Relationship of bone morphogenetic protein expression during osteoblast differentiation to wild type p53

We have previously shown p53 to have a specific role in osteoblast differentiation by its ability to regulate expression of certain bone specific proteins. In this study, we show mineralized matrix formation in vivo to be directly related to the presence of wild type p53 in osteoblastic osteosarcoma cells. In order to further understand the importance of p53 in differentiation, we investigated the relationship between p53 and Bone Morphogenetic Proteins (BMPs) (BMP 1, 2, 3A, 3B (GDF-10), 4, 5, 6, 7, 8A and 8B) during osteoblast differentiation. The expression of several BMPs were tested using RNase Protection Assay in differentiating ROS17/2.8 osteoblastic osteosarcoma cells. The expression of BMPs 1, 2, 3a, 3b and 7 showed time dependent modulation during in vitro differentiation. In order to determine if p53 has a role in this process, we used a murine osteosarcoma cell line stably expressing a temperature sensitive p53. Cells were exposed to ascorbic acid and glycerophosphates to hasten in vitro osteoblast differentiation and maintained either at 32 or 37 degrees C for expression of the wild type or mutant p53 phenotype. The expression of BMP-2, BMP-4 and BMP-7 were modulated in a p53 dependent fashion. We were able to confirm the p53 dependency of BMP-2 independently by RT-PCR. While BMP-2 expression was evident in the presence of both wild type and mutant p53, regulated expression was seen only in cells expressing wild type p53. Transient over expression of wild type p53 did not result in the same BMP-2 response as stable expression showing that the presence of p53 may be important for an orderly development of osteoblast differentiation rather than a direct effect on gene expression. The functional relationship between p53 and these bone specific markers is discussed.     

复合BMP同种异体半关节异位植入的免疫研究

目的探讨B M P骨诱导与免疫反应之间的关系。方法将复合B M P5m g的冻干异体半关节植入兔腰大肌肌袋内,并采用原位杂交技术对术后不同时间各组IL-1和IL-6m R N A表达强度进行比较。结果骨移植术后,实验组较对照组成骨作用增强。植骨局部在移植术后第4周起即可见IL-1和IL-6m R N A的阳性表达,实验组与对照组植骨局部均可见IL-1和IL-6m R N A的阳性表达,不同时间点各组间IL-1和IL-6m R-N A表达强度无明显差异。结论B M P作为外源性介质,在异位发挥诱导成骨作用的同时并不加重免疫排斥反应。     

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.     

骨缺损对脂肪源干细胞骨形态发生蛋白信号通路相关分子的影响

目的探讨脂肪源干细胞(ADSCs)是否存在骨形态发生蛋白(BMP)信号通路,以及骨缺损对大鼠内源性ADSCs中BMP信号通路分子表达的影响。 方法取30只Wistar大鼠随机分为空白对照组（A组5只）和实验组（B组25只）。A组不做处理,直接取腹股沟脂肪垫进行ADSCs培养。B组制造骨缺损模型,分别于造模后1、3、7、14、21d取腹股沟脂肪组织进行ADSCs培养,每一时间点取5只大鼠。原代培养7d,提取ADSCs总RNA,采用实时定量逆转录聚合酶链反应技术检测BMP受体(BMPR)和Smads的mRNA表达变化。实验数据用95％可信区间表示。 结果实验组骨缺损后1、3、7、14、21 d BMPR1a表达量均较对照组明显增加,差异有统计学意义(P＜0.05);骨缺损后7d表达量(2.532,2.552)增加达高峰。实验组骨缺损后1、3、7、14 d BMPRIb表达量较对照组明显增加,差异有统计学意义(P＜0.05);骨缺损后14 d表达量(6.628,6.648)增加达高峰。实验组骨缺损3、7、14、21 dBMPR2、Smad5、Smad8表达量较对照组明显增加,差异均有统计学意义(p＜0.05);骨缺损后14dBMPR2、Smad5表达量(3.538,3.658;8.055,8.149)增加达高峰,骨缺损后3 d Smad8表达量(3.657,3.759)增加达高峰。实验组骨缺损后7、14、21 d Smad1表达量较对照组明显增加,差异均有统计学意义(P＜0.05);骨缺损后7d表达量(3.641,3.771)增加达高峰。 结论 ADSCs表达BMPR,可能存在BMP信号通路,骨缺损可引起大鼠ADSCs中BMP信号通路相关分子的表达增加。     

人工骨腰椎后外侧融合过程中BMP基因的表达

目的研究利用人工骨做脊柱后外侧融合时局部BMP的表达情况,探讨人工骨在此过程中是否具备骨诱导作用。方法选用36只健康成年雄性新西兰白兔,制作L4、L5双侧横突间植骨融合模型,一侧采用磷酸钙人工骨(CPC),另一侧采用自体骨作为对照。按术后动物不同处死时间(0、2、4d、1、2、3、4、5、6、10周、6、10个月)随机平分为12组。采用RTPCR法检测融合组织BMP2mRNA、BMP4mRNA的表达水平。结果CPC材料内部在融合的各个时间段均未检测到BMP2mRNA和BMP4mRNA的表达。与紧密结合的植骨床及融合交界面中BMP2mRNA和BMP4mRNA的表达水平随时间变化均没有明显的增高,与自体骨相比,CPC融合过程中不同时间段内BMP2mRNA、BMP4mRNA的表达水平均明显低于自体骨(P<0.05)。结论单纯利用CPC做脊柱融合时,局部缺乏BMP的有效表达,可能导致融合失败率的增加。     

Interaction of bone morphogenetic proteins with cells of the osteoclast lineage: review of the existing evidence

The present review evaluates the existing scientific proofs of this supplementary role of the BMPs and summarises its clinical implications. Bone regeneration is a process consisting of bone formation and bone resorption, two different but closely coupling pathways, which in most circumstances proceed simultaneously. Plenty of evidence has also characterised the bone morphogenetic proteins (BMPs) as inducing factors of bone formation. However, there is also evidence that these multifunctioning proteins affect bone resorption and the osteoclast homeostasis utilising various pathways. The present review evaluates the existing scientific evidence of this supplementary role of the BMPs, and summarises its clinical implications. Note: This work is attributed to the Academic Unit, Trauma and Orthopaedic Surgery, Clarendon Wing, Leeds General Infirmary, Great George Street, Leeds, LS1 3EX, UK.     

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     

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.     

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.     

负载人骨形态发生蛋白-7基因的兔脂肪干细胞向成骨细胞分化的实验研究

目的 探讨人骨形态发生蛋白-7(hBMP-7)基因修饰对兔脂肪干细胞(ADSCs)成骨能力的影响. 方法 原代培养兔脂肪干细胞,免疫组织化学方法检测细胞表面抗原CD44、CD49d、CD106的表达,对细胞进行鉴定;阳离子脂质体介导hBMP-7基因转染ADSCs,绿色荧光蛋白表达观察转染效率、逆转录-聚合酶链反应(RT-PCR)检测目的基因hBMP-7的表达;ALP定量测定,Western blot检测Ⅰ型胶原、骨钙素的表达,以评价转基因ADSCs向成骨细胞分化的情况. 结果 从兔脂肪组织中分离出来的ADSCs CD44、CD49d表达呈阳性,CD106表达呈阴性.RT-PCR检测筛选后的ADSCs稳定表达hBMP-7.转染后7、10、14 d ALP定量测定转染组明显高于未转染组,差异有统计学意义(P<0.05);成骨标志物Ⅰ型胶原、骨钙素的表达转染组明显高于未转染组.结论 从脂肪组织中分离出的ADSCs是一种较好的组织工程种子细胞.hBMP-7重组质粒转染ADSCs后表达目的蛋白,并诱导ADSCs向成骨细胞分化.     

转染BMP-2基因的兔BMSCs种植PLA/PCL支架体外构建组织工程骨

目的:用腺病毒载体将人骨形态发生蛋白-2(BMP-2)基因导入兔骨髓基质干细胞(BMSCs),种植PLA/PCL(聚乳酸/聚己内酯)支架体外构建组织工程骨.方法:蛋白印迹法检测转染后细胞BMP-2的表达,流式细胞仪和ALP活性检测分析基因转染对细胞增殖分化的影响.然后将转染后细胞接种到PLA/PCL支架上,扫描电镜观察细胞贴附、生长状况.结果:转染后,BMSCs表达BMP-2,S期细胞比例和ALP活性明显增高.扫描电镜见转染细胞分布均匀,伸展良好.结论:BMP-2基因转染BMSCs,可促进细胞增殖分化.转染后细胞在PLA/PCL支架上生长良好,BMP-2基因治疗的组织工程骨构建成功.     

Morphometric and histological analysis of low-power laser influence on bone morphogenetic protein in bone defects repair

Bone morphogenetic proteins (BMPs) are secreted signaling molecules belonging to the transforming growth factor-β (TGF-β) superfamily. The objective of this study was to determine how gallium–aluminum–arsenium (GaAlAs) 650 nm laser influenced the action of BMPs on bone defects created in rat femurs. The sample consisted of 24 male albino Wistar rats. Group 1 was composed of rats with bone defects filled with bone-inducing substance, with the application of low-power laser. Group 2 contained rats with bone defects filled with a bone-inducing substance, without the application of low-power laser. Group 3 rats had bone defects not filled with a bone-inducing substance, with the application of low-power laser. Group 4 rats had bone defects and no treatment (control group). A bone defect was produced with drills. In groups 1 and 2 the defects were filled with a bone-inducing substance. The animals were treated with GaAlAs (50 mW) laser, energy density 4J/cm², for 80 ss on a 1 cm² area. Groups 2 and 4 were used as control. Bone samples were removed for histological procedures and morphometric analysis on the 7th, 14th and 21st days after surgery. Results obtained were subjected to statistical analysis. Rejection level for the null hypothesis was 0.05. Statistical differences were found in the comparison between group 1 (G1), G2, G3 and G4 [analysis of variance (ANOVA); P < 0.0134]. There was a statistically significant correlation between groups 1 and 4 (P < 0.01). The results of other correlations by Tukey’s post-hoc test were: group 1 vs group 3 (P = 0.341), group 1 vs group 2 (P = 0.862), group 2 vs group 4 (P = 0.061), group 2 vs group 3 (P = 0.744), and group 3 vs group 4 (P = 0.249). We concluded that the association of low-power laser with a bone-inducing substance produced better results than when low-power laser or BMPs were used alone.     

辐照对rhBMP-2骨诱导活性的影响

目的探讨不同剂量γ射线辐照对基因重组人骨形态发生蛋白2(rhBM P-2)骨诱导活性的影响及胶原载体的作用。方法以胶原海绵为载体制成rhBM P-2胶原复合物,分为4类:(Ⅰ)对照组:复合物未辐照;(Ⅱ)复合物辐照组:剂量为15、20、25和50kGy;(Ⅲ)载体辐照组:胶原载体接受50kGy辐照,然后复合未辐照的rhBM P-2;(Ⅳ)BM P辐照组:将rhBM P-2辐照25和50kGy,然后复合于胶原载体。各类BM P胶原复合物植入大鼠股骨肌袋内,术后2周和4周取出包块,进行大体、X线和组织学观察,通过组织形态分析对成骨进行定量分析。结果术后2周,复合物辐照15、20和25kGy组与对照组包块的大体、X线和组织学所见相似,包块内见大量未成熟的编织骨,成骨定量分析见20kGy组与对照组差异无统计学意义;复合物辐照25kGy组新生骨量明显低于复合物辐照15kGy组(P<0.05);复合物辐照50kGy和BM P辐照组无新骨形成。术后4周,各组均出现含有骨髓的成熟骨组织,其中复合物辐照25kGy组与对照组相似;复合物辐照50kGy组有成熟骨但数量较少;载体辐照和BM P辐照组新生骨量明显少于对照组。结论20和25kGy剂量辐照对rhBM P-2骨诱导活性没有影响或仅有一过性抑制;在辐照过程中,胶原载体对BM P具有保护作用。     

同种异体微小颗粒骨复合BMP胶原修复骨缺损的实验研究

目的探讨以同种异体微小颗粒骨复合BMP胶原修复节段性骨缺损的效果.方法取34只新西兰白兔,于两侧桡骨干制成长1.5 cm骨膜骨缺损,左侧(A组)植入由200mg同种异体微小颗粒骨、10 mg BMP与0.2 ml胶原的复合物;右侧(B组)植入复合0.2 ml胶原的200mg同种异体微小颗粒骨.分别于术后2、4、8、12周进行影像学,组织学检查;术后8、12周行骨密度检测;术后12周行生物力学检查.另取8只仅加入0.2 ml胶原作为空白对照组(即C组).结果放射学以及组织学检查结果显示A,B两组骨缺损区均得到了比较彻底的修复,但A组在成骨速度、骨再生量、再生髓腔结构等方面均优于B组.8、12周骨密度测试结果显示A组的骨密度值高于B组.12周的生物力学实验结果显示三点弯曲实验结果表明A组极限强度值明显高于B组,而且A组的弯曲刚度也好于B组;轴向压缩实验结果说明A组的抗压刚度优于B组;扭转实验结果指出A组的抗扭转刚度和最大扭矩分别高于B组,差异均有显著性意义(P<0.05).骨密度测定和生物力学实验结果证实了两组成骨情况的差异,这与新生骨的成熟程度不同有关.复合了BMP的A组因髓腔再通时间早,骨改建塑形更完善,新生板层骨的成熟度亦较高,耐受应力情况也更好.C组则不能产生骨性愈合.结论同种异体微小颗粒骨修复节段性骨缺损的成骨作用令人满意,复合BMP后效果更佳.     

Gene-mediated osteogenic differentiation of stem cells by bone morphogenetic proteins-2 or -6.

Bone marrow-derived mesenchymal stem cells (BMDMSC) hold promise for targeted osteogenic differentiation and can be augmented by delivery of genes encoding bone morphogenetic proteins (BMP). The feasibility of promoting osteogenic differentiation of BMDMSC was investigated using two BMP genes in monolayer and three-dimensional alginate culture systems. Cultured BMDMSC were transduced with E1-deleted adenoviral vectors containing either human BMP2 or BMP6 coding sequence under cytomegalovirus (CMV) promoter control [17:1 multiplicities of infection (moi)] and either sustained in monolayer or suspended in 1 mL 1.2% alginate beads for 22 days. Adenovirus (Ad)-BMP-2 and Ad-BMP-6 transduction resulted in abundant BMP-2 and BMP-6 mRNA and protein expression in monolayer culture and BMP-2 protein expression in alginate cultures. Ad-BMP-2 and Ad-BMP-6 transduced BMDMSC in monolayer had earlier and robust alkaline phosphatase-positive staining and mineralization and were sustained for a longer duration with better morphology scores than untransduced or Ad-beta-galactosidase-transduced cells. Ad-BMP-2- and, to a lesser degree, Ad-BMP-6-transduced BMDMSC suspended in alginate demonstrated greater mineralization than untransduced cells. Gene expression studies at day 2 confirmed an inflammatory response to the gene delivery process with upregulation of interleukin 8 and CXCL2. Upregulation of genes consistent with response to BMP exposure and osteogenic differentiation, specifically endochondral ossification and extracellular matrix proteins, occurred in BMP-transduced cells. These data support that transduction of BMDMSC with Ad-BMP-2 or Ad-BMP-6 can accelerate osteogenic differentiation and mineralization of stem cells in culture, including in three-dimensional culture. BMP-2-transduced stem cells suspended in alginate culture may be a practical carrier system to support bone formation in vivo. BMP-6 induced a less robust cellular response than BMP-2, particularly in alginate culture.     

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BMP-4对兔骨髓基质干细胞生物学行为的影响

目的应用原核细胞基因工程方法生产出有活性的骨形态发生蛋白-4(bonemorphoge-neticprotein-4,BMP-4),观察其对骨髓基质干细胞生物学行为的影响。方法利用RT-PCR技术,从成熟的人胎盘组织中扩增出长0.34kb编码人BMP-4成熟肽的基因序列,装入表达载体pET-22b( ),转化大肠杆菌BL-21菌株,并诱导目的蛋白表达,SDS-PAGE检测表达蛋白。获得的蛋白制品经小鼠异位成骨测活证实后,诱导培养的兔骨髓基质干细胞,观测细胞形态变化、碱性磷酸酶和骨钙素的含量。结果大肠杆菌中目的蛋白表达量达菌体蛋白的15%,该蛋白可诱导骨髓基质干细胞向成骨细胞分化,形成钙结节,碱性磷酸酶和骨钙素的含量也明显增加。结论大肠杆菌可表达出有活性的BMP-4,该蛋白可诱导骨髓基质干细胞向成骨细胞分化。     

Ectopic bone formation of human bone morphogenetic protein-2 gene transfected goat bone marrow-derived mesenchymal stem cells in nude mice

onemorphogenetic proteins (BMPs)haveapowerfulcapacitytoelicitnewboneformation .ThereareseveraldeliverymethodsofBMPsintreatingbonedefects ,oneofwhichisgenetherapy .Retrovirus,adenovirusandadeno associatedvirushavebeenutilizedtodeliverBMPgene .1,2 Sincethedirectuseofthesevectorshasseveraldisadvantages ,wehavedevelopedexvivo genetherapytechniquewhichinvolvestheisolationandcultivationofautologousbonemarrow derivedmesenchymalstemcells (MSCs) ,transfectionofthecellsinvitroandimplantationofthesec…     

基因治疗对兔下颌骨牵引过程中骨形成蛋白-2表达的影响

目的 探索电穿孔介导的基因治疗对兔下颌骨牵引成骨过程中骨形成蛋白-2(bone morphogenetic protein-2,BMP2)表达的影响.方法 新西兰大白兔双侧下颌骨截骨,术后3d开始牵引,连续牵引7d后,将实验动物分为A、B、C、D、E5组,A、B、C组分别在牵引区注射2 μg(0.1 μg/μl)重组质粒pIRES-hVEGF165-hBMP2、pIRES-hBMP2、pIRES-hVEGF165.D组与E组分别注射相同剂量的空质粒(pIRES)和生理盐水(NS).各组分别于固定期第7、14、28天处死动物,取材行免疫组织化学检测BMP2的表达情况,并利用病理图像分析系统进行分析.结果 BMP2主要在肉芽组织中的炎细胞及新生幼稚骨小梁表面的细胞组织中表达.固定7d时表达最强烈,各时点基因治疗组明显强于对照组.结论 电脉冲介导的基因治疗能使BMP2在牵引区的表达增强和表达时限延长并促进细胞的分裂增殖与分化,促进牵引区细胞基质的形成和新骨生成.