骨诱导性磷酸钙陶瓷为支架的组织工程骨修复狗下颌骨箱状缺损*☆◆

背景：应用不外加生长因子或细胞而具有骨诱导性的生物材料,在非骨部位构建骨移植物,即体内组织工程骨,其在修复箱状及节段性骨缺损方面,具有更可行的前景。 目的：采用骨诱导性钙磷陶瓷材料构建体内组织工程化类骨移植物,探索其应用于修复实验动物下颌骨箱状骨缺损的可行性。 方法：以骨诱导性磷酸钙陶瓷材料为支架植入狗肌肉内构建体内组织工程骨,同期在狗自体下颌骨左右两侧各拔除牙弓中段牙2颗,形成约20 mm无牙区。8周后在无牙区形成箱状缺损,同期取出支架即刻移植入一侧自体下颌骨缺损区,对侧骨缺损区直接移植入未经体内构建的磷酸钙陶瓷作为对照。 结果与结论：经肌肉内构建的体内组织工程骨移植物的力学性能较单纯磷酸钙陶瓷有明显提高。颌骨缺损区的核吸收强度明显强于对照区,其移植物内长入的骨组织较多,两者的成骨面积差异有非常显著性意义(P < 0.01)。说明在修复颌骨大范围缺损中,体内组织工程骨移植物较单纯骨磷酸钙陶瓷替代材料表现出明显的力学和生物学优势,修复效果显著,有良好的应用前景。关键词：骨诱导性磷酸钙陶瓷;体内组织工程骨;骨缺损;支架;组织工程 缩略语注释：BCP：biphasic calcium phosphate,双相磷酸钙 doi:10.3969/j.issn.1673-8225.2012.16.001     

大段负重骨移植中的生物力学实验研究

用大段自体骨和同种异体骨在羊负重的胫骨上移植后,植骨要产生替代和再生过程。由于组织抗原与移植骨固有性的差异,对上述过程要有不同的力学反应。结果表明：植入骨的愈合,生长过程与其力上关。不同类型移植骨在愈合、生长过程生物力学性质和明显差异。同时也也证明深冷冻、辐照处理的同种异体骨是代替自体移植骨最可受的材料。     

An initial investigation of photocurable three-dimensional lactic acid based scaffolds in a critical-sized cranial defect

Degradable polymer networks formed by the photoinitiated polymerization of multifunctional monomers have great potential as in situ forming materials, especially for bone tissue engineering. In this study, one specific chemistry was analyzed with respect to bone formation in a critical-sized defect model with and without adsorbed osteoinductive growth factors present. The scaffolds degraded in approximately 8 months and possessed an elastic modulus similar to that of trabecular bone. A porous scaffold fabricated with approximately 80% porosity and pore diameters ranging from 45 to 150 mm was implanted in a critical-sized cranial defect in rats. When implanted alone, the scaffolds were filled primarily with fibrous tissue after 9 weeks with only mild inflammation at the defect site. When the scaffolds released osteoinductive growth factors, statistically more bone filled the scaffold. For instance, 65.8+/-9.4% (n=5) of the defects were filled with radiopaque tissue in the osteoinductive releasing scaffolds, whereas only 24.2+/-7.4% (n=5) of the defects were filled in the untreated defects 9 weeks after implantation. These results illustrate not only the benefits of delivering osteoinductive factors when developing synthetic bone grafts, but the potential of these materials for supporting the infiltration and development of bone in large defects.     

Improved osteoconduction of cortical bone grafts by biodegradable foam coating

Alteration of the geometrical surface configuration of cortical bone allografts may improve incorporation into host bone. A porous biodegradable coating that would maintain immediate structural recovery and subsequently allow normal graft healing and remodeling by promoting bony ingrowth could provide an osteoconductive surface scaffold. We investigated the feasibility of augmenting cortical bone grafts with osteoconductive biodegradable polymeric scaffold coatings. Three types of bone grafts were prepared: Type I--cortical bone without coating (control), Type II--cortical bone coated with PLGA-foam, Type III--cortical bone coated with PPF-foam. The grafts were implanted into the rat tibial metaphysis (16 animals for each type of bone graft). Post-operatively the animals were sacrificed at 2 weeks and 4 weeks (8 animals for each type of bone graft at each time point). Histologic and histomorphometric analysis of grafts showed that the amount of new bone forming around the foam-coated grafts was significantly higher than in the control group (uncoated; p < 0.02). Although both foam formulations were initially equally osteoconductive, PLGA-based foam coatings appeared to have degraded at two weeks postoperatively, whereas PPF-based foam coatings were still present at 4 weeks postoperatively. While significant resorption was present in control allografts with little accompanying reactive new bone formation, PLGA-coated bone grafts showed evidence of bone resorption and subsequent bony ingrowth earlier than those coated with PPF-based foams suggesting that PPF-coated cortical bone grafts were longer protected against host reactions resulting in bone resorption.     

The effect of sterilization methods on the osteoconductivity of allograft bone in a critical-sized bilateral tibial defect model in rabbits

Clinically, allogeneic bone graft is used extensively because it avoids the donor site morbidity associated with autograft. However, there are concerns over the optimal sterilization method to eliminate immunological risks whilst maintaining the biological efficacy of the graft. This study compared the effect of Supercritical fluid (SCF) treatment and gamma irradiation at 25 kGy on the osteoconductivity of allograft bone in a bilateral critical sized defect rabbit model. Osteoconductivity was evaluated at 2 and 4 weeks using X-ray, CT, histology (qualitative and quantitative) and immunohistochemistry (Alkaline Phosphatase and Cathepsin-K). Both grafts were well tolerated and osteoconductive. At 2 weeks, there was decreased bone volume and density in the gamma irradiated graft compared to the SCF treated graft, corresponding with a greater inflammatory response histologically and increased Cathepsin-K expression. Catabolic activity predominated at 4 weeks, with both grafts undergoing significant resorption and remodeling inside the defect. Alkaline Phosphatase expression was greater in the SCF group at both time points indicative of a more anabolic response. Allograft bone sterilized with either gamma irradiation or SCF treatment was osteoconductive and capable of healing a critical sized tibial defect in a rabbit. Gamma irradiated allografts elicited an acute inflammatory reaction when implanted which may increase the amount of graft resorption compared to the SCF treated bone.     

Effect of adipose tissue-derived osteogenic and endothelial cells on bone allograft osteogenesis and vascularization in critical-sized calvarial defects

The use of processed bone allograft to repair large osseous defects of the skull has been limited, given that it lacks the osteogenic cellularity and intrinsic vascular supply which are essential elements for successful graft healing and, at the same time, the areas to be targeted through tissue-engineering applications. In this study, we investigated the effect of predifferentiated rat adipose tissue-derived osteoblastic cells (OBs) and endothelial cells (ECs) on calvarial bone allograft healing and vascularization using an orthotopic critical-sized calvarial defect model. For this purpose, thirty-seven 8?mm critical calvarial defects in Lewis rats were treated with bone allografts seeded with no cells, undifferentiated adipose tissue-derived stem cells (ASC), OBs, ECs, and OBs and ECs simultaneously. After 8 weeks, the bone volume and mineral density were calculated using microcomputed tomography and the microvessel formation using immunohistochemical staining and imaging software. The amount of bone within the 8?mm defect was significantly higher for the allografts treated with ECs compared with the allografts treated with OBs (p=0.05) and simultaneously with the two cell lineages (p=0.02). There were no significant differences in bone formation between the latter two groups and the control groups (allografts treated with no cells and undifferentiated ASC). There were no significant differences in bone mineral density among the groups. The amount of microvessels was significantly higher in the group treated with ECs relative to all groups (p=相似文献   

Iron lactate-induced osteomalacia in association with osteoblast dynamics

Osteomalacia was induced in rats fed a diet containing 50,000 ppm (5%) iron lactate for 13 weeks. The histopathology and histomorphometrical dynamics of osteoblasts under this condition were examined. Bone histomorphometry of the proximal tibial metaphysis revealed that the osteoblast surface, osteoid volume, osteoid surface and labeled surface ratio, which are the parameters of bone formation had increased. The blood chemistry revealed the greatest elevation in the osteocalcin level; however, the parathyroid hormone (PTH) secretion and inorganic phosphorus level were very low. From the serum biochemical, histopathological and histomorphometrical findings, the bone lesion in iron lactate-overloaded rats was considered to be similar to low turnover osteomalacia showing decreased trabeculae in secondary spongiosa and increased lamellar osteoid. Furthermore, an iron-positive reaction was detected at the interface between osteoid and mineralized bone. In the bone lesions induced by chronic iron overload, osteoblast recruitment exceeded that of mineralization or, alternatively, the iron within osteoblasts along the trabecular bone suppressed the remodeling and led to an increase in osteoid thickness.     

重组人血管内皮生长因子165/重组人骨形态发生蛋白2/脱蛋白骨与深低温冷冻骨成骨能力的比较

背景：载体+骨诱导因子+生长因子模式人工骨已被证实是理想的人工骨材料。 目的：验证血管内皮生长因子+骨形态发生蛋白与脱蛋白骨复合成重组合人工骨的再血管化及成骨作用,并与深低温冷冻骨比较。 方法：新西兰大白兔左前臂制成桡骨15 mm骨缺损模型,随机分成2组,实验组植入重组血管内皮生长因子165/重组骨形态发生蛋白2/脱蛋白骨;对照组植入深低温冷冻骨。 结果与结论：16周,实验组骨缺损区骨性愈合,移植物密度接近周围正常骨组织;对照组：断端间可见较多骨痂生成,移植物密度稍高于周围正常骨组织。实验组移植物-受体介面无明显分界,达到骨愈合;对照组移植物-受体分界线模糊,部分骨愈合。第3天及第1,2,4,8周,墨汁灌注微血管分析结果显示,实验组血管生成明显多于对照组(P < 0.01);生物力学测试结果显示,实验组三点抗弯曲应力负荷明显强于对照组(P < 0.01)。结果表明,重组血管内皮生长因子165/重组骨形态发生蛋白2/脱蛋白骨重组合人工骨能诱导断端间骨痂形成,加快移植物的血管化速度,且具有良好的生物学功能及生物力学功能。      

The effect of mesenchymal stem cell sheets on structural allograft healing of critical sized femoral defects in mice

Structural bone allografts are widely used in the clinic to treat critical sized bone defects, despite lacking the osteoinductive characteristics of live autografts. To address this, we generated revitalized structural allografts wrapped with mesenchymal stem/progenitor cell (MSC) sheets, which were produced by expanding primary syngenic bone marrow derived cells on temperature-responsive plates, as a tissue-engineered periosteum. In vitro assays demonstrated maintenance of the MSC phenotype in the sheets, suggesting that short-term culturing of MSC sheets is not detrimental. To test their efficacy in vivo, allografts wrapped with MSC sheets were transplanted into 4-mm murine femoral defects and compared to allografts with direct seeding of MSCs and allografts without cells. Evaluations consisted of X-ray plain radiography, 3D microCT, histology, and biomechanical testing at 4- and 6-weeks post-surgery. Our findings demonstrate that MSC sheets induce prolonged cartilage formation at the graft-host junction and enhanced bone callus formation, as well as graft-host osteointegration. Moreover, a large periosteal callus was observed spanning the allografts with MSC sheets, which partially mimics live autograft healing. Finally, biomechanical testing showed a significant increase in the structural and functional properties of MSC sheet grafted femurs. Taken together, MSC sheets exhibit enhanced osteogenicity during critical sized bone defect repair, demonstrating the feasibility of this tissue engineering solution for massive allograft healing.     

Structural bone allograft combined with genetically engineered mesenchymal stem cells as a novel platform for bone tissue engineering

The presence of live periosteal progenitor cells on the surface of bone autografts confers better healing than devitalized allograft. We have previously demonstrated in a murine 4 mm segmental femoral bone-grafting model that live periosteum produces robust endochondral and intramembraneous bone formation that is essential for effective healing and neovascularization of structural bone grafts. To the end of engineering a live pseudo-periosteum that could induce a similar response onto devitalized bone allograft, we seeded a mesenchymal stem cell line stably transfected with human bone morphogenic protein-2/beta-galactosidase (C9) onto devitalized bone allografts or onto a membranous small intestinal submucosa scaffold that was wrapped around the allograft. Histology showed that C9-coated allografts displayed early cartilaginous tissue formation at day 7. By 6 and 9 weeks, a new cortical shell was found bridging the segmental defect that united the host bones. Biomechanical testing showed that C9-coated allografts displayed torsional strength and stiffness equivalent to intact femurs at 6 weeks and superior to live isografts at 9 weeks. Volumetric and histomorphometric micro-computed tomography analyses demonstrated a 2-fold increase in new bone formation around C9-coated allografts, which resulted in a substantial increase in polar moment of inertia (pMOI) due to the formation of new cortical shell around the allografts. Positive correlations between biomechanics and new bone volume and pMOI were found, suggesting that the biomechanical function of the grafted femur relates to both morphological parameters. C9-coated allograft also exhibited slower resorption of the graft cortex at 9 weeks than live isograft. Both new bone formation and the persistent allograft likely contributed to the improved biomechanics of C9-coated allograft. Taken together, we propose a novel strategy to combine structural bone allograft with genetically engineered mesenchymal stem cells as a novel platform for bone tissue engineering.     

Augmentation of osteoinduction with a biodegradable poly(propylene glycol-co-fumaric acid) bone graft extender. A histologic and histomorphometric study in rats

We investigated the feasibility of enhancing the regeneration of skeletal tissues by augmenting bone grafts with a composite biodegradable bone graft extender material based on the polymer poly(propylene fumarate), PPF. The material was mixed with autograft and allograft and placed directly into a cylindrical metaphyseal defect made in the rat tibia. These formulations were compared to defects without any graft material, autografts, allografts and PPF alone. Nine animals were included in each group. Animals were sacrificed at 1 and 4 weeks postoperatively. Implantation sites were then evaluated using histologic and histomorphometric methods. Results of this study showed that defects did not heal in sham operated animals. In the experimental groups, there was early new woven bone formation in the autograft group with near complete healing of the defect at four weeks. When PPF was used alone, gradual ingrowth of new bone was seen. Mixing of the PPF bone graft extender with either allograft or autograft material resulted in enhancement of new bone formation with both allo- and autograft. However, significantly more new bone formation than in the autograft group was only seen when the PPF bone graft extender was mixed with fresh autograft. Histomorphometry corroborated these findings. Results of this study suggest that a PPF-based material may be used to increase the volume of smaller amounts of bone grafts supporting the concept of "bone graft extenders" by application of engineered biodegradable porous scaffolds.     

同种异体冷冻骨移植结合外固定架支撑修复大段骨缺损

背景：各种低温、细胞、免疫和灭菌去抗原技术的进步,使得大量的同种异体骨进入临床应用,给各类大段骨缺损的患者带来了希望。 目的：评价临床应用同种异体冷冻骨移植结合外固定架支撑治疗大段骨缺损的效果。 方法：利用同种异体冷冻骨移植治疗大段骨缺损36例,同时应用外固定架加压固定,提供力学支撑,在固定后定期回访。 结果与结论：随访1~24个月,失访1例,其余35例骨移植区骨质生长良好,9~20个月植骨骨化,无骨不连发生,各关节活动及肢体负重良好。部分患者肢体轻度肿胀,其中3例切口周围轻微红肿疼痛,4例患者伤口渗出较多,细菌培养阴性,涂片染色检查有浆细胞及淋巴细胞,中性粒细胞少,考虑为慢性免疫排斥反应所致,经过换药及应用泼尼松 3 d后分泌物减少,所有患者切口均一期愈合。结果可见同种异体冷冻骨移植结合外固定架加压固定是治疗大段骨缺损的有效治疗方法之一。     

Effects of preservation on the mechanical strength and chemical composition of cortical bone: an experimental study in sheep femora

Preservation methods have enabled bone banks to furnish cortical bone grafts to orthopaedic surgeons. However, cortical bone preserved by freezing and autoclaving, may be weakened by these treatments. To test this hypothesis we compared the ultimate tensile strength of freshly harvested sheep femora with that of femora which were frozen at -20 degrees C for 60 days, or autoclaved at 134 degrees C for 8 min. We measured the collagen and mineral contents (calcium, phosphorus, magnesium) and hydroxyproline of the specimens and tested for changes induced by preservation. Mechanical three point tests showed that frozen femora were significantly stronger than either fresh or autoclaved femora (p<0.05). Frozen specimens also had the highest phosphorus level, indicating these measures are related to strength. Cortical bone is not significantly weakened by autoclaving or freezing. This result does not imply that preserved grafts are clinically interchangeable with fresh grafts, rather, it suggests that future studies should focus on post surgical issues, such as the rate of remodeling and integration, which may be sensitive to preservation technique.     

Tissue engineered vascularized bone formation using in vivo implanted osteoblast-polyglycolic acid scaffold

Repair of skeletal defects with vascularized bone grafts has many advantages over non-vascularized free grafts, but the availability of these grafts is extremely limited. This study was designed to determine whether new vascularized bone could be engineered by transplantation of osteoblasts around existing vascular pedicles using biodegradable, synthetic polymer as a cell delivery vehicle. Cells were isolated from the periosteum of fetal bovine humerus, and then seeded onto non-woven multifilament, polyglycolic acid polymer. The polymers provided three dimensional support during in vitro culture. The cell-polymer constructs were maintained in vitro for two weeks and then implanted around the right femoral vessels of twelve athymic nude rats. The polymer templates without the cells were implanted around left femoral vessels of each mouse as a control. Twelve rats were sacrificed at the following intervals: three rats at six,and nine rats at nine weeks. New bone formation was evident in 10 out of the 12 periosteal-derived cell seeded implants. At six weeks, the tissue was primarily composed of what appeared both grossly and histologically to be cartilage enveloping small islands of osteoid. The degree of osteoid and bone formation progressed with time, as blood vessels invaded the tissue. This tissue ultimately underwent morphogenesis to become an organized trabeculated bone with a vascular pedicle. We believe that this technique may prove to be useful in the reconstruction of bony defect.     

Bone formation in a long bone defect model using a platelet-rich plasma-loaded collagen scaffold

Platelet-rich plasma (PRP), a platelet concentrate made of autogenous blood, has been used in recent years to improve bone defect healing particularly in maxillofacial reconstructions. The aim of the present study was to assess the effect of PRP on new bone formation in a critical diaphyseal long bone defect. A critical size defect (2.5 cm) in the tibial diaphysis of 16 sheep was supplied either with autogenous PRP in a collagen carrier or with collagen alone (controls). Platelets were enriched about 3.5 fold compared to normal blood in the PRP. After 12 weeks, the explanted bone specimens were quantitatively assessed by X-ray, computed tomography (CT), biomechanical testing and histological evaluation. Bone volume, mineral density, mechanical rigidity and histology of the newly formed bone in the defect did not differ significantly between the PRP treated and the control group, and no effect of PRP upon bone formation was observed. It was suggested that PRP does not enhance new bone formation in a critical size defect with a low regenerative potential. Such bone defects might require more potent stimuli, e.g. combinations of functional biomaterials or autografts, precursor cells or osteoinductive growth factors.     

Recent advances in gene delivery for structural bone allografts

In this paper, we review the progress toward developing strategies to engineer improved structural grafting of bone. Three strategies are typically used to augment massive bone defect repair. The first is to engraft mesenchymal stem cells (MSCs) onto a graft or a biosynthetic matrix to provide a viable osteoinductive scaffold material for segmental defect repair. The second strategy is to introduce critical factor(s), for example, bone morphogenetic proteins (BMPs), in the form of bone-derived or recombinant proteins onto the graft or matrix directly. The third strategy uses targeted delivery of therapeutic genes (using viral and nonviral vectors) that either transduce host cells in vivo or stably transduce cells in vitro for subsequent implantation in vivo. We developed a murine femoral model in which allografts can be revitalized via recombinant adeno-associated virus (rAAV) gene transfer. Specifically, allografts coated with rAAV expressing either the constitutively active BMP type I receptor Alk2 (caAlk2), or the angiogenic factor vascular endothelial growth factor (VEGF) combined with the osteoclastogenic factor receptor activator of NF-kappa B ligand (RANKL) have remarkable osteogenic, angiogenic, and remodeling effects that have not been previously documented in healing allografts. Using histomorphometric and micro computed tomography (muCT) imaging we show that rAAV-mediated delivery of caAlk2 induces significant osteoinduction manifested by a mineralized callus on the surface of the allograft, which resembles the healing response of an autograft. We also demonstrate that the rAAV-mediated gene transfer of the combination of VEGF and RANKL can induce significant vascularization and remodeling of processed structural allografts. By contrast, rAAV-LacZ coated allograft controls appeared similar to necrotic allografts and lacked significant mineralized callus, neovascularization, and remodeling. Therefore, innovations in gene delivery offer promising therapeutic approaches for tissue engineering of structural bone substitutes that can potentially have clinical applications in challenging indications.     

Reaming debris as a novel source of autologous bone to enhance healing of bone defects

Reaming debris is formed when bone defects are stabilized with an intramedullary nail, and contains viable osteoblast-like cells and growth factors, and might thus act as a natural osteoinductive scaffold. The advantage of using reaming debris over stem cells or autologous bone for healing bone defects is that no extra surgery is needed to obtain the material. To assess the clinical feasibility of using reaming debris to enhance bone healing, we investigated whether reaming debris enhances the healing rate of a bone defect in sheep tibia, compared to an empty gap. As golden standard the defect was filled with iliac crest bone. Bones treated with iliac crest bone and reaming debris showed larger callus volume, increased bone volume, and decreased cartilage volume in the fracture gap, and increased torsional toughness compared to the empty gap group at 3 weeks postoperative. In addition, bones treated with reaming debris showed increased torsional stiffness at 6 weeks postoperatively compared to the empty defect group, while bending stiffness was marginally increased. These results indicate that reaming debris could serve as an excellent alternative to iliac crest bone for speeding up the healing process in bone defects that are treated with an intramedullary nail.     

Porous poly(propylene fumarate) foam coating of orthotopic cortical bone grafts for improved osteoconduction

A porous biodegradable scaffold coating for perforated and demineralized cortical bone allografts could maintain immediate structural recovery and subsequently allow normal healing and remodeling by promoting bony ingrowth and avoiding accelerated graft resorption. This new type of osteoconductive surface modification should improve allograft incorporation by promoting new bone growth throughout the biodegradable scaffold, hence encasing the graft with the recipient's own bone. We investigated the feasibility of augmenting orthotopically transplanted cortical bone grafts with osteoconductive biodegradable polymeric scaffold coatings. Five types of bone grafts were prepared: type I, untreated fresh-frozen cortical bone grafts (negative control); type II, perforated and partially demineralized cortical bone grafts without additional coating (positive control); type III, perforated and partially demineralized cortical bone coated with a low-porosity poly(propylene fumarate) (PPF) foam; type IV, perforated and partially demineralized cortical bone coated with a medium-porosity PPF foam; and type V, perforated and partially demineralized cortical bone coated with a high-porosity PPF foam. Grafts were implanted into the rat tibial diaphysis. Fixation was achieved with an intramedullary threaded K-wire. Two sets of animals were operated on. Animals were killed in groups of eight with one set being killed 12 weeks, and the other 16 weeks, postoperatively. Radiographic, histologic, and histomorphometric analyses of grafts showed that the amount of new bone forming around the foam-coated grafts was significantly higher than that in the type I control group (uncoated) or that in type II group (perforated and partially demineralized cortical bone grafts). Although all foam formulations appeared initially equally osteoconductive, histologic evaluation of medium-porosity PPF foam-based coatings appeared to result in a sustained response 16 weeks postoperatively. Significant resorption was present in perforated and partially demineralized cortical bone graft allografts, with some accompanying new bone formation occurring primarily within the laser holes. Therefore, PPF foam-coated cortical bone grafts appeared to be better protected from excessive bone resorption, as frequently seen with invasion of fibrovascular tissue. Biomechanical analysis of the PPF foam-coated grafts corroborated findings of the morphometric analysis in that the failure strength at the allograft-host bone junction sites of all PPF-coated cortical bone grafts was higher than in the uncoated controls.     

Bioactivity and osseointegration study of calcium phosphate ceramic of different chemical composition

Synthetic hydroxyapatite (HA) and tricalcium phosphate (TCP) are promising bone-substitute materials in the orthopaedic and dental fields, as their chemical composition is similar to that of bone. This study investigated the osseointegration performance of carbonated biphasic calcium phosphate (CBCP) ceramics containing carbonated hydroxyapatite and tricalcium phosphate prepared by microwave irradiation, in femoral defects of dogs. The defects were created as 3-mm holes on the lateral aspect of femur and filled with the implant material. The serum was collected postoperatively and biochemical assays for alkaline phosphatase activity levels were carried out. The animals' defective sites were radiographed at 4, 8, and 12 weeks. The radiographic results showed that the process of ossification started after 4 weeks and the defect was completely filled with new bone after 8 weeks. Histological examination of the tissue showed the osteoblastic activity inducing the osteogenesis in the defect. The complete haversian system with osteoblastic and osteoclastic activity and bone remodelling process were observed after 12 weeks. The alkaline phosphatase activity levels also correlated with the formation of osteoblast cells. This calcium phosphate ceramic has proved to work well as a biocompatible implant and as an osteoconductive and osteoinductive material for the filling of bone defects.     

BMP-7 and CBFA1 in allograft bone in vivo bone formation and the influence of gamma-irradiation

An initial study showed that morselized human bone grafts were osteoconductive and osteoinductive when implanted in nude rat tibial window defects, and 25 kGy of gamma-irradiation significantly reduced those properties. The mechanism of the osteoinductivity and the influence of gamma-irradiation required further investigation. In this study we assessed the paraffin sections of seven morselized human bone grafts implanted into rat tibial defects for 3 weeks after being treated with 0, 15, or 25 kGy gamma-radiation respectively. Osteoclast-like cell counting and protein expressions of bone morphogenetic protein-7 (BMP-7), core binding factor alpha1 (CBFA1), and proliferating cell nuclear antigen (PCNA) were investigated and the positive signals were quantitatively analyzed. More new bone formation was observed in the 0 and 15 kGy groups compared with 25 kGy groups. The newly formed bones were found mainly from the intact cortex into the defects bridged by the implanted grafts. A dense staining of BMP-7 and CBFA1 was noted in the osteoblast-like cells in those areas. The BMP-7 and CBFA1 staining was also seen in the cells surrounding the implanted grafts in the centre areas of the defects in distance from the intact cortex. Quantitative analysis of immunohistochemical staining of the centre areas of the defects showed that gamma-irradiation (15 and 25 kGy) significantly reduced the expression of CBFA1 and BMP-7. In conclusion, morselized human bone grafts may contain some factors, which induced osteoblast lineage differentiation and bone formation and gamma-irradiation damages those bone inducing factors.