管型与非管型脱钙骨基质支架材料成骨活性差别的实验研究

目的探讨管型与非管型骨缺损修复材料成骨活性的差别。方法以兔同种异体尺桡骨脱钙骨基质为原料,制成管型(A组)及非管型(B组)修复材料各30只,随机植入30只兔双侧桡骨中上段1.5 cm骨缺损中,分别于术后1、2、4、8、12周随机处死6只取材进行X线、组织学切片、大体观察、单位湿重Ca值等检查,结果进行统计学处理。结果早期(术后2周)A组具有更多的骨痂生成(P<0.05)、更高的单位湿重Ca值(P<0.05);中期(术后4周)A组编织骨具有更高的孔隙率(P<0.05);后期(术后8、12周)A组表现为更薄的板层骨、更好的髓腔再通率、更好的塑形、更成熟的骨髓形成(P<0.05)、更高的单位湿重Ca值(P<0.05)。结论实验显示管型脱钙骨基质较非管型具有更好的节段性骨缺损修复能力,提示管型人工骨治疗节段性骨缺损具有美好的前景。     

hBMP-2转染自体兔骨髓基质细胞附和异体兔脱钙骨基质的成骨实验研究

目的　探讨人骨发生蛋白 2 (hBMP - 2 )转染兔自体骨髓基质细胞 (rMSCs)的骨生成诱导作用以及附和异体兔脱钙骨基质 (DBM )后的成骨效能。方法　取兔自体骨髓基质细胞培养扩增 ,用脂质体介导的方法转染人骨发生蛋白 2基因 ,将转染和未转染人骨发生蛋白 2基因的自体骨髓基质细胞附和于异体兔脱钙骨基质形成新的生物植骨材料 ,连同空白及单纯脱钙骨基质对照组植入兔前肢肌袋和桡骨缺损。 2、 4、 6、 8周分别取材进行大体、放射线和病理观察。结果　肌袋试验 6周后 ,转染人骨发生蛋白 2基因的自体骨髓基质细胞附和异体兔脱钙骨基质材料中出现骨组织细胞 ,未转染组和单纯异体兔脱钙骨基质组为纤维组织。骨缺损试验中 ,三组均能产生骨的形成和缺损的修复 ,但仍然是转染复合材料组的骨修复再生能力最大。结论　局部应用hBMP - 2基因转染的rMSCs-DBM材料可以诱导骨形成促进骨修复     

hBMP-2转染自体兔骨髓基质细胞附和异体兔脱钙骨基质的成骨实验研究

目的 探讨人骨发生蛋白2(hBMP-2)转染兔自体骨髓基质细胞(rMSCs)的骨生成诱导作用以及附和异体兔脱钙骨基质(DBM)后的成骨效能。方法 取兔自体骨髓基质细胞培养扩增，用脂质体介导的方法转染人骨发生蛋白2基因，将转染和未转染人骨发生蛋白2基因的自体骨髓基质细胞附和于异体兔脱钙骨基质形成新的生物植骨材料，连同空白及单纯脱钙骨基质对照组植入兔前肢肌袋和桡骨缺损。2、4、6、8周分别取材进行大体、放射线和病理观察。结果 肌袋试验6周后，转染人骨发生蛋白2基因的自体骨髓基质细胞附和异体兔脱钙骨基质材料中出现骨组织细胞，未转染组和单纯异体兔脱钙骨基质组为纤维组织。骨缺损试验中，三组均能产生骨的形成和缺损的修复，但仍然是转染复合材料组的骨修复再生能力最大。结论 局部应用hBMP-2基因转染的rMSCs—DBM材料可以诱导骨形成促进骨修复。     

表面脱钙骨基质明胶修复大块骨缺损的实验与临床研究

目的 探讨表面脱钙骨基质明胶修复大块骨缺损的疗效及相关问题。方法 将32只成年家兔随机分成A、B两组，双侧桡骨制成1cm骨缺损，分别植入表面脱钙骨基质明胶(SDBMG)、全脱钙骨基质明胶(VDBMG)及SDBMG、自体骨，术后定期进行X线、组织学检查(A组)及生物力学测定(B组)；临床采用人SDBMG治疗大块骨缺损31例，术后定期进行X线检查。结果 SDBMG呈由表及里的“渐进”诱导成骨过程；SDBMG的骨修复作用与自体骨相近，SDBMG侧抗压强度与自体骨侧的差异无显著性意义；临床随访示除1例因适应症选择不当而失败外，其它30例均治愈。结论 SDBMG具有良好的成骨作用和可靠的力学性能，可作为自体骨理想的替代材料修复大块骨缺损，但对活动期感染性骨缺损宜慎用。     

脱钙骨基质复合牛骨形态发生蛋白与单纯脱钙骨基质修复兔桡骨缺损比较

目的比较脱钙骨基质(DBM)复合牛骨形态发生蛋白(bBMP)和单纯DBM修复节段性骨缺损的能力.方法32只新西兰大白兔采用桡骨15 mm节段性骨缺损模型,随机分为2组:A组植入异体DBM与bBMP(10 mg)复合材料;B组植入异体DBM.术后4、8、12、16周,进行放射学检查、病理组织学检查和计算机图象分析新生骨面积.结果X线和组织学检查显示异体DBM与bBMP复合材料组的新骨生成、修复骨缺损能力优于异体DBM组,组织切片的计算机图象分析提示DBM bBMP组修复新骨面积大于DBM组,两者之间差异有显著性(4、8、12周P＜0.05,16周P＜0.01).结论异体DBM复合bBMP材料通过骨诱导和骨传导两种方式修复骨缺损,其修复骨缺损的能力要优于单纯DBM材料,是一种较为理想,具有高效成骨活性的植骨材料.     

异位成骨材料修复新西兰兔桡骨干骨缺损

目的探索异位成骨材料修复新西兰兔桡骨干骨缺损的可行性。方法13只新西兰白兔(4周龄,体质量1.5 kg)作为实验动物。取新西兰白兔骨髓间充质干细胞进行成骨诱导,2周后予碱性磷酸酶染色确认其成骨活性。取诱导后的间充质干细胞悬液与β-磷酸三钙生物陶瓷共培养,3天后植入4只新西兰白兔股骨骨膜旁。于植入后第8周收取异位成骨材料,行骨组织切片检查。取8只新西兰兔,制作桡骨干骨缺损动物模型,并将其均分为实验组(4只)和对照组(4只),分别植入异位成骨材料和β-磷酸三钙生物陶瓷修复骨缺损,于术后4、8、12周行手术侧X线摄片,术后12周取修复处骨,行组织切片检查。结果异位成骨植入第8周见植入物成骨完整,获取的成骨材料组织切片显示与股骨干骨细胞结构相似。骨缺损修复动物实验术后X线摄片显示,实验组修复效果显著优于对照组;术后12周修复处骨组织切片显示,实验组新生骨与周边骨组织边界消失,融合度好,骨小梁密度高,新生血管较多。结论异位成骨材料可用于兔桡骨干骨缺损修复。     

家兔自体骨髓移植对骨痂中钙镁铜含量的影响

目的 探讨新骨的形成在骨髓移植术后2周尤为显著现象的机制。方法 用34只家兔剥除双侧桡骨骨膜3cm，截除剥离骨膜后的桡骨中段1cm，1小时后去除骨缺损区血肿，从股骨大转子处抽取1ml骨髓，随机注入一侧桡骨缺损区作为实侧，另一侧注入等量生理盐水作为对照侧，术后2、4、6、8周分批做放射学、组织学和骨痂中钙、镁、铜含量的检查。结果 骨髓移植有肯定的成骨效果，术后2周实验侧骨痂中镁、铜及术后4周镁含量高于对照侧（P＜0．01）。结论 铜和镁在骨髓移植促进骨缺损愈合早期可能具有重要作用，移植术后的2周内补充铜和镁有可能提高骨髓移植的成骨能力。     

带血运骨膜管移植和骨充填物修复桡骨长段缺损的研究

目的：探讨联合应用带血运骨膜管移植和骨充填物治疗兔桡骨长段缺损的效果。方法：实验分两部分，分别选用幼兔和成年兔各40只，根据填充物的不同分为4组，将兔双侧桡骨干中段切除3cm制成骨长段缺损模型，保留切骨段骨膜，重新重原缝合后作带血运骨膜管移植模型，左侧分别用自体骨，同种异体脱钙骨，磷酸三钙陶瓷和羟基磷灰石进行填充，右侧不行任何填作为对照。观察3个月。通过X线片，髓强度，骨密度和组织学检查等方法，了解骨缺损的修复效果。结果：幼兔术后6周，所有实验组双侧的骨缺损均得到修复，术后12周，磷酸三钙陶瓷和羟基磷灰石组桡骨抗弯曲强度较差与自体骨组、同种异体脱钙骨组和对照侧比较具有统计学意义（P＜0．05）；骨愈合为膜内成骨和软骨成骨，以膜内成骨为主，成年兔；各组实验侧骨缺损修复率分别为：自体骨组50％；同种异体脱钙骨组40％；磷酸三钙陶瓷和羟基磷灰石组为30％。对照侧骨缺损修复率为42．5％，结论：幼兔单行单血运骨管移植或结合应用骨充填物均可有效修复骨长段缺损，但置换较慢的骨充填物不利于再生骨强度的恢复，成年兔带血运骨膜移植联合应用骨填充物不能有效修复骨长段缺损。     

基因修饰的组织工程骨联合带血管蒂骨膜移植修复长段骨缺损的研究

目的评价骨形态发生蛋白2（bone morphogenetic protein2,BMP-2）基因修饰的组织工程骨联合带血管蒂骨膜移植修复长段骨缺损的效果。方法分离培养兔骨髓基质干细胞,经BMP-2基因转染后复合异种骨支架体外构建基因修饰的组织工程骨（gene modified tissue engineering bone,GMB）。建立兔双侧桡骨缺损（长2.5cm）模型,采用5种方法修复。A组：GMB＋带血管蒂骨膜移植;B组：GMB＋血管束植入;C组：GMB＋游离骨膜移植;D组：GMB;E组：单纯支架。于术后第4、8、12周行X线、组织学、生物力学测定和微血管墨汁灌注等观察血管形成及成骨情况。结果①A组血运建立快,第8周时即可修复骨缺损,其修复机制包括膜内成骨和软骨成骨两种机制;②B组血管束发出分支向移植骨内长入,但中心区成骨缓慢,第12周时骨缺损得到完全修复;③C组第4周时游离骨膜成活并发出微小血管,第8周时形成薄层外骨痂,第12周时骨缺损基本修复;④D组在BMP-2基因诱导下成骨速度和质量优于E组,可在第12周时使骨缺损部分修复,但中心区呈＂空心＂现象;而E组第12周时形成骨不连,缺损区内被纤维组织填充。结论带血管蒂骨膜与BMP-2基因修饰的组织工程骨联合移植,既提供了血运又提供了骨膜成骨细胞,同时具有良好的骨生成、骨诱导和骨引导作用,是治疗节段性骨缺损较为理想的方法。     

异种脱蛋白骨管复合不同移植材料修复大段骨缺损的实验研究

目的 应用异种脱蛋白骨管复合骨基质明胶(BMG)、骨形态发生蛋白(BMP)、骨膜细胞与复合脱钙骨基质(DBM)、自体红骨髓(RBM)作比较,观察修复猪大段骨缺损的成骨差异.方法 建立猪双侧胫骨大段骨缺损模型.24只成年猪,随机分成2组,实验组A:异种骨管+BMG、BMP、骨膜细胞;实验组B:异种骨管+DBMLRBM;两组均采用锁定钢板及异种皮质骨板固定.分别于术后4、8、12、24周时通过大体标本观察.放射学及组织学观察评估其修复猪大段骨缺损的成骨情况.结果 植入后24周,异种骨管复合BMG、BMP、骨膜细胞完全修复.成骨活跃程度,骨再生量和重建髓腔结构等方面均显著;复合脱钙骨基质、自体红骨髓成骨能力较弱.结论 异种脱蛋白皮质骨管复合骨基质明胶、骨形态发生蛋白及骨膜细胞能够有效修复大段骨缺损.     

海绵状、泥灰状脱钙骨基质修复骨缺损

目的探讨海绵状脱钙骨基质(DBM)和泥灰状DBM修复骨缺损的能力。方法将兔四肢长骨骨干脱脂、脱钙后制成长度为500～1000 μm的纤维状及直径为200～400μm的颗粒状DBM,并与2%明胶混合分别制成海绵状DBM和泥灰状DBM。在9只兔双侧桡骨中段做一长10mm的骨膜骨缺损,分别植入海绵状DBM和泥灰状DBM及空白对照,每组各6个缺损,术后观察6周,于4、6周时拍摄X线片,并对实验动物的大体标本、骨密度、生物力学、新生骨矿化率及病理组织学改变进行观察。结果术后4、6周X线片显示两实验组骨缺损均修复,骨髓腔完整;空白对照组无一例修复骨缺损。骨密度测量显示海绵状DBM组新生骨骨密度与正常桡骨间差异无显著性(P >0.05),但泥灰状DBM组的新生骨骨密度与正常桡骨间差异有显著性(P< 0.05)。术后6周生物力学测定显示海绵状DBM组新生骨极限压缩强度值与正常桡骨差异无显著性(P >0.05),泥灰状DBM组低于正常桡骨且差异有显著性(P< 0.05)。两实验组新生骨矿化率差异无显著性(P >0.05)。组织学观察显示两实验组中DBM绝大部分被吸收,形成板状骨骨小梁及完整的骨髓腔,塑形完整, 新生骨内可见骨单位及局部尚未完全骨化的新生骨。结论海绵状DBM和泥灰状DBM均具有诱导成骨活性和骨传导能力,使用方便,新生骨塑形完整,生物力学强度高,矿化     

复合血管内皮祖细胞的组织工程骨在修复大段骨缺损实验中的血管化评价

目的 评价复合了血管内皮祖细胞(EPCs)的组织工程骨在修复兔桡骨大段骨缺损实验中的血管化情况.方法 自体骨髓通过不同方法的体外培养,获得EPCs及经成骨诱导的骨髓基质干细胞(BMSCs),与脱钙骨基质(DBM)构建组织工程骨修复兔桡骨大段骨缺损.用墨汁灌注、放射性核素骨显像、血管内皮细胞生长因子和Ⅷ因子相关抗原的免疫组化方法观察术后不同时期实验组(EPCs+BMSCs+DBM)、自身对照组(BMSCs+DBM)、阴性对照组(DBM)的骨组织血管化情况.结果 各组手术后2周骨缺损区新生血管数量增加,血流量开始升高,4周和8周时达到峰值,12周后开始下降.实验绀的新生血管数量和局部血流量在术后2、4、8周明显高于自身对照组和阴性对照组,血管排列更整齐.结论 复合EPCs的组织工程骨在修复大段骨缺损时,能促进早期血管化,从而进一步促进成骨.     

Demineralized bone matrix and hydroxyapatite/tri-calcium phosphate mixture for bone healing in rats

Purpose: Hydroxyapatite/tri-calcium phosphate (HA/TCP) mixture is an osteoconductive material used as a bone graft substitute, and demineralised bone matrix (DBM) is an osteoinductive material. A combination of DBM and HA/TCP mixture would probably create a composite with both osteoconductive and osteoinductive properties. The purpose of this study was to determine the effect of the combination of DBM and HA/TCP mixture on healing of rat radius segmental defects. Methods: Twenty-four adult male Wistar rats were used. Bilateral radial defects were created in each animal. Radial defects were implanted with DBM, HA/TCP mixture and a combination of both substances. Control defects were left unfilled. Ten weeks after implantation, the animals were sacrificed, and the radii were evaluated by radiograhic and histopathological studies. Results: The use of DBM alone demonstrated improved healing on radiographic and histological studies compared to other groups and the control group. There were no differences between the other two groups and the control group. Conclusion: The DBM group showed the best healing response. Combined use of DBM and HA/TCP mixture did not improve bone healing, and the osteoinductive properties of DBM were inhibited by HA/TCP mixture.     

Enhanced guided bone regeneration with a resorbable chamber containing demineralized bone matrix

BACKGROUND: The effectiveness of a nonporous poly-DL-lactide tubular chamber in guiding bone regeneration through a long bone defect had already been assessed in an experimental model using the rabbit radius. The injection of bone marrow stem cells into the chamber had proven to enhance bone regeneration. METHODS: The present study reports on the development of the above research project in a subsequent stage. Demineralized bone matrix (DBM) obtained by milling New Zealand rabbit femoral and tibial diaphyses was placed into a tubular chamber. A 10-mm defect was bilaterally created in the radii of 10 rabbits. On the left side (chamber side) the defect was treated by means of a poly-DL-lactide chamber filled with DBM, whereas DBM alone was used on the right side (control). RESULTS: Controls were performed at 3 and 6 months by radiographs and histomorphometry and demonstrated better bone growth on the chamber side versus the control side. A comparison with the results previously obtained by stem cell injection into the chamber revealed significant acceleration of bone regrowth in the first 3 months because of the addition of DBM to the chamber. However, no significant difference was found between the two sides after 6 months. CONCLUSION: These results have confirmed the effectiveness of the chamber as a container for the factors promoting bone regeneration, probably because the osteogenetic activity is maintained in situ.     

自体外周血干细胞/脱钙骨复合移植治疗兔桡骨缺损的实验研究

目的 观察自体外周血干细胞(Autologous Pexipheral Blood Stem Cell—APBSC)/脱钙骨(Decalcified Bone Matrix-DBM)复合移植治疗骨缺损的疗效。方法 36只家兔桡骨做成1cm骨缺损,随机分为DBM组自体红骨髓(Autologous red bone marrow—ARBM)/DBM组APBSC/DBM组,每组12只。分别进行X线观察、生物力学检查和组织学观察的对比研究。结果 术后第2、4、8、14周APBSE/DBM组X线观察改进的Gary评分和光镜观察均优于DBM组,与ARBM/DBM组相似,术后第14周APBSC/DBM组整骨破坏载荷和骨缺损修复,形态学评分也优于DBM组,与ARBM/DBM组比较,前者没有显著性差异,后者有显著性差异。结论 APBSC/DBM复合移植治疗骨缺损的疗效优于单纯DBM移植,与ARBM/DBM组相似。结合临床我们认为具有良好的应用前景。     

Repair of large segmental bone defects using bone marrow stromal cells with demineralized bone matrix

Objective: The aim of the present study was to evaluate the effect of tissue‐engineered constructs on repair of large segmental bone defects in goats. Methods: Allogenic demineralized bone matrix (aDBM) was seeded with autologous marrow stromal cells (aMSC) for seven days to construct DBM–MSC grafts prior to implantation. 24 goats were randomly divided into three groups (eight in each). In each group, 3 cm diaphyseal femoral defects were created unilaterally, and subsequently filled with the DBM‐MSC grafts, DBM alone and an untreated control, respectively. Radiological analysis and biomechanical evaluation were performed at 12 and 24 weeks after operation. Results: Obvious increases in radiological scoring and biomechanical strength were found in the DBM‐MSC group when compared to the DBM group. X‐ray examination showed excellent bone healing in the DBM‐MSC group, whereas only partial bone repair was seen in the DBM group, and no healing in untreated controls. Histologically, a tendency to bone regeneration and remodeling was far more obvious for the DBM‐MSC group than the DBM only and untreated controls. Conclusion: Our results strongly suggest that transplantation of bone MSC within a DBM could have advantages for the bone repair of large segmental defects.     

Demineralized bone matrix composite grafting for posterolateral spinal fusion

In a prospective series, the 24-month fusion status was evaluated radiographically among patients undergoing instrumented posterolateral lumbosacral spinal fusion. Seventy-three patients had a diagnosis of degenerative disk disease or degenerative spondylolisthesis and had supplemental bone grafting with demineralized bone matrix (DBM) putty (Grafton DBM; Osteotech, Eatontown, NJ) enriched with aspirated bone marrow (DBM bone marrow), DBM putty combined with iliac crest autograft (DBM autograft), or autograft. Overall, approximately 63% (12 of 19) of DBM bone marrow, 70% (19 of 27) of DBM autograft, and 67% (18 of 27) of autograft patients were fused at 24 months (P = .875). These findings suggest that both DBM composites offer similar performance to autograft in posterolateral spinal fusion.     

Augmented demineralized bone matrix: a potential alternative for posterolateral lumbar spinal fusion

Variable osteoinductive potential has been reported between and within production lots of different demineralized bone matrix (DBM) products. This study compared fusion rates of different manufactured lots and augmented formulations of DBM with a dose-response curve of recombinant human bone morphogenetic protein 2 (rhBMP-2) on inactivated DBM carrier in a posterolateral fusion rat model. Lumbar fusions were performed in 145 rats. In the control rats, we implanted autograft, graft alternative, including inactivated DBM, or nothing (ie, no graft). In the study rats, we implanted 1 of 2 BioSETR (RTI Biologics, Alachua, Florida) DBM lots, growth factor-enriched DBM, and inactivated DBM plus rhBMP-2 in different concentrations. Manual palpation revealed fusion rates of 25% (autograft), 0% (inactivated DBM), 17% (DBM donor A), and 36% (DBM donor B). The fusion rate of the most enhanced donor B graft (83%) was higher (P<.05) than that of autograft or unenhanced DBM. Inactivated DBM plus rhBMP-2 fused between 45% and 100%. There was no significant difference between DBM plus rhBPM-2 and the highest enrichment group of donor B. Differences between 2 DBM lots in an athymic rat ectopic bone formation model also were found in the spine fusion model. Enhanced DBM formulations were comparable with inactivated DBM plus rhBMP-2 with respect to performance and could represent a bone graft alternative in spine fusion.     

Grafting long bone fractures with demineralized bone matrix putty enriched with bone marrow: pilot findings

In this pilot study, the preliminary effectiveness of a composite graft consisting of demineralized bone matrix (DBM) putty (Grafton DBM) and aspirated bone marrow was evaluated for treating long bone fractures. Patients were ssigned randomly to treatment with the DBM putty composite (n = 10) or iliac crest autograft (n = 8), and had a minimum of 12 months of radiographic follow-up. Ninety percent of DBM patients (9/10) achieved full bone formation compared to 75% of autograft patients (6/8) (P = .41). Additionally, all 10 DBM patients were healed compared with 63% of autograft patients (5/8) (P = .07). These findings suggest that DBM putty enriched with bone marrow may be comparable to autograft for treating long bone fractures.