Abstract
Background: There is great interest in the use of bone substitutes
to improve bone repair. We compared the osteogenic potential of lyophilized
bone chips combined with platelet gel, or with platelet gel and bone marrow
stromal cells, with that of lyophilized bone chips alone in the healing of a
high tibial osteotomy.
Methods: A prospective, randomized, controlled study was performed,
and a standardized clinical model was applied. Thirty-three patients
undergoing high tibial osteotomy to treat genu varum were enrolled and
assigned to three groups. During the osteotomy, lyophilized bone chips with
platelet gel were implanted into eleven patients (Group A), lyophilized bone
chips with platelet gel and bone marrow stromal cells were implanted in twelve
patients (Group B), and lyophilized bone chips without gel were placed in ten
patients as controls (Group C). Six weeks after surgery, computed
tomography-guided biopsies of the grafted areas were performed and the
specimens were analyzed by histomorphometry. Clinical and radiographic
evaluation was performed at six weeks, twelve weeks, six months, and one year
after surgery.
Results: Histomorphometry at six weeks showed significantly
increased osteoblasts and osteoid areas in both Group A (p = 0.006 and p =
0.03, respectively) and Group B (p = 0.009 and p = 0.001) in comparison with
controls, as well as increased bone apposition on the chips (p = 0.007 and p =
0.001, respectively), which was greater in Group B than in Group A (p <
0.05). Group B showed significantly higher revascularization than the controls
(p = 0.004). Radiographs revealed a significantly higher rate of
osseointegration in Groups A and B than in the controls at six weeks (p <
0.005 and p < 0.0001, respectively). At the final evaluation at one year,
the osseointegration was still better in Groups A and B than in Group C;
however, all patients had complete clinical and functional evidence of
healing.
Conclusions: Adding a platelet gel or a platelet gel combined with
bone marrow stromal cells to lyophilized bone chips increases the osteogenetic
potential of the lyophilized bone chips and may be a useful tool in the
treatment of patients with massive bone loss.
Level of Evidence: Therapeutic Level I. See Instructions
to Authors for a complete description of levels of evidence.
In recent years, much work has been done in order to obtain biocompatible
substitutes to improve bone
healing1-4.
Autologous cancellous bone graft is the most effective grafting material, but
the limited quantity of bone available for harvesting has led to the use of
alternatives. Freeze-dried bone allograft has decreased antigenicity and does
not induce biochemical changes; however, it has a reduced osteogenic
potential5.
Grafting procedures involving the use of growth factors, platelet-rich
plasma, and platelet-derived growth
factors6-12,
as well as platelet gel obtained by adding calcified thrombin to platelet-rich
plasma6,8,13,14,
have been studied and have shown promising results.
Platelet-rich plasma added to a bone graft has been reported to
substantially accelerate the bone repair process, particularly in
fracture-healing and bone implant
fixation15.
However, some studies have contradicted these
results16,17,
and composites of platelet-rich plasma and bone graft with autologous
bone-marrow stromal
cells18 have been
proposed to further improve the repair-promoting
effect19-21.
In this context, an experimental study in rabbits was performed at our
institute in order to compare the bone-healing potential of bone marrow
stromal cells, platelet-rich plasma, and freeze-dried bone allograft
separately and in
combination22. The
research showed that the composite of freeze-dried bone allograft,
platelet-rich plasma, and bone marrow stromal cells induced faster
bone-healing and bone-remodeling processes compared with platelet-rich plasma
and bone-marrow stromal cell treatments without freeze-dried bone
allograft.
The present study was performed in order to compare the early osteogenetic
potential of lyophilized bone chips alone, which is the standard surgical
procedure, and chips supplemented with platelet gel or with platelet gel and
bone marrow stromal cells in patients who underwent a high tibial osteotomy to
treat genu varum and osteoarthritis of the knee.
Patients
Thirty-three patients who had undergone a unilateral opening-wedge high
tibial osteotomy for genu varum and osteoarthritis, with an opening defect of
>1 cm on the medial
side23, were
enrolled and assigned to three groups through random-number generation, to
achieve balance between the groups. Lyophilized bone chips with platelet gel
were used to fill the defect in Group A, lyophilized bone chips with platelet
gel and bone marrow stromal cells were used in Group B, and lyophilized bone
chips alone were used in Group C. Patients who were less than twenty-five
years old or more than sixty-five years old and those with rheumatic or
metabolic diseases, a previous fracture, or deformity correction of <1 cm
were excluded from the study. Two patients (one in Group A and one in Group C)
who had a complete osteotomy with resultant instability were excluded also.
Three patients (one in Group A and two in Group B) declined to participate in
the study. Patient data are reported in
Table I. The institutional
ethics committee on human research approved the clinical study, and the
patients gave their informed consent for participation in the study.
The opening-wedge osteotomy was performed proximal to the tibial
tuberosity, from the medial to the lateral side, leaving intact the lateral
10% of the tibia. The anteromedial side of the osteotomy was fixed with a
titanium spacer plate and screws, just medial to the tibial tuberosity, to
protect it from breaking through the lateral side. The femorotibial angle,
measured on an anteroposterior single-leg radiograph made with the patient in
a standing position, was used to evaluate limb alignment preoperatively and
postoperatively. All of the subjects were instructed to avoid weight-bearing
up to the sixth week postoperatively. The same surgeon (D.D.) performed all
operations under radiographic control with an image intensifier, and an
osteotome was used to create the osteotomy in order to avoid thermal necrosis
at the graft site. During the high tibial osteotomy, lyophilized bone chips
with platelet gel were packed densely into the defects in eleven patients
(Group A), and lyophilized bone chips with platelet gel and bone marrow
stromal cells were used in twelve patients (Group B). In ten patients who
served as controls (Group C), lyophilized bone chips alone were used and were
packed densely as in the other groups. Radiographic evaluation during surgery
demonstrated that all of the defects were completely filled with chips without
any major gaps.
Patients were studied with use of a previously reported, standardized
clinical and laboratory
method24.
Preparation of Lyophilized Human Bone Chips, Platelet Gel, and Bone
Marrow Stromal Cells
Human lyophilized bone chips were produced in the bone bank of our
institute by a previously described freeze-drying
technique24 and
were mixed with platelet-rich plasma and autologous cryoprecipitate, obtained
from autologous fresh-frozen plasma by freeze-thaw precipitation of proteins
and subsequent resuspension in plasma. The platelet gel was formed by
combining 4 mL of thrombin and 16 mL of platelet-rich plasma from the patient
(approximately 1 × 106/µL of platelets). Autologous
bone-marrow stromal cells were obtained from the iliac crest during surgery.
The bone marrow was centrifuged, and 50 mL of buffy coat was obtained. Thirty
milliliters of concentrated bone marrow containing a mean (and standard
deviation) of 24.35 × 103 ± 8.1 nucleated bone
marrow-derived cells (range, 13.0 to 34.5 × 103 cells) per
microliter were used for each sample and were further concentrated to a mean
of 59.1 × 103 ± 22 nucleated cells (range, 28.3 to
90.0 × 103 cells) per microliter. Cells were cultured in
vitro and were characterized for their osteogenic
potential25 and
expression of the osteoblastic phenotype.
The technical details regarding the bone chip, platelet gel, and
bone-marrow stromal cell preparation are presented in the Appendix.
Clinical and Radiographic Evaluation
At six weeks, twelve weeks, six months, and one year after surgery, the
patients underwent a clinical and radiographic evaluation. One year after
surgery, the Knee Society scoring system, as modified by Insall, was used to
define the
outcome26.
Anteroposterior standard radiographs were made at six weeks, twelve weeks, six
months, and one year after surgery with use of a standard technique (65 kV, 20
mA, a focus-film distance of 100 cm, and an equivalent effective dose of
<0.01 mSv within ± 2%). The osteotomy areas on these radiographs
were compared with the same areas on radiographs performed during surgery (the
baseline radiographs). The percentage of integration, based on the density
evaluation on the radiographs, was estimated by blinded observers.
The integration was studied on the medial side of the defect. Bone-graft
radiographic junctions, that is, the lines at the proximal and distal level
where the graft had a different density compared with the host bone, were
considered. Specifically, the distances between the proximal line and the
distal line of the osteotomy were calculated and summarized for all samples in
each group at each follow-up evaluation. The mean values were obtained and
were converted into percentages of integration. These values were used to
attribute a semiquantitative score, with 0+ indicating <10% integration;
1+, 10% to 30%; 2+, 30% to 50%; 3+, 50% to 80%; and 4+, >80%. Six weeks
after surgery, a computed tomography (single-slice spiral scan)-guided needle
biopsy of the graft insertion site was performed. The tissue was removed at
the level of the proximal junction between the bone graft and the host bone,
from the anteromedial aspect of the tibia in front of the plate, with the
Trapsystem device (HS Hospital Service, Rome, Italy). The bone biopsy was
obtained at a depth of 2 cm from the medial cortical margin.
Histology and Histomorphometry
The bone-tissue biopsy specimens were fixed in 10% formalin solution
buffered at pH 7.2. Half of each formalin-fixed sample was decalcified and
embedded in paraffin, and half was dehydrated and embedded in
methylmethacrylate. Five to 10-µm-thick sections of the decalcified
fragments were stained with hematoxylin and eosin (Bio-Optica, Milan, Italy),
and 10 to 20-µm-thick slices of the resin-embedded specimens were stained
with Paragon (Merck, Darmstadt, Germany), Goldner trichromic (Bio-Optica), and
solochrome cyanine (Sigma-Aldrich, Milan, Italy). Two blinded evaluators
performed the histopathological evaluation according to a standardized
protocol24.
Osteogenesis, angiogenesis, and inflammation were estimated (see
Appendix).
Calculations and Statistical Analysis
Measurable parameters were expressed as the arithmetic mean and the
standard error, with the range and median value. A retrospective power
analysis and the analysis of variance test were performed, and the study was
found to be underpowered for vascular buds, osteoclasts, and fibrous tissue
(0.25, 0.31, and 0.08, respectively, and p = 0.27, 0.21, and 0.77,
respectively). Consequently, nonparametric tests were performed.
The Kruskal-Wallis test was used to detect the effects of the graft
material on the bone healing, as well as on the Knee Society score and the
Knee Society functional
score26. The
Mann-Whitney U test was applied to highlight specific differences between
groups. The frequency distribution of osseointegration levels on the
radiographs (+0 to +4) was also calculated, and the Pearson chi-square test
was done. When the number of events collected in a cell of the frequency-table
was =5, the Fisher exact test was applied for statistical differences among
groups. Differences were considered significant if the p value was
<0.05.
Clinical Findings
All patients had relief from knee pain and improvement in walking ability
after the osteotomy, and all were able to walk an unlimited distance at the
latest follow-up examination. The mean range of knee flexion was 118.7°
± 10.5° before the osteotomy and 127.6° ± 18° at the
time of the latest follow-up. Six knees had a residual flexion contracture of
<15°. The final range of motion had increased compared with the
preoperative range in eleven knees, was unchanged in fourteen knees, and was
decreased in three knees.
The clinical and functional evaluation was performed, according to the Knee
Society scoring
system26
(Table II), at twelve months.
With the numbers studied, no significant difference among the groups was found
either preoperatively or postoperatively.
There were no infections or neurological complications. Drainage from the
surgical wound was present fifteen days after surgery in one knee from Group
A, but the culture was negative.
Radiographic Findings
Six weeks after surgery, the osseointegration process
(Fig. 1) was significantly
increased in Groups A and B in comparison with the controls (Group C) (p <
0.005 and p < 0.0001, respectively). In addition, the osseointegration
process was better in Group B than in Group A, as demonstrated by the
osseointegration semiquantitative score. Three of the nine knees in Group A
and seven of the ten knees in Group B had a 30% osseointegration rate; this
trend almost reached significance (p = 0.06).
At twelve weeks after surgery, seven knees from Group A showed a 30% to 50%
graft integration rate (p < 0.0003 compared with the controls) and nine
knees from Group B had a 50% to 80% integration rate (p < 0.0001 compared
with the controls). Moreover, the osseointegration process was faster in Group
B than in Group A (p < 0.05) (Fig.
2).
At six months and one year after surgery, the osseointegration level was
higher in Groups A and B than in Group C, but the significance decreased with
time (from p < 0.001 at six months to p < 0.05 at one year for Group A
compared with the controls, and from p < 0.001 at six months to p < 0.05
at one year for Group B compared with the controls). At both six months and
one year, with the numbers studied, no significant difference was detected
between Groups A and B.
Histology and Histomorphometry
There was no evidence of acute or chronic infection surrounding the bone
grafts: polymorphonuclear cells, lymphocytes, and plasma cells were absent in
all of the groups.
At six weeks after surgery, Group A had extensive bone formation throughout
the defect. At the level of the junction between the graft and the host bone,
a discrete amount of bone marrow-like tissue that filled the spaces among the
bone chips was observed and new linings of osteoblasts in the trabeculae and
vascular buds were also seen (Fig.
3-A). In Group B, bone remodeling was found to be more active in
comparison with that in Group A; tissue filling the defect was rich in
vascular buds and a discrete amount of new mature bone was present
(Fig. 3-B). In contrast, the
specimens in Group C often had fibrous tissue around the bone chips and, in
some cases, there was a histiocytic and/or giant-cell reaction to the grafts,
which were surrounded and phagocytized by cells.
The results of the histomorphometric analysis confirmed a more active
osteogenetic process in Group A (lyophilized bone chips and platelet gel) and
Group B (lyophilized bone chips with platelet gel and bone marrow stromal
cells) in comparison with the controls. The Kruskal-Wallis test, which was
applied to compare all of the groups cumulatively, demonstrated significant
results for new bone apposition areas (p = 0.0008), osteoid areas (p = 0.003),
and vascular buds (p = 0.03). On the contrary, the results for osteoblasts (p
= 0.07), osteoclasts (p = 0.20), and fibrous tissue (p = 0.73) were not
significant, on the basis of the numbers.
When the Mann-Whitney U test was applied to highlight specific differences
between the groups, the values concerning osteoblast number and osteoid areas
in patients who received gel (Group A) and in patients who received gel and
bone marrow stromal cells (Group B) were not significantly different. However,
the values in both groups were significantly increased in comparison with the
controls. Moreover, there was a substantial increase in the percentage of bone
apposition on chips in both Groups A and B, but Group B showed a higher
percentage of bone apposition in comparison with the controls (p = 0.007 for
Group A, p = 0.001 for Group B, and p = 0.05 for Group A compared with Group
B). Similarly, Group B showed greater revascularization compared with the
controls (p = 0.004). The results are given in
Table III.
In orthopaedic surgery, research has focused on procedures that may enhance
bone repair in combination with replacement graft materials. Platelet-rich
plasma has been shown to stimulate osteoblast proliferation in
vitro27 and to
enhance bone
repair8,13,28-31,
presumably because of the high levels of autologous growth factors that do not
induce immunological
reactions28.
Platelet-rich plasma supplemented with fibrin glue to obtain a platelet gel
might confine growth factor secretion to a chosen site, especially when used
in combination with graft materials. Bone marrow stromal cells, which are able
to form osteogenetic and angiogenetic tissue in the presence of stimulatory
factors, such as platelet-derived growth factors, might improve this
effect.
To our knowledge, there is only one preliminary clinical study regarding
the efficacy of a platelet
gel18. Our study is
the first randomized case-control study that we are aware of that has assessed
the effect of platelet gel or platelet gel and bone-marrow stromal cell
supplementation to bone chips used in a high tibial osteotomy to treat genu
varum.
A limitation of our study was that it was underpowered; therefore,
nonparametric statistical tests were performed. However, it is possible that
with larger numbers significant differences could be found.
The same surgeon performed all of the operations, and the bone substitute
preparation was standardized, as was the computed tomography-guided needle
biopsy. Moreover, a semiquantitative analysis of the radiographs and a
histomorphometric evaluation of the bone tissue were performed, always at the
same level as the osteotomy.
It is important to point out that our methods were able to detect slight
differences in early bone formation that were due to the presence of platelet
gel or platelet gel and bone marrow stromal cells in the bone chips; in fact,
the period of direct influence of platelet-derived growth factors is less than
five days. After the initial burst of platelet-rich plasma-related growth
factors, the platelets synthesize and secrete additional growth factors for
the remaining seven days of their life
span9,32,33.
Histomorphometry performed forty-five days after surgery demonstrated that
the addition of the platelet gel to bone chips increased the osteogenesis rate
and enhanced bone formation, as shown by the higher number of osteoblasts,
vascular buds, and bone-forming areas. The addition of bone marrow stromal
cells further accelerated the remodeling process, as shown by the higher
number of bone-forming areas and in comparison with bone chips with platelet
gel alone. Similarly, Group B showed higher revascularization compared with
the other groups. The enhanced bone formation might indirectly depend on such
increased vascularization, perhaps achieved by the exogenous delivery of
angiogenic growth factors by
platelets34. The
improved vascularity might provide better nutrition and increasing resorption
and substitution by healthy tissue. In fact, bone formation is preceded by
vascular invasion, and osteogenesis takes place near newly formed vessels,
which mediate osteoprogenitor cell delivery, secrete mitogens for osteoblasts,
and transport nutrients and
oxygen35.
Regarding the radiographic analysis, we used a score that depended on the
density evaluation as an indirect index of graft osseointegration. It is
noteworthy that, at six weeks, the radiographic evaluation was in agreement
with the histomorphometric data; radiographically, the osseointegration
percentage was significantly increased by the addition of platelet gel or
platelet gel and bone marrow stromal cells in comparison with the controls,
and it confirmed faster graft integration in Group B than in Group A. This
difference seen on the radiographs remained at three months and disappeared by
six and twelve months after surgery. However, both groups maintained a better
pattern of osseointegration in comparison with the controls who, in some
cases, at twelve months, did not show complete graft osseointegration, even if
the patient had good mechanical stability without signs of pseudoarthrosis or
pain.
The findings of this clinical study confirmed the results of other studies,
both in animals36
and in humans18,
and our preliminary
findings23, and
they disagreed with results suggesting that the addition of platelet-rich
plasma does not enhance bone formation in either autografts or
allografts16,17.
These discrepancies might be due to the different platelet-rich plasma
concentrations within the graft. Platelet-derived growth factor levels are
known to influence bone
healing37, and the
amount of platelets required for a positive effect from the platelet-rich
plasma seems to span a relatively low range of concentrations, from
approximately 500,000 to 1,800,000 platelets (median, 1,000,000 platelets) per
microliter of platelet-rich plasma. Below this range, the effect of
platelet-rich plasma is suboptimal. Also, beyond this range, particularly for
concentrations of >2,000,000 platelets per microliter of platelet-rich
plasma, an inhibitory effect has been observed on bone regeneration and on
osteoblast
activity38.
In our study, platelet-rich plasma with a median value of 1 ×
106 platelets per microliter was used. Other authors have
demonstrated an inhibitory effect of such high platelet concentrations in
three different species (miniature pig, dog, and
human)8,12,39.
No explanation has been given for these differences; they could be due to the
inhibitory effects of platelet growth factors. In fact, a
concentration-regulated antimitogenic effect of the transforming growth
factor-beta (TGF-ß), a major growth factor of platelet a-granules,
has been previously
reported40.
Furthermore, TGF-ß1 plays a role in cell migration to the site of future
skeletogenesis, in inducing mesenchymal stem cell differentiation to
osteoblasts, and in inhibiting alkaline phosphatase activity and
mineralization. All of these effects are affected by the TGF-ß1 dose and
its production
site41.
In our patients, the final clinical outcome of lyophilized bone chips with
added platelet gel or platelet gel and bone marrow stromal cells did not
differ from that obtained with the use of bone chips alone. We believe that
this was due to the small size of the defects. Nevertheless, better healing of
the bone gap at the osteotomy site was obtained, while the use of lyophilized
bone chips alone in some patients did not achieve complete
osseointegration.
Cell therapy for bone regeneration with lyophilized bone supplemented with
platelet gel or platelet gel and bone marrow stromal cells is particularly
attractive in the treatment of patients with large bone defects or patients
with a decreased ability to heal spontaneously. We believe that cell therapy
has the potential for clinical applications because both components are
autologous, nontoxic, and nonimmunoreactive, and the treatment is easy to
perform, with minimal side effects. It may be a useful tool for patients with
massive bone loss, as it could shorten the period necessary for bone
regeneration because of higher osteogenetic and angiogenetic activity. Since
this procedure was used during an elective operation in cancellous bone in
well-vascularized areas, we cannot extrapolate these results to cortical bone,
particularly in the presence of trauma or bone tumor, and further research is
necessary in this regard.
A detailed description of the bone substitute preparation, histology and
histomorphometry methods, as well as figures showing bone-staining examples,
are available with the electronic versions of this article, on our web site at
jbjs.org (go to the article citation and click on "Supplementary
Material") and on our quarterly CD-ROM (call our subscription
department, at 781-449-9780, to order the CD-ROM). ?
Note: The authors thank Cristina Tarabusi for her contribution
to the histological specimen preparation and evaluation.
Ito K, Yamada Y, Nagasaka T, Baba S,
Ueda M. Osteogenic potential of injectable tissue-engineered bone: a
comparison among autogenous bone, bone substitute (Bio-oss), platelet-rich
plasma, and tissue-engineered bone with respect to their mechanical properties
and histological findings. J Biomed Mater Res A.
2005;73:
63-72.7363
2005
[PubMed]
Hernigou P, Medevielle D, Debeyre J,
Goutallier D. Proximal tibial osteotomy for osteoarthritis with varus
deformity. A ten to thirteen-year follow-up study. J Bone Joint Surg
Am. 1987;69:
332-54.69332
1987
Goulet JA, Senunas LE, DeSilva GL,
Greenfield ML. Autogenous iliac crest bone graft. Complications and functional
assessment. Clin Orthop Relat Res.
1997;339:
76-8.33976
1997
[PubMed][CrossRef]
Koshino T, Murase T, Saito T. Medial
opening-wedge high tibial osteotomy with use of porous hydroxyapatite to treat
medial compartment osteoarthritis of the knee. J Bone Joint Surg
Am. 2003;85:
78-85.8578
2003
[CrossRef]
Gazdag AR, Lane JM, Glaser D, Forster
RA. Alternatives to autogenous bone graft: efficacy and indications. J
Am Acad Orthop Surg. 1995;3:
1-8.31
1995
Kassolis JD, Rosen PS, Reynolds MA.
Alveolar ridge and sinus augmentation utilizing platelet-rich plasma in
combination with freeze-dried bone allograft: case series. J
Periodontol. 2000;71:
1654-61.711654
2000
[CrossRef]
Landesberg R, Roy M, Glickman RS.
Quantification of growth factor levels using a simplified method of
platelet-rich plasma gel preparation. J Oral Maxillofac Surg.
2000;58:
297-300.58297
2000
[PubMed][CrossRef]
Marx RE, Carlson ER, Eichstaedt RM,
Schimmele SR, Strauss JE, Georgeff KR. Platelet-rich plasma: Growth factor
enhancement for bone grafts. Oral Surg Oral Med Oral Pathol Oral Radiol
Endod. 1998;85:
638-46.85638
1998
[CrossRef]
Marx RE. Platelet-rich plasma (PRP):
what is PRP and what is not PRP? Implant Dent.
2001;10:
225-8.10225
2001
[PubMed][CrossRef]
Sonnleitner D, Huemer P, Sullivan DY. A
simplified technique for producing platelet-rich plasma and platelet
concentrate for intraoral bone grafting techniques: a technical note.
Int J Oral Maxillofac Implants.
2000;15:
879-82.15879
2000
[PubMed]
Cenni E, Ciapetti G, Pagani S, Perut F,
Giunti A, Baldini N. Effects of activated platelet concentrates on human
primary cultures of fibroblasts and osteoblasts. J Periodontol.
2005;76:
323-8.76323
2005
[PubMed][CrossRef]
Kim SG, Chung CH, Kim YK, Park JC, Lim
SC. Use of particulate dentinplaster of Paris combination with/without
platelet-rich plasma in the treatment of bone defects around implants.
Int J Oral Maxillofac Implants.
2002;17:
86-94.1786
2002
[PubMed]
Whitman DH, Berry RL, Green DM. Platelet
gel: an autologous alternative to fibrin glue with applications in oral and
maxillofacial surgery. J Oral Maxillofac Surg.
1997;55:
1294-9.551294
1997
[PubMed][CrossRef]
Bhanot S, Alex JC. Current applications
of platelet gels in facial plastic surgery. Facial Plast Surg.
2002;18:
27-33.1827
2002
[PubMed][CrossRef]
Lowery GL, Kulkarni S, Pennisi AE. Use
of autologous growth factors in lumbar spinal fusion. Bone.
1999;25(2 Suppl):
47S-50S.2547S
1999
[PubMed][CrossRef]
Choi BH, Im CJ, Huh JY, Suh JJ, Lee SH.
Effect of platelet-rich plasma on bone regeneration in autogenous bone graft.
Int J Oral Maxillofac Surg.
2004;33:
56-9.3356
2004
[PubMed][CrossRef]
Jensen TB, Rahbek O, Overgaard S,
Søballe K. Platelet rich plasma and fresh frozen bone allograft as
enhancement of implant fixation. An experimental study in dogs. J
Orthop Res. 2004;22:
653—8.22653
2004
[CrossRef]
Kitoh H, Kitakoji T, Tsuchiya H,
Mitsuyama H, Nakamura H, Katoh M, Ishiguro N. Transplantation of
marrow-derived mesenchymal stem cells and platelet-rich plasma during
distraction osteogenesis—a preliminary result of three cases.
Bone. 2004;35:
892-8.35892
2004
[PubMed][CrossRef]
Cancedda R, Mastrogiacomo M, Bianchi G,
Derubeis A, Muraglia A, Quarto R. Bone marrow stromal cells and their use in
regenerating bone. Novartis Found Symp.
2003;249: 133-47,
170-4, 239-41.249133
2003
[PubMed]
Caplan AI, Bruder SP. Mesenchymal stem
cells: building blocks for molecular medicine in the 21st century.
Trends Mol Med. 2001;7:
259-64.7259
2001
[PubMed][CrossRef]
Lucarelli E, Beccheroni A, Donati D,
Sangiorgi L, Cenacchi A, Del Vento AM, Meotti C, Bertoja AZ, Giardino R,
Fornasari PM, Mercuri M, Picci P. Platelet-derived growth factors enhance
proliferation of human stromal stem cells. Biomaterials.
2003;24:
3095-100.243095
2003
[PubMed][CrossRef]
Dallari D, Fini M, Stagni C, Torricelli
P, Nicoli Aldini N, Giavaresi G, Cenni E, Baldini N, Cenacchi A, Bassi A,
Giardino R, Fornasari PM, Giunti A. In vivo study on the healing of bone
defects treated with bone marrow stromal cells, platelet-rich plasma, and
freeze-dried bone allografts, alone and in combination. J Orthop
Res. 2006;24:
877-88.24877
2006
[CrossRef]
Little DG, McDonald M, Bransford R,
Godfrey CB, Amanat N. Manipulation of the anabolic and catabolic responses
with OP-1 and zoledronic acid in a rat critical defect model. J Bone
Miner Res. 2005;20:
2044-52.202044
2005
[CrossRef]
Savarino L, Cenni E, Tarabusi C, Dallari
D, Stagni C, Cenacchi A, Fornasari PM, Giunti A, Baldini N. Evaluation of bone
healing enhancement by lyophilized bone grafts supplemented with platelet gel:
a standardized methodology in patients with tibial osteotomy for genu varus.
J Biomed Mater Res B Appl Biomater.
2006;76:
364-72.76364
2006
[PubMed]
Ciapetti G, Ambrosio L, Marletta G,
Baldini N, Giunti A. Human bone marrow stromal cells: In vitro expansion and
differentiation for bone engineering. Biomaterials.
2006;27:
6150-60.276150
2006
[PubMed][CrossRef]
Slater M, Patava J, Kingham K, Mason RS.
Involvement of platelets in stimulating osteogenic activity. J Orthop
Res. 1995;13:
655-63.13655
1995
[CrossRef]
Anitua E. Plasma rich in growth factors:
preliminary results of use in the preparation of future sites for implants.
Int J Oral Maxillofac Implants.
1999;14:
529-35.14529
1999
[PubMed]
Knighton DR, Ciresi KF, Fiegel VD,
Austin LL, Butler EL. Classification and treatment of chronic nonhealing
wounds. Successful treatment with autologous platelet-derived wound healing
factors (PDWHF). Ann Surg.
1986;204:
322-30.204322
1986
[PubMed][CrossRef]
Steed DL. The role of growth factors in
wound healing. Surg Clin North Am.
1997;77:
575-86.77575
1997
[PubMed][CrossRef]
Wang HJ, Wan HL, Yang TS, Wang DS, Chen
TM, Chang DM. Acceleration of skin graft healing by growth factors.
Burns. 1996;22:
10-4.2210
1996
[PubMed][CrossRef]
Mundy GR. Regulation of bone formation
by bone morphogenetic proteins and other growth factors. Clin Orthop
Relat Res. 1996;324:
24-8.32424
1996
[CrossRef]
Marx RE. Platelet-rich plasma: evidence
to support its use. J Oral Maxillofac Surg.
2004;62:
489-96.62489
2004
[PubMed][CrossRef]
Rhee JS, Black M, Schubert U, Fischer S,
Morgenstern E, Hammes HP, Preissner KT. The functional role of blood platelet
components in angiogenesis. Thromb Haemost.
2004;92:
394-402.92394
2004
[PubMed]
Eckardt H, Bundgaard KG, Christensen KS,
Lind M, Hansen ES, Hvid I. Effects of locally applied vascular endothelial
growth factor (VEGF) and VEGF-inhibitor to the rabbit tibia during distraction
osteogenesis. J Orthop Res.
2003;21:
335-40.21335
2003
[PubMed][CrossRef]
Yamada Y, Ueda M, Naiki T, Takahashi M,
Hata K, Nagasaka T. Autogenous injectable bone for regeneration with
mesenchymal stem cells and platelet-rich plasma: tissue-engineered bone
regeneration. Tissue Eng.
2004;10:
955-64.10955
2004
[PubMed][CrossRef]
Lee MB. Bone morphogenetic proteins:
background and implications for oral reconstruction. A review. J Clin
Periodontol. 1997;24:
355-65.24355
1997
[CrossRef]
Weibrich G, Hansen T, Kleis W, Buch R,
Hitzler WE. Effect of platelet concentration in platelet-rich plasma on
peri-implant bone regeneration. Bone.
2004;34:
665-71.34665
2004
[PubMed][CrossRef]
Gruber R, Varga F, Fischer MB, Watzek G.
Platelets stimulate proliferation of bone cells: involvement of
platelet-derived growth factor, microparticles and membranes. Clin Oral
Implants Res. 2002;13:
529—35.13529
2002
[CrossRef]
Pollard JW. Tumour-stromal interactions.
Transforming growth factor-beta isoforms and hepatocyte growth factor/scatter
factor in mammary gland ductal morphogenesis. Breast Cancer
Res. 2001;3:
230-7.3230
2001
[CrossRef]
Kanaan RA, Kanaan LA. Transforming
growth factor beta1, bone connection. Med Sci Monit.
2006;12:
RA164-9.12RA164
2006
[PubMed]