自体微小颗粒骨复合骨形成蛋白修复兔桡骨缺损

目的探讨自体微小颗粒骨复合I型胶原以及骨形成蛋白(bone morphogenetic protein，BMP)移植修复节段性兔桡骨缺损的效果。方法新西兰大耳白兔56只，切取自体髂骨研磨成微小颗粒，分别与BMP及I型胶原复合，实验分成4组（n=16)。A组：自体微小颗粒骨复合BMP、I型胶原，B组：自体微小颗粒骨复合I型胶原，C组：自体微小颗粒骨，用于修复兔桡骨干1．5cm缺损的动物模型。D组：空白对照组(n=8)，双侧桡骨缺损不作处理。术后2、4、8和12周，行X线片、组织学观察，骨密度及生物力学检测，比较各移植物修复节段性骨缺损的疗效。结果X线片显示，A组术后8周即可使骨缺损完全修复，而B组术后12周使骨缺损完全修复。术后8、12周骨量测定A组成骨量最多，12周生物力学测定显示移植物修复后的骨缺损具有最佳生物力学表现，而C组则不能完全修复骨缺损。结论自体微小颗粒骨复合BMP、I型胶原及自体微小颗粒骨复合I型胶原均能有效修复节段性骨缺损，以复合BMP移植效果更理想。     

微小颗粒骨复合细胞移植治疗大鼠颅骨缺损

目的 观察自体微小颗粒骨复合骨髓基质细胞(MSC)或成骨细胞移植修复大鼠颅骨缺损的治疗效果。方法 制作大鼠颅骨缺损动物模型，培养同种异体新生大鼠的骨髓基质细胞及成骨细胞，复合自体颗粒骨植入骨缺损区，X线摄片观察骨愈合情况；逆转录一聚合酶链反应(RT-PCR)检测骨钙素、转化生长因子-β1(TGF-β1)表达水平。结果植入微小颗粒骨复合成骨细胞组颅骨缺损愈合最快，骨钙素、TGF-β1表达出现早，与另两组差异有统计学意义(P＜0．05)。植入微小颗粒骨复合骨髓基质细胞组颅骨缺损愈合时间居中，骨钙素、TGF-β1表达早于单纯微小颗粒骨组(P＜0．05)。结论 微小颗粒骨复合细胞移植修复颅骨缺损，细胞因子表达早，缺损愈合明显加快，具有较好的治疗效果。     

煅烧骨与诱导的MSCs复合的实验研究

目的：观察大鼠骨髓基质干细胞（MSCs）与经胶原涂层处理的煅烧骨（CCB）复合行体外培养时贴附、增殖能力的变化,探讨构建组织工程骨体外培养的最适时间。方法：制备鼠尾胶原（I型胶原）,无菌条件下,将煅烧骨置于胶原液中,2h后取出骨块,置滤网中将孔内外的胶原溶液沥干。体外培养的大鼠MSCs经矿化液诱导向成骨细胞分化。将分化后的成骨细胞滴加到涂胶原的煅烧骨块上,同时以未涂胶原的煅烧骨块为对照组。3天,7天时取材,扫描电镜下观察实验组与对照组成骨细胞在煅烧骨表面的贴附数量及形态,用细胞计数法测定细胞在两组材料的生长曲线,组织化学方法检测成骨细胞的ALP活性。结果：扫描电镜发现实验组成骨细胞的贴附数量显著高于对照组（P〈0．01）。细胞计数认为实验组细胞在材料上粘附数量多,增殖速度快。实验组ALP活性明显高于对照组（P〈0．01）。结论：胶原修饰的煅烧骨具有良好的组织相容性,能明显提高成骨细胞的贴附,并且促进成骨细胞活性维持,体外培养7～8天时,材料上的细胞数达到最大,应尽快植入体内。     

自体成骨细胞的诱导培养及其生物学特性

目的 诱导培养来自自体骨髓间充质干细胞的成骨细胞或成骨细胞的前体细胞，进行自体成骨细胞异体骨复合移植实验研究。方法 抽取动物骨髓、抗凝稀释，以适当条件培养诱导，分化为成骨细胞并分析细胞生物学特性。成骨细胞生长在同种异体骨制成的载体上，形成复合物，植入人工缺损处，观察骨修复情况。结果 可以诱导培养出大量的成骨细胞或成骨细胞的前体细胞。自体成骨细胞异体松质骨复合物修复骨缺损的效果明显优于对照组。结论 本实验为自体成骨细胞异体骨复合移植的临床应用提供一个简单有效的方法。     

复合骨形态发生蛋白(BMP)的微小颗粒骨修复兔桡骨缺损的实验研究

目的探讨自体或深低温冷冻同种异体微小颗粒骨复合胶原、骨形态发生蛋白(BMP)修复节段性兔桡骨缺损的效果。方法将兔自体或同种异体骨研磨成微小颗粒，分别与BMP及Ⅰ型胶原复合，并采用兔桡骨干1．5cm缺损的动物模型，通过X线、组织学、骨密度、生物力学等检测手段，与自体微小颗粒骨复合胶原修复节段性骨缺损的疗效比较。结果自体或深低温冷冻同种异体微小颗粒骨复合BMP胶原比自体微小颗粒骨复合胶原成骨效果优良，其中复合BMP组在8周即可使骨缺损修复，髓腔通畅，在骨缺损修复各时期，其成骨速度及成骨量均好于未复合BMP组。结论自体或深低温冷冻同种异体微小颗粒骨复合胶原BMP均可有效地修复节段性骨缺损，两种方法促进新骨形成无明显差异，异体微小颗粒骨复合胶原BMP是良好的骨缺损修复材料。     

生物玻璃结合自体红骨髓复合移植的实验研究

目的：研究生物玻璃结合自体红骨髓复合移植在骨缺损修复过程中对于成骨细胞活性的影响情况。方法：应用影像学观察、病理切片组织学观察、多媒体彩色病理分析系统监测成骨细胞活性水平的表达情况。结果：在影像学方面,自体红骨髓结合人工骨组术后3周即可见可能的骨连接迹象,术后1.5个月可见植入物与骨的确实性结合迹象。在组织学方面,自体红骨髓结合人工骨线可明显促进成软骨细胞和骨祖细胞向 成骨细胞的分化,在骨小梁的重建及钙质沉 答方面 明显优于单独自体红髓髓或人工骨移植。多媒体彩色病理分析活动性成骨细胞表现长（AOS）亦可得出相同结果。结论：生物玻璃结合自体红骨髓复合移植结合了无机材料和有机材料的共同优点,对于骨缺损的修复有明显的作用,骨肿瘤缺损充填及人工假体固定等临床应用方面有广阔的前景。     

骨膜成骨细胞修复骨缺损的实验研究

为了进一步验证骨膜成骨细胞培养建立“成骨细胞库”用于修复骨缺损的可能性，取８只家兔胫前骨膜进行成骨细胞分离培养，体外繁殖后，以明胶海绵为载体，回植于自体的桡骨缺损部。对侧以明胶海绵吸附Ｈａｎｋｓ液为对照。分别在细胞植入后第２，４，８和１２周拍摄Ｘ线片，按Ｌａｎｅ方法计分，评价其骨缺损修复愈合的能力。结果表明，植入的成骨细胞因自身的成骨能力而促进骨缺损的愈合。提示自体“成骨细胞库”的建立和移植在骨缺损的修复中是一种可以继续探索的方法     

成骨细胞、血管内皮细胞复合组织工程骨修复骨缺损的实验研究

目的采用异种骨脱细胞基质（heterogenous bone acellular matrix，HBACM）复合细胞构建组织工程骨并植入动物体内，观察其对骨缺损的修复作用。方法培养与鉴定兔成骨细胞（osteoblast，0B）、血管内皮细胞（vascular endothelial cell，VEC）以及两种细胞的混合细胞。使用猪肋骨制备HBACM，并分别与0B、VEC以及两种细胞的混合细胞共同培养制备组织工程骨。27只新西兰大白兔随机分为三组，切除兔双前肢桡骨制成骨缺损动物模型，左侧植入HBACM与各组细胞复合培养的组织工程骨，右侧植入HBACM为对照。分别于术后3、6、12周取材，观察骨折愈合情况。结果大体形态观察和X线检查表明，组织工程骨修复骨缺损的速度和程度为：联合培养细胞组〉OB组〉VEC组〉单纯HBACM组。BrdU标记细胞示踪检测结果显示：6周时，HBACM上仍可检测到体外培养的标记细胞，并可见新生软骨形成。常规组织学检查可见12周时组织工程骨与正常骨之间的髓腔再通，骨折愈合。Ⅰ型胶原免疫组化研究发现三组均表达Ⅰ型胶原，VEC组略弱。组织切片图像分析示血管面积联合培养细胞组〉VEC组〉OB组，差异有统计学意义。结论联合培养细胞作为组织工程骨种子细胞可以缩短骨折愈合时间、促进骨缺损的修复。     

自体骨膜骨髓复合移植修复肿瘤性骨缺损

目的 介绍自体骨膜骨髓复合移植修复少儿委性骨肿瘤切除后骨缺损的方法。方法 抽取自体髂骨骨髓，自然形成凝块后植入骨缺损处，自体胫前骨膜切片１～２ｍｍ＾２邮票状，均匀植入骨髓凝块中。结果 随访４～１０个月，术后１个月新生骨形成，术后３～４个月１４例骨缺损处新生骨骨密度与周围组织一致，术后４个月恢复正常的关节功能和肢体负重功能。结论 自体游离骨膜骨髓复合移植，成骨能力强，方法简便，是治疗骨肿瘤切除后骨缺     

组织工程化人工骨移植修复长骨干缺损的成骨研究

目的 研究仿生制备的组织工程化人工骨移植修复长骨干缺损的成骨性能、修复效果及可能的修复机制。方法 采用人工合成双相羟基磷灰石(HA／β-TCP)为支架材料，与聚-DL-乳酸(PDLLA)复合后再复合Ⅰ型胶原及重组合人类骨形态发生蛋白-2(rhBMP-2)，与兔骨膜成骨细胞及肾脏血管内皮细胞复合培养。将仿生制备的组织工程化人工骨移植到日本大耳白兔桡骨完全骨膜-骨缺损区，分别于术后第4、8、12周进行大体解剖观察、X线观察、HE染色及Masson三色法染色组织学观察、图像分析、扫描电镜、X线能谱分析，研究骨缺损的修复情况。结果 术后第4周可见新生板层骨；第8周植入物与自体骨呈皮质骨融合，有新骨髓长入；第12周植入物外周被新生皮质骨完全替代，组织学新生骨呈数个连续过渡的条带样分布区。新生骨定量4周组与8周组及12周组比较差异有显著性，8周组与12周组差异无显著性。随植入时间延长，植入体中钙／磷比值趋向于自体皮质骨。结论 仿生构建的组织工程化人工骨植入体内修复长骨干缺损，修复效果好，其骨再生机制为软骨内化骨。     

Bone regeneration in the rat mandible with bone morphogenetic protein-2: a comparison of two carriers.

OBJECTIVE: To compare mandibular bone regeneration with bone morphogenetic protein-2 (BMP-2) delivered with two carriers: a hyaluronic acid polymer (HY), and a collagen carrier complexed with calcium hydroxyapatite and tricalcium phosphate (collagen/HA/TCP). STUDY DESIGN: Defects were created in the bilateral mandibular bodies of 16 Sprague-Dawley rats. The defects were filled with the HY carrier, the HY carrier loaded with BMP-2, the collagen/HA/TCP carrier, or the collagen/HA/TCP carrier loaded with BMP-2. Animals were euthanatized after 6 weeks, and the hemi-mandibles were analyzed histomorphologically. RESULTS: Specimens containing BMP-2 had significantly larger new bone and marrow volumes than control specimens. Specimens in the hyaluronan/BMP-2 group tended to have larger volumes of new bone and osteoid than collagen/HA/TCP/BMP-2 specimens, though these differences were not statistically significant. CONCLUSION: The HY and collagen/HA/TCP carriers had comparable efficacy for bone healing with BMP-2. SIGNIFICANCE: Bone morphogenetic proteins can be delivered with commercially available alloplasts as osteogenic bone substitutes for the repair of craniofacial bone defects. EBM rating: B-2.     

牛脱蛋白松质骨、胶原、BMP、RBM复合物修复骨缺损的实验研究

目的探讨牛脱蛋白松质骨、胶原、骨形态发生蛋白(BMP)、自体红骨髓(RBM)复合物修复骨缺损的能力及其作为自体骨移植替代材料的合理性、可行性.方法在42只家兔双侧桡骨中段制备骨缺损模型,分为4组牛脱蛋白松质骨、胶原、BMP、RBM复合物植入组(A1组20侧)、单纯牛脱蛋白松质骨植入组(A2组20侧)、自体骨植入组(B1组20侧)、空白对照(B2组20侧).在2、4、8、12周分别进行大体标本观察、X线检查、病理组织学检查及生物力学测试.结果 12周时,A1组、B1组骨缺损均已完全修复,而A2组、B2组骨缺损未修复;生物力学测试术后4周A1组的最大扭矩和抗扭刚度均高于B1组,而术后8周后则无显著性差异.结论牛脱蛋白松质骨、胶原、BMP、RBM复合物具有较强的成骨能力,可修复骨缺损,并能作为自体骨移植的一种替代材料.     

聚磷酸钙纤维/磷酸钙骨水泥/微小颗粒骨修复骨缺损的实验研究

目的探讨聚磷酸钙纤维(calcium polyphosphate fibers,CPPF)、磷酸钙骨水泥(calcium phosphate cement,CPC)、自体微小颗粒骨复合材料修复骨缺损的能力。方法选用新西兰大白兔72只,双侧桡骨制成1.5cm骨缺损模型,随机分成6组分别植入。A组:CPPF/CPC/微小颗粒骨复合材料;B组:CPC/微小颗粒骨复合材料;C组:单纯微小颗粒骨;D组:CPPF/CPC复合材料;E组:单纯CPC;F组:空白对照组,不植入任何物质。在2、4、8、12周各时相点,分别进行大体观察、X线照片、组织学切片、扫描电镜观察及力学测试。结果A组在12周可使骨缺损修复,在骨缺损修复各时期,其成骨速度及成骨量均好于其他各组。结论CPPF/CPC/微小颗粒骨复合材料有良好的成骨能力、力学特性和生物相容性,有望成为骨组织工程中修复骨缺损的理想材料。     

A comparison of two biomaterial carriers for osteogenic protein-1 (BMP-7) in an ovine critical defect model

Critical size defects in ovine tibiae, stabilised with intramedullary interlocking nails, were used to assess whether the addition of carboxymethylcellulose to the standard osteogenic protein-1 (OP-1/BMP-7) implant would affect the implant's efficacy for bone regeneration. The biomaterial carriers were a 'putty' carrier of carboxymethylcellulose and bovine-derived type-I collagen (OPP) or the standard with collagen alone (OPC). These two treatments were also compared to "ungrafted" negative controls. Efficacy of regeneration was determined using radiological, biomechanical and histological evaluations after four months of healing. The defects, filled with OPP and OPC, demonstrated radiodense material spanning the defect after one month of healing, with radiographic evidence of recorticalisation and remodelling by two months. The OPP and OPC treatment groups had equivalent structural and material properties that were significantly greater than those in the ungrafted controls. The structural properties of the OPP- and OPC-treated limbs were equivalent to those of the contralateral untreated limb (p > 0.05), yet material properties were inferior (p < 0.05). Histopathology revealed no residual inflammatory response to the biomaterial carriers or OP-1. The OPP- and OPC-treated animals had 60% to 85% lamellar bone within the defect, and less than 25% of the regenerate was composed of fibrous tissue. The defects in the untreated control animals contained less than 40% lamellar bone and more than 60% was fibrous tissue, creating full cortical thickness defects. In our studies carboxymethylcellulose did not adversely affect the capacity of the standard OP-1 implant for regenerating bone.     

新型复合异种骨的研制及其修复骨缺损的免疫组化观察

将人重组骨形成蛋白—2(rhBMP—2)与经综合化学处理的新生小羊骨松质(CB)结合,制备成新型复合异种骨.将其移植修复兔下颌骨缺损,免疫组织化学观察结果表明:复合异种骨移植后,术后各时期移植区成骨活跃,成骨细胞及软骨细胞呈强阳性染色;而单纯松质骨组移植区新骨形成少,染色浅淡,从而证实,rhBMP—2对复合异种骨内新骨形成具有决定性意义.这种新型复合异种骨是一种较理想的植骨材料.     

同种异体脂肪干细胞复合脱钙骨修复大鼠尺骨缺损

目的探讨同种异体脂肪干细胞修复管状骨缺损的可行性。方法获取SD大鼠的腹股沟处脂肪,分离培养脂肪干细胞（Adipose-Derived Stem Cells,ADSCs）;鼠第3代ADSCs与脱钙骨复合,24 h后进行成骨诱导培养。检测细胞在材料表面的生长及成骨分化能力。建立鼠两侧尺骨缺损模型,分别植入鼠ADSCs-脱钙骨复合物（实验侧）和单纯脱钙骨材料（对照侧）;8周、24周后取样,行DR和组织学检测,观察成骨情况。结果 ADSCs能在脱钙骨上很好地黏附和生长,并维持成骨分化能力。细胞-材料复合物植入24周后,DR显示实验侧有新生骨基质长成,对照侧未见骨组织生成。组织学检测显示,实验侧缺损区被典型的骨组织取代,可见新生骨小梁附着于脱钙骨表面;对照侧只有少量的骨组织和纤维组织充填。结论 ADSCs-脱钙骨材料复合物植入,能成功修复临界大小的管状骨缺损。     

A Bioactive Coating Enhances Bone Allografts in Rat Models of Bone Formation and Critical Defect Repair

Bone allografts are inferior to autografts for the repair of critical‐sized defects. Prior studies have suggested that bone morphogenetic protein‐2 (BMP‐2) can be combined with allografts to produce superior healing. We created a bioactive coating on bone allografts using polycondensed deoxyribose isobutyrate ester (PDIB) polymer to deliver BMP‐2 ± the bisphosphonate zoledronic acid (ZA) and tested its ability to enhance the functional utility of allografts in preclinical Wistar rat models. One ex vivo and two in vivo proof‐of‐concept studies were performed. First, PDIB was shown to be able to coat bone grafts (BGs). Second, PDIB was used to coat structural allogenic corticocancellous BG with BMP‐2 ± ZA ± hydroxyapatite (HA) microparticles and compared with PDIB‐coated grafts in a rat muscle pouch model. Next, a rat critical defect model was performed with treatment groups including (i) empty defect, (ii) BG, (iii) collagen sponge + BMP‐2, (iv) BG + PDIB/BMP‐2, and (v) BG + PDIB/BMP‐2/ZA. Key outcome measures included detection of fluorescent bone labels, microcomputed tomography (CT) quantification of bone, and radiographic healing. In the muscle pouch study, BMP‐2 did not increase net bone volume measured by microCT, however, fluorescent labeling showed large amounts of new bone. Addition of ZA increased BV by sevenfold (p < 0.01). In the critical defect model, allografts were insufficient to promote reliable union, however, union was achieved in collagen/BMP‐2 and all BG/BMP‐2 groups. Statement of clinical significance: These data support the concept that PDIB is a viable delivery method for BMP‐2 and ZA delivery to enhance the bone forming potential of allografts. © 2019 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 37:2278–2286, 2019     

Osteogenic activity of OP-1 bone morphogenetic protein (BMP-7) in a human fibular defect.

We performed a prospective, randomised double-blind study in 24 patients undergoing high tibial osteotomy to evaluate the effectiveness of human recombinant osteogenic protein (OP-1) on a collagen type-I carrier in a critically-sized fibular defect. The study had two phases, each evaluated by clinical, radiological and DEXA methods during the first postoperative year. The first concerned the validation of the model of the fibular defect, using positive (demineralised bone) and negative (untreated) controls. The second phase concerned the osteogenic potential of OP-1 on collagen type-I v collagen type-I alone. The results of the first phase established the critically-sized nature of the defect. In the untreated group no bony changes were observed while, in the demineralised bone group, formation of new bone was visible from six weeks onwards. The results of the second phase showed no significant formation of new bone in the presence of collagen alone, while in the OP-1 group, all patients except one showed formation of new bone from six weeks onwards. This proved the osteogenic activity of OP-1 in a validated critically-sized human defect.     

Effect of puerarin on bone formation

OBJECTIVE: Puerarin is one of the major phytoestrogens isolated from Pueraria lobata, a Chinese medicine known as Gegen. Our laboratory compared the amount of new bone produced by puerarin in collagen matrix (carrier) to that produced by the collagen matrix alone. METHOD: Eighteen bone defects, 5mm by 10mm were created in the parietal bone of nine New Zealand White rabbits. In the experimental group, six defects were grafted with puerarin solution mixed with collagen matrix. In the control groups, six defects were grafted with collagen matrix alone (active control) and six were left empty (passive control). Animals were killed on day 14 and the defects were dissected and prepared for histological assessment. Serial sections were cut across each defect. No new bone was formed in the passive control group. Quantitative analysis of new bone formation was made on 100 sections (10 sections in each defect, in five defects randomly selected in each of the experimental group and active control group) using image analysis. RESULTS: A total of 554% more new bone was present in defects grafted with puerarin in collagen matrix than those grafted with the collagen matrix alone. CONCLUSION: Puerarin in collagen matrix has the effect of increasing new bone formation locally and can be used for bone grafting or for bone induction often required in surgery.     

自体成骨细胞--nBGC复合物修复犬胫骨骨缺损

目的：评价搭载自体骨细胞的纳米仿生材料在体内的组织反应性和骨修复作用。方法：4只犬的双侧胫骨各制作一直径l0mm的单侧皮质骨缺损，一侧骨缺损植人复合自体成骨细胞的仿生生物材料作为实验组，另一侧植人自固化磷酸钙(CPC)作为对照组。于术后8周取材，观察各组X线片、组织学、四环素荧光标记变化，比较骨缺损修复情况。结果：X线检查显示nBGC支架与周围骨组织很快融合，界限模糊；而对照组骨缺损界限仍清晰。nBGC支架内大量新骨组织形成，而在CPC植人组新生骨只沉积于植入物的表面，CPC周围有厚纤维组织条带包裹。结论：nBGC支架在体内既能降解又具有生物活性，能融入骨重塑过程，具有良好的组织反应性和骨修复作用，是一种皂好的组织工程骨支架材料．