
recipient site, and within the graft (Andrade et al., 2010). In a monkey model study,
Fonseca et al. (1980) observed that autogenous bone grafts have completely integrated
and become histologically indistinguishable with the recipient bone after 4-6 months of
healing. In humans, further insight could be gained if the bone cores were collected from
the buccal aspect of the graft and penetrated into the recipient bone to capture this
interface.

51

Chapter 5: Conclusion

Within the limits of the first human histology study using ramus or symphysis
graft alone, the following conclusions could be drawn. Varying degrees of angiogenesis
and osteogenesis were present in all samples. Vital bone is largely a result of
osteogenesis and graft turnover rather than cellular survival. Remodeling is ongoing at 4,
5, 7 months and was more pronounced in symphysis grafts than ramus grafts. Further
studies will investigate the mechanism of graft healing in humans and factors that
influence bone graft remodeling.

52

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Abstract (if available)

Abstract Background: Autogenous block bone grafting is a treatment option for ridge deformities. Currently, the healing process of autogenous bone grafting in humans has not been histologically reported and the vitality of bone after graft placement has been challenged. ❧ Objective: The purpose of this study is to histologically evaluate vitality of autogenous bone grafts in patients who have been treated for ridge deformities in preparation for implant placement. ❧ Materials and Methods: At 4, 5, and 7 months after bone augmentation and at the time of implant placement, a core of 2x3 mm bone graft was trephined for histological analysis (n=4

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Asset Metadata

Creator Kim, Andrew (author)

Core Title Human histological analysis of autogenous block grafts

School School of Dentistry

Degree Master of Science

Degree Program Craniofacial Biology

Publication Date 09/01/2012

Defense Date 07/17/2012

Publisher University of Southern California (original), University of Southern California. Libraries (digital)

Tag autogenous bone,Histology,OAI-PMH Harvest,ramus,symphysis

Language English

Contributor Electronically uploaded by the author (provenance)

Advisor Zadeh, Homayoun H. (committee chair), Navazesh, Mahvash (committee member), Rich, Sandra (committee member)

Creator Email andykdds@gmail.com,kimandre@usc.edu

Permanent Link (DOI) https://doi.org/10.25549/usctheses-c3-93776

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Legacy Identifier etd-KimAndrew-1179.pdf

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Type texts

Source University of Southern California (contributing entity), University of Southern California Dissertations and Theses (collection)

Access Conditions The author retains rights to his/her dissertation, thesis or other graduate work according to U.S. copyright law. Electronic access is being provided by the USC Libraries in agreement with the a...

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