带血供肌瓣作为骨形态发生蛋白载体修复骨缺损的实验研究

目的 探讨带血供肌瓣作为骨形态发生蛋白（BMP）载体修复骨缺损的可行性。方法 观察带血从肌瓣复合BMP和单纯BMP组修复骨缺损时的成骨情况;对纤维蛋白粘合剂、带血供肌瓣、无血运肌瓣、同种异体脱钙骨4种不同BMP载体的成骨能力进行比较。结果 以指深屈肌支为蒂制备的带血供肌瓣复合BMP修复骨缺损,效果优于单纯BMP组;带血供肌瓣联合纤维蛋白粘合剂复合BMP修复骨缺损,效果优于单纯BMP组;带血供肌瓣联合纤维蛋白粘合剂复合BMP组修复骨缺损,效果优于其它载体。结论 带血供肌瓣可作为BMP的良好载体。带血供肌瓣联合纤维蛋白粘合剂作BMP的载体效果更优。     

Intraosseous BMP implants in rabbits. Inhibitory effect on bone formation

The bone harvest chamber is a model for rapid spontaneous bone healing in rabbits. We have previously shown inhibition of bone formation by using BMP-2 on a collagen carrier in this intraosseous model, despite bone formation when depositing BMP-2 on a similar carrier subfascially in the same animals. The doses were 12 and 0.6 micrograms/5 mm3 chamber volume. As these findings conflicted with most other experiments dealing with the skeletal response to BMP-2, we repeated the previous experiments with variations. We studied: 1) a lower BMP-2 dose, 2) a different type of BMP (BMP-7/OP-1), 3) a different carrier (hydroxyapatite), 4) a different chamber construction allowing contact with extraskeletal tissue and 5) BMP-2 on the original collagen carrier in an acutely inserted chamber in rats. We also studied the border between the BMP-2 implant and the preexisting bone to see whether BMP-2 caused premature differentiation of the callus so that proliferation was stopped and a bone cyst formed. The low dose of BMP-2 reduced tissue ingrowth and tended to reduce bone formation. BMP-7 showed the same inhibitory effects as BMP-2. BMP-2 on a hydroxyapatite carrier also inhibited bone formation in the chamber. In the chamber that allowed contact with extraskeletal tissue, we observed no effects of BMP-2. The border between the BMP-2 implant and the preexisting bone did not look like a cyst wall. BMP-2, from the same batch, on a similar collagen carrier, regularly increased bone formation in the acutely inserted bone chamber in rats, thereby excluding major defects in the BMP-2 implants. The inhibition in this specific model is a consistent finding and not due to an overdose, a specific BMP-type, a specific carrier or premature callus differentiation.     

Differentiation of cartilage on three substrata under the influence of an aggregate of morphogenetic protein and other bone tissue noncollagenous proteins (BMP/iNCP)

A cellulose acetate membrane was fashioned into a cone to serve as a substratum for an outgrowth of connective tissue from normal neonatal muscle, and a container for diffusion of bone morphogenetic protein (BMP) of an aggregate of BMP and cold water insoluble noncollagenous protein (BMP/iNCP). The BMP/iNCP was prepared by dissociative extraction and differential precipitation with other bone matrix proteins that slowly become soluble and diffusible in culture media at 37 degrees. The BMP/iNCP was applied either on, within, or beneath the surface of the explants or suspended in the culture medium. Under the influence of BMP in tissue cultures, without any bone matrix or bone collagen in the system, connective tissue outgrowths of muscle differentiate into cartilage on three substrata: (1) cellulose acetate membranes with pore size of 0.45-5.0 micron; (2) remnants of undissolved BMP/iNCP; and (3) degenerating myofibers. The cartilage developed in the interior of muscle, possibly by phenotypic cell transformation, when the pore size of the membrane was 0.1-0.22 micron too small to sustain anchorage of the explant. Cartilage developed on particle surfaces when the muscle tissue and BMP/iNCP particle were minced and mixed before explantation. The cartilage preferentially grew out directly onto the cellulose acetate membrane when the pore size was optimal for anchorage and the BMP/iNCP was suspended on the surface of the explant to either simultaneously percolate through the explant or diffuse through the culture medium. The biosynthetic activity of cells proliferating before and associated with cell differentiation was measured by 35S uptake in total glycosaminoglycan (GAG) per microgram of DNA. When the pore size was 8.0 micron, large enough to permit cells to migrate across the membrane, a thick plate of fibrous connective tissue developed on the undersurface of the membrane without any evidence of cartilage cell differentiation in any location. Repeated doses of BMP/iNCP with each change of culture medium produced a greater incidence and quantity of cartilage than a single dose, but the 35S incorporation into GAG always reached peak levels, in the interval between four and ten days, irrespective of the schedule of administration or dosage. These observations suggested that the exogenous or endogenous noncollagenous proteins are a carrier for BMP and can substitute for whole bone matrix or bone collagen.     

Experimental spinal fusion with use of recombinant human bone morphogenetic protein 2.

STUDY DESIGN: Posterolateral lumbar spinal fusion with use of recombinant human bone morphogenetic protein 2 (rhBMP-2) was tested in rabbits by implanting composites of rhBMP-2 and collagen carrier. OBJECTIVES: To examine the bone-formation-inducing activity of rhBMP-2 and find the optimal amount of rhBMP to add to a collagen carrier to constitute bone-formation-inducing implants to be substituted for bone graft in posterolateral spinal fusion in rabbits. SUMMARY OF BACKGROUND DATA: In animal models, rhBMP-2--impregnated collagen has been successfully used for posterolateral spinal fusion, indicating that it is a potential substitute for the autogenous corticocancellous bone graft currently used most routinely in posterolateral lumbar spinal fusion. METHODS: Nine rabbits were divided into three equal groups. The bilateral L4-L5 transverse processes were exposed, and collagen strips impregnated with rhBMP-2 (10, 50, or 200 micrograms) were placed on the left transverse processes, and collagen strips alone were inserted on the right. All rabbits were killed 24 weeks after surgery. The implanted sites were assessed for new bone formation and bony fusion by radiography and histologic examination. RESULTS: New bone formation was noted in intertransverse spaces on the left side of all rabbits except one (10 micrograms rhBMP-2). Twelve weeks after implantation, no new bone formation was seen on the right side of all animals. The newly formed bone masses were significantly larger in the 50-microgram and 200-microgram rhBMP-2 groups than in the 10-microgram rhBMP-2 group (P < 0.01), but there was no significant difference between bone formation in the 50-microgram and 200-microgram groups (P = 0.647). CONCLUSIONS: The rhBMP-2/collagen composite implant was an effective bone graft substitute for achieving posterolateral spinal fusion. When combined with a collagen carrier, the optimal rhBMP-2 dose for achieving posterolateral spinal fusion seemed to be approximately 50 micrograms per segment in rabbits.     

Osteogenesis by recombinant human bone morphogenetic protein-2 at skeletal sites

Osteogenesis was evaluated in the mandibular bone by combinations of various dosages of recombinant human bone morphogenetic protein-2, atelopeptide Type I collagen, and porous hydroxyapatite (four groups: Group I, 2 micrograms recombinant human bone morphogenetic protein-2, atelopeptide Type I collagen, and porous hydroxyapatite; Group II, 10 micrograms recombinant human bone morphogenetic protein-2, atelopeptide Type I collagen, and porous hydroxyapatite; Group III, 50 micrograms recombinant human bone morphogenetic protein-2, atelopeptide Type I collagen, and porous hydroxyapatite; Control Group, only atelopeptide Type I collagen and porous hydroxyapatite). The prepared materials were implanted in the mandibular bone hole (7 mm in diameter, 2 mm deep). Three weeks later, the alkaline phosphatase activity in the implanted region was determined, and the histologic features of the excised tissue were examined. There were significant differences in histologic and biochemical findings among the four groups. In the recombinant human bone morphogenetic protein-2 implanted groups, osteogenesis increased with the dosage of recombinant human bone morphogenetic protein-2, as assessed by alkaline phosphatase activity and histologic findings. The results suggest that atelopeptide Type I collagen is an effective carrier for recombinant human bone morphogenetic protein-2 and that porous hydroxyapatite would be advantageous for clinical application as a material to maintain its original form after implantation.     

Low dose fibroblast growth factor-2 (FGF-2) enhances bone morphogenetic protein-2 (BMP-2)-induced ectopic bone formation in mice

To examine how fibroblast growth factor-2 (FGF-2) affects the BMP signaling pathway during bone morphogenetic protein-2 (BMP-2)-induced ectopic bone formation, we implanted type I collagen disks containing constant amounts of BMP-2 (5 micrograms) and varying amounts of FGF-2 onto the back muscles of adult male mice. We then performed histological analyses and histomorphometry, and measured bone mineral density and radiopaque area on the discs 1, 2, and 3 weeks after implantation. We also determined the expression profiles of several genes involved in bone formation and the BMP signaling pathway in the muscle that had been adjacent to the implanted disc and in muscle-derived primary culture cells that had similarly been treated with a constant concentration of BMP-2 and a varying concentration of FGF-2. In the presence of a constant amount of BMP-2, we confirmed that low doses of FGF-2 increased ectopic bone formation in vivo and high doses inhibited bone formation. Northern and/or Western blots of recovered muscle from the in vivo experiment and treated muscle-derived primary culture cells from the in vitro experiment revealed that low doses of FGF-2, but not high doses, increased the expression BMP receptor (BMPR)-1B, phosphorylated Smad1, Noggin, and Osteocalcin. Our results indicate that low-dose FGF-2 may facilitate BMP-2-induced ectopic bone formation by altering the expression of BMPRs on the surface of bone forming progenitor cells. They also indicate that the inhibitory effect of high-dose FGF-2 is not mediated via increased expression of the BMP inhibitor Noggin.     

Repair of Osteochondral Defects in a Rabbit Model Using a Porous Hydroxyapatite Collagen Composite Impregnated With Bone Morphogenetic Protein‐2

Articular cartilage has a limited capacity for spontaneous repair, and an effective method to repair damaged articular cartilage has not yet been established. The purpose of this study was to evaluate the effect of transplantation of porous hydroxyapatite collagen (HAp/Col) impregnated with bone morphogenetic protein‐2 (BMP‐2). To evaluate the characteristics of porous HAp/Col as a drug delivery carrier of recombinant human BMP‐2 (rhBMP‐2), the rhBMP‐2 adsorption capacity and release kinetics of porous HAp/Col were analyzed. Porous HAp/Col impregnated with different amounts of rhBMP‐2 (0, 5, and 25 μg) was implanted into osteochondral defects generated in the patellar groove of Japanese white rabbits to evaluate the effect on osteochondral defect regeneration. At 3, 6, 12, and 24 weeks after operation, samples were harvested and subjected to micro‐computed tomography analysis and histological evaluation of articular cartilage and subchondral bone repair. The adsorption capacity was 329.4 μg of rhBMP‐2 per cm³ of porous HAp/Col. Although 36% of rhBMP‐2 was released within 24 h, more than 50% of the rhBMP‐2 was retained in the porous HAp/Col through the course of the experiment. Defects treated with 5 μg of rhBMP‐2 showed the most extensive subchondral bone repair and the highest histological regeneration score, and differences against the untreated defect group were significant. The histological regeneration score of defects treated with 25 μg of rhBMP‐2 increased up to 6 weeks after implantation, but then decreased. Porous HAp/Col, therefore, is an appropriate carrier for rhBMP‐2. Implantation of porous HAp/Col impregnated with rhBMP‐2 is effective for rigid subchondral bone repair, which is important for the repair of the smooth articular surface.     

Intraosseous BMP implants in rabbits Inhibitory effect on bone formation

The bone harvest chamber is a model for rapid spontaneous bone healing in rabbits. We have previously shown inhibition of bone formation by using BMP-2 on a collagen carrier in this intraosseous model, despite bone formation when depositing BMP-2 on a similar carrier subfascially in the same animals. The doses were 12 and 0.6 μg/ 5 mm³ chamber volume. As these findings conflicted with most other experiments dealing with the skeletal response to BMP-2, we repeated the previous experiments with variations. We studied: 1) a lower BMP-2 dose, 2) a different type of BMP (BMP-7/OP-1), 3) a different carrier (hydroxyapatite), 4) a different chamber construction allowing contact with extraskeletal tissue and 5) BMP-2 on the original collagen carrier in an acutely inserted chamber in rats. We also studied the border between the BMP-2 implant and the preexisting bone to see whether BMP-2 caused pre mature differentiation of the callus so that proliferation was stopped and a bone cyst formed. The low dose of BMP-2 reduced tissue ingrowth and tended to reduce bone formation. BMP-7 showed the same inhibitory effects as BMP-2. BMP-2 on a hydroxyapatite carrier also inhibited bone formation in the chamber. In the chamber that allowed contact with extraskeletal tissue, we observed no effects of BMP-2. The border between the BMP-2 implant and the preexisting bone did not look like a cyst wall. BMP-2, from the same batch, on a similar collagen carrier, regularly increased bone formation in the acutely inserted bone chamber in rats, thereby excluding major defects in the BMP-2 implants. The inhibition in this specific model is a consistent finding and not due to an overdose, a specific BMP-type, a specific carrier or premature callus differentiation.     

骨胶原的制备及作为骨形成蛋白载体修复骨缺损的研究

在提取BMP的过程中,从废去的中间产物骨基质胶中同时提取胶原作为BMP的载体用于修复兔颅骨缺损.40只大白兔随机分为A、B两组,在兔颅顶骨两侧对称各制造直径为1cm的圆形骨缺损,A组兔颅顶骨缺损左侧植入骨胶原/BMP,右侧植入单纯骨胶原,B组兔左侧植入BMP,右侧空白对照,组织学检查显示:植入后7夭,骨胶原/BMP组即有软骨细胞岛出现,旦在各期的成骨面积均大于单纯BMP组和胶原组.另外,移植材料内的灰量测定结果及X线摄片结果也表明,骨胶原/BMP的成骨量明显大于单纯骨胶原组和BMP组(P<0.01),认为骨胶原作为BMP的载体有更多的优越性.     

Augmentation of fracture healing by hydroxyapatite/collagen paste and bone morphogenetic protein‐2 evaluated using a rat femur osteotomy model

In fracture treatment, biological bone union generally depends on the bone's natural fracture healing capacity, even in surgically treated cases. Hydroxyapatite/collagen composite (HAp/Col) has high osteoconductivity and stimulates osteogenic progenitors. Furthermore, it has the potent capacity to adsorb bone morphogenetic proteins (BMPs). In this study, we prepared an injectable HAp/Col paste and evaluated its augmentation of bone union. Furthermore, the effect of HAp/Col paste combined with BMP‐2 was also evaluated. We used a rat femur osteotomy model with a defect size of 1 mm. Male Wistar rats were assigned to one of the following four groups; a control group without any implant, a HAp/Col implant group, a group that received an absorbable collagen sponge (ACS) implant impregnated with BMP‐2 (1 μg), and a group that received a HAp/Col implant impregnated with BMP‐2 implant. Micro‐CT analysis, three‐point bending tests, and histological evaluation were performed. Bone union was achieved in two of eight cases in the HAp/Col group, five of eight cases in the ACS + BMP‐2 group, and all cases in the HAp/Col + BMP‐2 group at 8 weeks post‐surgery. The control group did not achieve bone union. In addition, in the HAp/Col + BMP‐2 group, the biomechanical strength of the fused femurs was comparable to that of the contralateral intact femur; the ratio of the mechanical load at the breaking point of the osteotomy side relative to that of the contralateral side was 1.00 ± 0.151 (SD). These results indicate that HAp/Col paste with or without BMP‐2 augments bone union. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 36:129–137, 2018.     

Immunolocalization of bone morphogenetic protein and its receptors in degeneration of intervertebral disc.

STUDY DESIGN: Immunohistochemical study of expression and localization of bone morphogenetic protein (BMP)-2/4 and type I and II receptors on intervertebral disc. OBJECTIVES: To determine the biologic functions of BMPs and their receptors in the process of degeneration of the intervertebral disc. SUMMARY OF BACKGROUND DATA: Biologic and pathologic processes in the cell during the degeneration of the intervertebral disc are as yet poorly understood. METHODS: The cervical spines of 15 male senescence-accelerated mice aged 8, 24, or 50 weeks were used for histologic and immunohistochemical examination of BMP-2/4 and BMP receptors IA, IB, and II. Immunostaining was performed with the avidin-biotin-peroxidase complex method. RESULTS: Degenerative change was recognized within intervertebral discs of senescence-accelerated mice aged 50 weeks. BMP-2/4 and its receptors were abundant in hyaline cartilaginous cells within the endplate of the vertebrae at 8 and 24 weeks of age. However, the expression of BMP-2/4 and its receptors moved from the hyaline cartilage of the endplate of the vertebrae to fibrous cells within the anulus and to the calcified cartilage at the site of enthesis of mice aged 50 weeks. CONCLUSIONS: BMP-2/4 and its receptors may play roles in degenerative change of intervertebral disc.     

Induction of callus formation by implants of bone morphogenetic protein and associated bone matrix noncollagenous proteins

Callus formation in the periosteal bone interface in response to bone morphogenetic protein (BMP) and associated bone matrix noncollagenous proteins (BMP/NCP) was investigated in mature adult rabbits. For controls byproducts of BMP/NCP purification, bone marrow, eight nonskeletal tissues, purified matrix gamma-carboxyglutamic acid-rich protein (MGP), and a composite of BMP/NCP and polylactic-polyglycolic acid polymer (PLA/PGA) were also implanted in the periosteal bone interface. Quantitative microcomputer image analysis and histologic studies were performed three weeks after the implantation. BMP/NCP and bone marrow or BMP/NCP implanted over a single drill hole into the marrow cavity produced three times more new bone than the bone marrow alone. BMP/NCP alone produced twice as much new bone as bone marrow alone. Control implants of bovine serum albumin or purified MGP produced no new bone. Autogeneic minced muscle and ten nonskeletal tissue controls produced little or no bone formation. Even at one-fifth of the dose of BMP/NCP, a composite of PLA/PGA incorporating BMP/NCP showed almost the same amount of new bone as BMP alone. Histologically, the response to BMP/NCP consisted of an external callus of calcifying cartilage and woven bone. The response to subperiosteal implants of BMP/NCP or BMP/NCP with bone marrow or with minced muscle occurred with the same sequence of developmental events as seen either in embryonic skeletogenesis or in fracture callus.     

Repair of large osteochondral defects in rabbits using porous hydroxyapatite/collagen (HAp/Col) and fibroblast growth factor‐2 (FGF‐2)

Articular cartilage has a limited capacity for self‐renewal. This article reports the development of a porous hydroxyapatite/collagen (HAp/Col) scaffold as a bone void filler and a vehicle for drug administration. The scaffold consists of HAp nanocrystals and type I atelocollagen. The purpose of this study was to investigate the efficacy of porous HAp/Col impregnated with FGF‐2 to repair large osteochondral defects in a rabbit model. Ninety‐six cylindrical osteochondral defects 5 mm in diameter and 5 mm in depth were created in the femoral trochlear groove of the right knee. Animals were assigned to one of four treatment groups: porous HAp/Col impregnated with 50 µl of FGF‐2 at a concentration of 10 or 100 µg/ml (FGF10 or FGF100 group); porous HAp/Col with 50 µl of PBS (HAp/Col group); and no implantation (defect group). The defect areas were examined grossly and histologically. Subchondral bone regeneration was quantified 3, 6, 12, and 24 weeks after surgery. Abundant bone formation was observed in the HAp/Col implanted groups as compared to the defect group. The FGF10 group displayed not only the most abundant bone regeneration but also the most satisfactory cartilage regeneration, with cartilage presenting a hyaline‐like appearance. These findings suggest that porous HAp/Col with FGF‐2 augments the cartilage repair process. © 2009 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 28:677–686, 2010     

一种新型生物活性人工骨的制备及成骨活性的研究

目的：研制CPC/BMP复合人工骨,检测其成骨活性。方法：制备CPC/BMP及CPC骨块,扫描电子显微镜观察表面结构。用小鼠肌袋植入实验观察材料的成骨活性。结果：BMP在CPC中呈微球状均匀分布。CPC植入小鼠肌袋内不能诱导,CPC/BMP植入后1周有软骨细胞出现,2周有编织骨,4周以后小梁骨生成,16周出现成熟的板层骨。同时材料出现降解迹象。有机质含量、碱性磷酸酶浓度在CPC/BMP组出现升高,扫描电镜结果同样证实有新骨形成。结论：CPC/BMP生物活性人工骨可异位诱导成骨,可望成为新型的骨缺损修复材料。     

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.     

The effect of different mineral frames on ectopic bone formation in mouse hind leg muscles induced by native reindeer bone morphogenetic protein

Introduction Bone morphogenetic proteins (BMPs) require carrier material for slow release and framing material for osteoconduction.Materials and methods The effect of a frame on early bone formation induced by partially purified native reindeer BMP in composite implants containing 3 mg of BMP, type IV collagen and tricalcium phosphate (TCP/Col/BMP) or hydroxyapatite (HA/Col/BMP) or biphasic tricalcium phosphate-hydroxyapatite (TCP/HA/Col/BMP) or biocoral (NC/Col/BMP) was evaluated using a mouse hind leg muscle pouch model. Collagen with native reindeer BMP (Col/BMP) and corresponding implants without native reindeer BMP served as controls. Evaluation was done by incorporation of ⁴⁵Ca, radiographically and histologically 3 weeks after the implantation.Results None of the implants without native reindeer BMP were able to induce new bone visible on radiographs. The area of new bone formation in the Col/BMP (p=0.026) and TCP/HA/Col/BMP (p=0.012) groups was significantly greater than in the TCP/Col/BMP group. The optical density of the new bone area was significantly greater in the TCP/HA/Col/BMP group than in the TCP/Col/BMP (p=0.036) or Col/BMP (p=0.02) groups. ⁴⁵Ca incorporation was many times greater in all the groups containing native reindeer BMP than in the corresponding groups without BMP. In the Col/BMP (p=0.046) and TCP/HA/Col/BMP (p=0.046) groups, ⁴⁵Ca incorporation was significantly greater than in the TCP/Col/BMP group. No significant differences were found in any parameters between HA/Col/BMP and NC/Col/BMP groups and the other BMP-containing groups.Conclusions Hydroxyapatite, biocoral and biphasic tricalciumphosphate-hydroxyapatite are equally good as framing material for native reindeer BMP, while tricalciumphosphate is somewhat worse. Osteoinduction of native reindeer BMP works well with collagen alone.     

PLGA和胶原海绵复合BMP修复兔关节软骨缺损的对比研究

目的以PLGA和胶原海绵为载体，分别复合rhBMP-2，比较两者修复兔关节软骨缺损的效果。方法将PLGA和胶原海绵制成直径4mm、厚3mm的圆柱形，分别与0．5mgrhBMP-2复合。2月龄新西兰兔24只，雌雄不拘，体重1．8～2．3kg，平均2．0kg。于24只兔双侧股骨髁部制造直径4mm的缺损，其中18只动物右侧缺损处植入PLGA／rhBMP-2复合物（实验1组），左侧缺损处植入胶原海绵／rhBMP-2复合物（实验2组）；余6只动物（12个膝关节）缺损不作任何处理，作为空白对照组。术后4、12和24周取材，行大体及组织学观察，并根据关节软骨半定量评分标准进行组织学评分。结果大体及组织学观察：4周，实验1组缺损内被半透明组织填充；实验2组缺损未被新生组织填满，两组形成的软骨组织与周围界限明显，软骨细胞分布较均一，排列无方向性；对照组中形成少量纤维组织。12周，实验1组缺损内完全充填白色半透明新生软骨组织，与正常软骨界限不清；实验2组缺损内形成白色半透明软骨组织，与周围正常软骨界限仍可辨认；两组新生软骨厚度逐渐接近正常软骨厚度，表面细胞平行排列，深层细胞排列较紊乱，基质异染，软骨下骨及潮线恢复，与正常软骨连接良好；对照组缺损边缘及底部形成少量软骨细胞，分布不均匀，底部纤维组织不连续。24周，实验1组缺损内形成半透明新生软骨组织，与正常软骨界限消失；实验2组形成的新生软骨组织颜色、质地与12周接近；两组新生软骨与正常软骨厚度相近，表面平整，细胞排列均一，但较紊乱，基质异染，软骨下骨及潮线恢复，与正常软骨连接良好；对照组缺损底部形成一层纤维组织，不连续。组织学评分：实验1组、实验2组与对照组在各时间点比较，差异均有统计学意义（P〈0．01）；两个实验组12、24周与4周比较差异有统计学意义（P〈0．01），但12周和24周比较差异无统计学意义（P〉0．05）；同一时间点两实验组间比较差异无统计学意义（P〉0．05）。结论PLGA和胶原海绵与rhBMP-2复合均可修复关节软骨缺损。     

BMP‐2 delivered via sucrose acetate isobutyrate (SAIB) improves bone repair in a rat open fracture model

Human bone morphogenetic proteins (BMPs) are an alternative to bone graft for the treatment of high‐energy open fractures. The standard delivery system for BMP‐2 is a porous collagen sponge, but we have previously found that the biocompatible, high viscosity carrier, Sucrose acetate isobutyrate (SAIB) is an effective and potentially less invasive alternative. The efficacy of SAIB as a BMP‐2 delivery system was examined in an open fracture model featuring a femoral osteotomy with periosteal stripping in 9‐week‐old male Sprague Dawley rats. SAIB containing BMP‐2 (SAIB/BMP‐2) was delivered into the fracture site during surgery and an additional group was further co‐treated with zoledronic acid and hydroxyapatite nanoparticles (SAIB/BMP‐2/HA/ZA). These were compared to untreated fractures and SAIB carrier alone (negative controls), and BMP‐2 loaded collagen sponge (positive control). The rate of radiographic union and the biomechanical properties of the healed fractures were compared after 6–week. Untreated and SAIB‐treated fractures showed poor repair, with 53% and 64%, respectively, not bridged at 6 week. In contrast, collagen/BMP‐2, SAIB/BMP‐2, and SAIB/BMP‐2/HA/ZA showed significantly increased union (100%, 100%, and 94%, respectively, p < 0.05). Four‐point bend testing revealed that collagen/BMP‐2 and SAIB/BMP‐2/HA/ZA restored the strength of fractured femora to that of intact femora by 6 week, whereas untreated and SAIB remained less than intact controls by 60% and 67%, respectively (p < 0.05). Overall, the SAIB/BMP‐2/HA/ZA formulation was comparable to BMP‐2 infused collagen sponge in terms of promoting open fractures repair, but with the additional potential for less invasive delivery. © 2015 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 34:1168–1176, 2016.     

Reindeer BMP extract in the healing of critical-size bone defects in the radius of the rabbit

Background?Native BMP extracts from reindeer effectively induce ectopic new bone formation in vivo, but their bone healing properties have not yet been evaluated. We investigated the effect of reindeer BMP extracts on the healing of long bone defects.

Methods?The implants tested contained 5?mg or 10?mg of unsterilized BMP extract from reindeer and 10?mg of gamma-sterilized BMP extract administered with collagen carrier (Lyostypt, B. Braun, Germany). 70 μg of rhBMP-2 with collagen carrier (InductOs; Wyeth Europa) served as positive control, and collagen implants (Lyostypt) and untreated defects served as negative controls. New Zealand White rabbits with 1.5?cm of critical-size radius bone defects were used, with 8 weeks of follow-up.

Results?Radiographic analysis showed bone formation (BF) to be higher in all groups containing BMPs than in the untreated controls. BF was also higher in the rhBMP-2 group, and marginally higher in the group treated with 10?mg of unsterilized reindeer BMP extract (p = 0.06) as compared to the collagen controls. Bone union (BU) was better in the unsterilized BMP extract groups and rhBMP-2 group than in the untreated controls. BU was also better in the implants with 10?mg of unsterilized reindeer BMP extract and rhBMP-2 than in the collagen-treated implants. The mean area of new bone at the site of the defect proved to be higher in all implants containing BMP than in the untreated defects. It was also higher in the groups with 10?mg of unsterilized reindeer BMP extract and rhBMP-2 than in the collagen-treated controls. Mechanical tests showed torsional stiffness of the bones to be higher in the group with 10?mg of unsterilized BMP extract than in the collagen group. The mean cross-sectional bone area measured by pQCT densitometry was higher in the rhBMP-2 group than in the collagen group. The mean bone density at the defect area was higher in the group with 10?mg of unsterilized BMP than in the rhBMP-2 group.

Interpretation?We conclude that both reindeer BMP extract and rhBMP-2 induced improved healing of the rabbit radius bone defects at the doses used. Gamma sterilization of reindeer BMP extract reduced osteoinductivity slightly, but not significantly.