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。     

rhBMP-2对兔肩袖损伤重建术后腱-骨愈合的组织学及生物力学研究

[目的]评价重组人骨形态发生蛋白2(rhBMP-2)对兔肩袖损伤重建术后的修复效果。[方法]48只8个月龄雄性新西兰大白兔双侧冈上肌腱行急性损伤后重建其在肱骨大结节上的止点,术后随机分为3组,每组16只,32侧,实验组在腱骨界面注射以纤维蛋白胶为载体的rhBMP-2;实验对照组仅填充纤维蛋白胶;空白对照组术后不给予任何干预。于术后8周取出各组8侧肩袖标本行组织形态学检查,分别于术后2、4、8周分批随机取出各组8侧肩袖标本行生物力学测试。[结果]组织学分析表明,术后8周实验组腱骨界面出现有明显的Sharpey′s纤维连接,出现类似直接止点的四层结构。而实验对照组及空白对照组腱骨界面以结缔组织为主,出现部分Sharpey′s纤维结构及新骨形成。生物力学测试显示,术后各组腱骨界面的抗拉强度及刚度均随着康复时间的延长而不断增强,而且各个时间点实验组腱骨界面的抗拉强度及刚度均显著高于实验对照组及空白对照组,差异有统计学意义(P0.01)。[结论]rh-BMP-2可以在术后明显增强腱骨界面的抗拉强度及刚度,促进腱骨界面形成类似直接止点的特有结构,可很好的修复肩袖损伤。     

Downregulation of the expression of bone morphogenetic protein 7 in experimental pyelonephritis

Bone morphogenetic protein 7 (BMP 7) is a member of the transforming growth factor (TGF) beta superfamily and is involved in regeneration, repair, and development of specific tissues, for example kidney, gut, lens, and skeleton. BMP 7 has emerged as a renotrophic factor and experimental studies have shown its protective role against fibrotic processes. Tubulointerstitial changes are present in the pyelonephritic kidney which progresses to fibrosis. Renal fibrosis may lead to significant morbidity in the form of hypertension, proteinuria, and loss of renal function. The objective of this study was to investigate BMP 7 expression in experimental acute and chronic pyelonephritis models. Eighteen Wistar rats were injected with 0.1 mL solution containing E. coli ATCC 25922 10¹⁰ cfu mL^–1 into left renal medullae. Six rats were used as a sham group and were given 0.1 mL 0.9% NaCl. Pyelonephritic rats were sacrificed 24 h (group I, n=6), 1 week (group II, n=6), and 6 weeks (group III, n=6) after E. coli injection. Serum creatinine levels were analyzed. Renal tissues were studied histopathologically by use of hematoxylin and eosin and scored for diagnosis of pyelonephritis. BMP 7 expression was studied semiquantitatively by immunohistochemical staining. Acute (group I) and chronic (group II and group III) pyelonephritic histopathological changes were observed in experimental pyelonephritic groups. A gradual decrease in BMP 7 expression was observed in the tubulointerstitial and tubular area of the pyelonephritic kidneys, mildest in the acute pyelonephritic group and most severe in the chronic pyelonephritic 6th week group. A statistically significant difference was observed between tubulointerstitial BMP 7 expression by groups I and III (P=0.017) and by groups III and IV (P=0.000). Tubular BMP 7 expression was statistically significantly different between groups II and IV (P=0.009) and between groups III and IV (P=0.002). The data imply that BMP 7 has a major role in chronic pyelonephritis. Tubulointerstitial and tubular BMP 7 expression also had a significant negative correlation with fibrosis, tubular, atrophy, and vascular changes. Serum creatinine levels of the study group were all normal. We conclude that the decrease in renal BMP 7 expression in experimental chronic pyelonephritis is one of the factors responsible for fibrotic changes in persistent renal damage.     

Distinct roles of bone morphogenetic proteins in osteogenic differentiation of mesenchymal stem cells.

Efficacious bone regeneration could revolutionize the clinical management of many bone and musculoskeletal disorders. Bone morphogenetic proteins (BMPs) can regulate the differentiation of mesenchymal stem cells into cartilage, bone, tendon/ligament, and fat lineages. Early data documented the osteogenic potential of rhBMP2 and rhBMP7/OP-1. However, prior to this work that summarized several of our recent studies, no comprehensive analysis had been undertaken to characterize relative osteogenic activity of all BMPs. Using recombinant adenoviruses expressing 14 BMPs, we have demonstrated that, besides BMP2 and BMP7, BMP6 and BMP9 exhibit the highest osteogenic activity both in vitro and in vivo. We further demonstrated that several BMPs may exert synergistic effect on osteogenic differentiation, and that osteogenic BMPs produce a distinct set of molecular fingerprints during osteogenic differentiation. The reported work should expand our current understanding of BMP functions during osteogenic differentiation. It is conceivable that osteogenic BMPs (i.e., BMP2, 4, 6, 7, and 9) may be used to formulate synergistic pairs among themselves and/or with other less osteogenic BMPs for efficacious bone regeneration in clinical settings.     

骨形成蛋白-脱钙骨基质颗粒-骨水泥复合材料修复狗股骨微波灭活骨缺损后的长期观察

目的 :探讨骨形成蛋白 (BMP) 脱钙骨基质颗粒 (DBM ) -骨水泥 (BC)复合材料修复狗股骨微波灭活骨缺损后的生物学过程和归宿。方法 :对BMP DBM BC复合材料进行综合评价和植入动物体内进行了初步观察后 ,进一步对实验动物进行X射线照相、99Tcm MDP骨显像、大体标本观察、组织学观察、土霉素荧光标记和印度墨汁灌注染色观察。结果 :X射线照相 :术后随时间延长复合材料与宿主骨边界模糊程度增加。99Tcm MDP骨显像 :术后 18个月时实验侧同位素浓集仍略高于正常骨组织。大体标本 :初期可见复合材料处有充血炎性反应 ,至 9个月时炎性反应消退。组织学观察 :随时间延长 ,材料内部新生骨增多 ,“生物铆定”增强。土霉素荧光标记 :复合材料内大部分DBM所在部位呈现较强的黄色荧光。印度墨汁灌注 :复合材料内有大量的墨染血管。结论 :BMP DBM BC复合材料修复狗股骨微波灭活骨缺损后能够诱导新骨形成 ,复合材料与宿主骨最终形成“生物铆定”。     

重组人骨形态发生蛋白-2诱发黄韧带骨化的实验模型

目的 :建立脊柱黄韧带骨化的实验模型。方法 :以中国大白兔为实验对象 ,采用重组人骨形态发生蛋白 2(rhBMP 2 )作为诱导物 ,分别将rhBMP 2 /明胶海绵植入双侧黄韧带腹侧的硬膜外腔 (E1组 ) ,或直接将rhBMP 2注射到双侧的黄韧带内 (E2组 ) ,每一侧植入物中含rhBMP 2 10 0 μg。设立相应的对照组 (C1组、C2组 )。对手术节段进行X线、CT扫描及病理组织学检查。结果 :脊柱CT扫描发现E1组 4周时手术节段后正中椎板前方出现结节状高密度增高影 ,8周时密度进一步增高。病理组织学检查发现E1组手术节段韧带细胞分化为软骨细胞并发生骨化。E2组仅见黄韧带轻度增生肥厚及少量散在的软骨细胞。C1组和C2组黄韧带组织均未见明显异常改变。结论 :rhBMP 2可诱导脊柱黄韧带骨化 ;明胶海绵可作为BMP诱导成骨的载体。     

Expression of osterix inhibits bone morphogenetic protein-induced chondrogenic differentiation of mesenchymal progenitor cells

Osteoblasts and chondrocytes arise from common bipotential mesenchymal progenitor cells. Although the differentiation of these two cell lineages can be induced by treatment with bone morphogenetic proteins (BMPs), the responses of mesenchymal progenitors to BMP differ from cell line to cell line. Here we demonstrate that C3H/10T1/2 cells preferred chondrogenic differentiation, primary bone marrow stroma cells (MSCs) tended to convert to osteoblasts, and ST-2 cells differentiated into both the osteoblastic and chondrocytic lineages simultaneously, suggesting that a molecular switch functions to select cell fate. Osterix, the secondary master regulator of osteoblastogenesis, was induced by BMP at high and low levels in MSCs and ST-2 cells, respectively; in contrast, C3H/10T1/2 cells demonstrated only faint expression. As osterix has been suggested as a negative regulator of chondrogenesis, we hypothesized that the intense chondrocyte differentiation of C3H/10T1/2 cells may have resulted from an absence of osterix. We therefore restored osterix gene expression in C3H/10T1/2 cells using an adenovirus vector. Following BMP treatment, infection with an osterix-encoding virus dramatically inhibited the chondrocytic differentiation of C3H/10T1/2 cells, resulting instead in prominent osteoblast differentiation. These results indicate the chondrogenic potential of C3H/10T1/2 cells was abrogated by osterix expression. Chondrocyte differentiation of MSCs, however, was not enhanced by silencing the osterix gene using lentivirus-mediated shRNA, despite successful suppression of osteoblast differentiation. These results suggest that the low levels of osterix expression remaining after knockdown are sufficient to block chondrogenesis, whereas higher expression may be required to promote osteoblastic differentiation.     

骨髓间充质干细胞在丝素蛋白/骨形态发生蛋白中的生物学行为观察

目的 观察丝素蛋白与牛骨形态发生蛋白的载体系统与骨髓间充质干细胞的体外相容性.方法 将多孔丝素蛋白修剪成20×10×5 mm长方体状物,置入6孔板中;将粉状bBMP用PBS液溶解制成2mg/ml混悬液,0.5ml无菌移液管滴定于多孔丝素蛋白上(0.5毫升/个).骨髓间充质干细胞以107/ml接种到载体系统上,复合4小时后加入生长液继续体外培养,每1、4、8天时进行碱性磷酸酶、矿化结节染色观察以及扫描电镜和激光共聚焦显微镜观察;另外同时设立单纯丝素蛋白与细胞复合培养对照组进行观察.结果 倒置显微镜、扫描电镜和共聚焦显微镜下观察到细胞在两种材料上生长形态、活性正常,体外培养24小时时细胞已贴附材料上,呈匍匐状伸展,以后逐渐融合生长,8天时大量细胞成片状攀附在材料表面,并分泌有大量的网状细胞外基质,实验组可见结节状基质生成;碱性磷酸酶、矿化结节染色观察发现实验组细胞碱性磷酸酶和矿化结节染色阳性反应,对照组阴性.结论 丝素蛋白是一种良好细胞外基质材料,也是一种良好生长因子的缓释载体.     

Increased accumulation of superficial zone protein (SZP) in articular cartilage in response to bone morphogenetic protein-7 and growth factors.

The purpose of this study was to investigate the role of bone morphogenetic proteins (BMPs), such as BMP-7, growth factors, and cytokines, in the accumulation of superficial zone protein (SZP) in bovine articular cartilage. Calf superficial articular cartilage discs and chondrocytes were obtained for explant and monolayer culture systems, respectively. Dose- and time-dependent actions of BMP-7 on SZP accumulation were investigated in both explant and monolayer culture systems. In addition, actions of various morphogens and growth factors [BMP-2, BMP-4, fibroblast growth factor 2 (FGF-2), insulin-like growth factor 1 (IGF-1), platelet-derived growth factor (PDGF), and transforming growth factor beta (TGF-beta1)], and cytokines [interleukin (IL)-1alpha, IL-1beta, and tumor necrosis factor (TNF-alpha)] alone, and in combination with BMP-7, on SZP accumulation were investigated in monolayer culture systems. SZP accumulation was quantified in both the cartilage and the medium using SDS-PAGE and subsequent immunoblotting. In both explant and monolayer cultures, BMP-7 increased SZP accumulation in a dose- and time-dependent fashion (p < 0.05). Furthermore, SZP accumulation was significantly increased in monolayer cultures by FGF-2, IGF-1, PDGF, and TGF-beta1 (p < 0.05). Both IL-1alpha and TNF-alpha significantly reduced SZP accumulation (p < 0.05). The inhibition of SZP accumulation by TNF-alpha was partially alleviated by concurrent treatment with BMP-7. The results of this investigation provide novel insights into the role of morphogens, especially BMP-7, growth factors, and cytokines in the accumulation of SZP in articular cartilage. This information has clinical implications because stimulation of SZP may ameliorate the pathology of joint function in arthritis. Furthermore, tissue engineering approaches to articular cartilage may depend on the optimal synthesis and assembly of SZP in the superficial zone to ensure functional tissue architecture.     

The effect of the delivery carrier on the quality of bone formed via bone morphogenetic protein-2

Bone morphogenetic protein-2 (BMP-2) can induce bone generation in vivo. Although many studies have demonstrated an increased quantity of regenerated bone after the delivery of BMP-2 using various carriers, little is known about the effect of the carrier type on the quality of the regenerated bone. In this study, we compared the quality of regenerated bone when BMP-2 was delivered with either β-tricalcium phosphate (β-TCP) or heparin-conjugated fibrin (HCF), both of which are shown to be excellent carriers for BMP-2. The profile of the release of BMP-2 was not significantly different between the delivery carriers. However, the alkaline phosphate activity of cultured osteoblasts was significantly higher when BMP-2 was delivered using HCF than when BMP-2 was delivered using β-TCP. To evaluate the quality of the regenerated bone, both types of BMP-2 carriers were implanted into critical-sized calvarial defects in mice. Eight weeks after implantation, the regenerated bone was examined by histomorphometry. Importantly, the treatment using HCF + BMP-2 and β-TCP + BMP-2 resulted in similar bone formation areas. However, the treatment using HCF + BMP-2 resulted in significantly higher bone density than the treatment using β-TCP + BMP-2. This study shows that a BMP-2 delivery carrier can modulate the quality of bone regenerated via BMP-2 delivery.     

Recombinant human bone morphogenetic protein 2 enhances bone healing in an experimental model of fractures at risk of non-union

INTRODUCTION: Identification of patients at risk of developing non-union and institution of procedures preventing non-union could be attractive in routine fracture management. We investigated whether recombinant human bone morphogenetic protein (rhBMP-2) delivered in a hyaluronic acid carrier could prevent non-union development in an experimental non-union model, which simulates the clinical situation of open mid-tibial fractures. METHODS: Sixteen rabbits underwent a standard non-union operation comprising mid-tibial osteotomy, excision of periosteum and endosteum, and plate fixation. Before closure of the wound eight rabbits received interfragmentary deposition of 200 microg rhBMP-2 delivered in a hyaluronan gel carrier, and eight rabbits received gel carrier alone. RESULTS: After 7 weeks, torsional failure moment of the osteotomy and energy absorbed at failure, macroscopic and radiographic appearance, callus area, and interfragmentary bone volume fraction confirmed that rhBMP-2 delivery significantly improved bone healing. Blood flow at the osteotomy site, measured using radiolabelled microspheres, was not higher in the united osteotomies than in non-united osteotomies. DISCUSSION: rhBMP-2 delivered in a hyaluronic acid carrier-induced formation of competent bone in an experimental model of compromised healing. We, therefore, propose interfragmentary deposition of rhBMP-2 delivered in a hyaluronic acid carrier to patients encountering fractures at risk of non-union or delayed union.     

Expression of bone morphogenetic proteins and rat distal-less homolog genes following rat femoral fracture

Expression of the genes encoding bone morphogenetic proteins (BMPs), BMP type IA receptor (BMPR-1A), and rat distal-less homolog (rDlx) was studied in bone, callus, and the surrounding soft tissue following rat femoral closed fracture, using RT-PCR-based techniques. Before fracture, the genes encoding BMP-5, BMP-6, and BMPR-1A were found to be expressed in both bone and the surrounding soft tissue, whereas the BMP-2 gene was expressed only in bone and BMP-7 was not expressed in either tissue. Expression of these genes was unaffected by fracture. The gene encoding BMP-4 was also expressed in both bone and the surrounding soft tissue before fracture. Moreover, although unchanged in bone, 6 h after fracture BMP-4 expression was increased tenfold in the surrounding soft tissue. The increased BMP-4 expression was transient and returned to prefracture levels within 72 h. Expression of rDlx was also increased in bone after fracture, but at later times than were observed with BMP-4: elevated rDlx expression was detected after 48 h and persisted for 30 days or more. No expression of rDlx was observed in the surrounding soft tissue before or after fracture. These findings indicate that BMP-4 and rDlx are selectively expressed following femoral fracture in the rat, and also suggest that they are involved in the formation of the callus at an early point during the postfracture healing of bone. Received: March 26, 1999 / Accepted: Sept. 20, 1999     

Enhancement of recombinant human bone morphogenetic protein-2 (rhBMP-2)-induced new bone formation by concurrent treatment with parathyroid hormone and a phosphodiesterase inhibitor,pentoxifylline

We investigated the enhancement of new bone |formation elicited ectopically by recombinant human bone morphogenetic protein-2 (rhBMP-2), using parathyroid hormone (PTH) and a phosphodiesterase inhibitor (PDEi), pentoxifylline (PTX), in an animal model. Collagen sponge sheet discs containing rhBMP were implanted onto the back muscles of mice. PTX alone (200mg/kg body weight [BW]), PTH(1–34) (10µg/kg BW), PTX plus PTH (200mg/kg BW and 10µg/kg BW, respectively), or vehicle (control) were injected subcutaneously daily for 3 weeks after implantation. At the end of this period, rhBMP-2-induced ectopic ossicles were harvested from each group of animals. Ossicles from the PTX-treated group were significantly larger in size, with unchanged bone mineral density (BMD), as compared with the ossicles from the controls. In contrast, the ossicles from the PTH-treated group had significantly higher BMD, but showed no difference in size when compared with those from the control animals. The ossicles of the PTX + PTH treatment group were significantly larger than those of the control and PTH treatment groups. In addition, the BMD of the harvested tissues from the PTX + PTH treatment group was signifi-cantly higher than that of tissues from the control and PTX treatment groups. Although the calcium content of ossicles was significantly higher in the PTX-, PTH-, and PTX + PTH-treated groups than in the control group, the Ca content of ossicles from the PTH + PTX-treated group was highest (two times that of controls), followed by the PTH- and PTX-treated groups.     

血管内皮细胞生长因子165及骨形态发生蛋白2促进兔骨缺损部位再血管化的研究

目的为大段骨缺损修复过程的血管再生探索一条可行的途径。方法成年大耳白兔30只,随机分为五组。右前肢为实验肢体,缺损长度约15 mm。第1～4组都采用胶原膜引导,结合纳米羟基磷灰石／聚酰胺骨水泥,不同的是在第1组中,复合血管内皮生长因子165(VEGF165)及骨形态发生蛋白2(BMP2)质粒;第2组复合VEGF165质粒;第3组复合BMP2质粒;第4组不复合任何质粒;第5组仅制作动物模型而不做任何处理,作为空白对照组。结果术后2周,核素断层摄影(ECT)结果显示第1组骨缺损修复后的局部血流量高于第2、3、4组(P<0．05);术后4周,X线片示第1组骨缺损处的骨痂明显增多;SEM显示在正常骨与工程骨交界处可见新生的骨小梁结构以及成骨细胞的附着;ECT结果照示第1组骨缺损的局部血流量高于第2、3、4组(P<0．01);第2组骨缺损的局部血流量高于第3、4组(P<0．01);术后8周,SEM显示第1组工程骨表面成骨细胞的附着多于其它各组, ECT结果显示与术后4周相同。结论在骨缺损的局部,联合应用表达VEGF165和BMP2质粒可以促进骨缺损局部的血液供应;附着质粒DNA的纳米羟基磷灰石／聚酰胺及引导性胶原膜在大段骨缺损局部的联合应用有助于新骨的形成。     

Recombinant human bone morphogenetic protein is a valid alternative to autologous bone graft for long bone non-unions: a systematic review and meta-analysis

ObjectiveTo compare the efficacy of recombinant human bone morphogenetic proteins (rhBMPs) and autologous bone graft (ABG) on the healing of long bone non-union.MethodsA systematic literature search was conducted on PubMed, Web of Science, Cochrane Library, and CNKI up to December 2021. Two authors independently screened the studies, extracted data, and assessed the quality of the trials. A Meta-analysis was performed using state software (version 12.0).ResultsA total of 14 studies were included in this meta-analysis. Overall, there was no significant difference between the rhBMPs group and the ABG group in terms of healing rate (RR = 1.04, 95% CI = 0.96–1.12, p = 0.365) and healing time (SMD = −0.31, 95% CI = −0.76–0.14, p = 0.175). Subgroup analysis showed rhBMPs lead to higher healing rates (RR = 1.35, 95% CI = 1.17–1.56, p < 0.001), and shorter healing time (SMD = −0.65, 95% CI = −1.08 to −0.22, p = 0.003) in the subgroup of moderate-quality studies. Sensitivity analysis proved that our conclusions were relatively robust. No significant publication bias was recognized in all studies (Begg’s test, p = 0.193; Egger's test, p = 0.307).ConclusionsRhBMPs or combined with allografts bone, inorganic bone was a valid alternative to ABG for the treatment of long bone non-union.     

Effects of OP-1 and PTH in a new experimental model for the study of metaphyseal bone healing.

The purpose of this study was to establish a reliable model of metaphyseal bone healing and to use this model to investigate the effect of recombinant human osteogenic protein 1 (rhOP-1; BMP-7) and parathyroid hormone fragment (PTH 1-34) on healing. A wedge-shaped osteotomy was created in the distal tibia of 16-week-old female New Zealand White rabbits (n = 20) and was bridged with a custom-made external fixator. Five experimental groups of four animals each were investigated. In groups 1-4 the osteotomy gap was filled with tricalcium phosphate (TCP), and the gap was left unfilled in group 5 ("normal healing"). In group 1, 200 microg OP-1 was mixed in with the TCP. Groups 2 and 3 received daily subcutaneous injections of 10 and 40 microg/kg PTH, respectively, beginning on postoperative day 1. Radiographs were taken weekly. Following sacrifice on postoperative day 28, peripheral quantitative computed tomography (pQCT), histology, and mechanical testing (axial compression and torsion) were performed. Only one animal failed to complete the full 4-week time course, and no infections were encountered. Bone healing occurred in all animals. OP-1 stimulated bone formation locally, while the lower dose of PTH enhanced bone formation systemically (p < 0.05). Tibiae treated with OP-1 exhibited higher torsional strength (p = 0.04) than those in the normal healing group. These results indicate that a reliable and reproducible surgical model of metaphyseal healing has been established. In addition, differences in systemic versus local effects of PTH and OP-1 in accelerating metaphyseal fracture healing were found.     

Intramuscular bone induction by human recombinant bone morphogenetic protein-2 with beta-tricalcium phosphate as a carrier: in vivo bone banking for muscle-pedicle autograft

 An ideal replacement for bone defects is auto-bone tissue, of which there is an ample supply with the required form and with vascularity. Our strategy for generating such bone tissue is as follows. First, bone tissue is induced in muscle by bone morphogenetic protein-2 (BMP-2) with beta-tricalcium phosphate as a carrier to maintain its form in the muscle. Second, the induced bone in the muscle pedicle is grafted to the bone defect to maintain vascularity. In the first experiment, 50 μg of recombinant human BMP-2 (rhBMP-2) was inoculated into the hip abductor muscle of rabbits with beta-tricalcium phosphate under anesthesia. Five weeks after the operation, intramuscular bone formation was observed in all of the samples, and the form and size of the induced bone tissue were identical to those of the carrier. Ten weeks after the operation, the induced bone was partly absorbed. In the second experiment, 50 μg of rhBMP-2 was inoculated in the same manner as previously. Five weeks after the operation, the muscle tissue around the induced bone was incised, leaving just the proximal part as a pedicle. Two or four weeks after the second operation, the induced bone tissue had rich vascularity and no empty lacunae. This indicates the possibility of in vivo bone banking to enable morphologically controlled and vascularized auto-bone grafts. Received: December 21, 2001 / Accepted: March 28, 2002     

Enhancement of recombinant human BMP‐7 bone formation with bmp binding peptide in a rodent femoral defect model

Bone morphogenetic binding peptide (BBP) is an 18.5 kDa fragment of a bone matrix protein peptide. A rat femoral defect model was used to test the effect of BBP combined with recombinant human bone morphogenetic protein‐7 (rhBMP‐7) to induced bone healing. Two doses of BBP (500 and 1000 µg) were tested with two doses of rhBMP‐7 (2 and 5 µg), and the results were compared with a positive control (10 µg rhBMP‐7). Bone healing was evaluated by radiology, manual palpation, microcomputed tomography, and histology. The high dose of 10 µg of rhBMP‐7 resulted in a consistent 100% bone union rate and a mature histological appearance on histology, and was used as a positive control. When 1000 µg of BBP was combined with lower doses of BMP‐7 (2 µg rhBMP‐7 or 5 µg rhBMP‐7) significant differences were seen in radiographic scores, manual palpation, and bone volume, when compared to 2 µg rhBMP‐7 or 5 µg rhBMP‐7 alone. The combination of 1000 µg of BBP and 5 µg rhBMP‐7 also achieved 100% fusion rate, induced a larger amount of bone formation, and yielded similar maturity of bone marrow when compared with the high dosage 10 µg rhBMP‐7 group. This study demonstrated that when combined together, BBP can enhance the bone healing of rhBMP‐7. Improved healing imparted by the addition of BBP may result in lesser amounts of rhBMP‐7 needed to achieve union in the clinical setting. © 2010 OrthopaedicResearchSociety.PublishedbyWileyPeriodicals, Inc.JOrthopRes29:753–759,2011     

In vivo molecular imaging of adenoviral versus lentiviral gene therapy in two bone formation models.

Regional gene therapy techniques are promising methods to enhance bone formation in large bone defects that would be difficult to treat with allograft or autograft bone stock. In this study, we compared in vivo temporal expression patterns of adenoviral- and lentiviral-mediated gene therapy in two bone formation models. Primary rat bone marrow cells (RBMC) were transduced with lentiviral or adenoviral vectors containing luciferase (Luc) or BMP-2 cDNA, or cotransduced with vectors containing Luc and bone morphogenetic protein 2 (BMP-2). In vitro protein production was determined with luciferase assay or ELISA (for BMP-2 production) weekly for 12 weeks. Two bone formation models were used -- a hind limb muscle pouch or radial defect -- in SCID mice. A cooled charged-coupled device (CCD) camera was used to image in vivo luciferase expression weekly for 12 weeks. In vitro, adenoviral expression of BMP-2 and luciferase was detected by ELISA or luciferase assay, respectively, for 4 weeks. Lentiviral expression of BMP-2 and luciferase was sustained in culture for 3 months. Using the CCD camera, we found that adenoviral vectors expressed luciferase expression for up to 21 days, but lentiviral vectors expressed target gene expression for 3 months in vivo in both bone formation models. There was no detectable difference in the amount of bone formed between the adenoviral and lentiviral groups. Lentiviral-mediated delivery of BMP-2 can induce long term in vitro and in vivo gene expression, which may be beneficial when developing tissue engineering strategies to heal large bone defects or defects with a compromised biologic environment.