骨形态发生蛋白7腺病毒基因转染对骨髓基质干细胞生物学功能的影响  

Effect of bone morphogenetic protein 7 adenovirus transfection on biological function of bone marrow stromal stem cells

作　　者：蒲志超[1] 谢伟勇[1] 王延斌[1] 薛剑[1] 张兴世 

机构地区：[1]武警广东省总队医院番禺分院,广东省广州市511430

出　　处：《中国组织工程研究》2015年第6期837-842,共6页Chinese Journal of Tissue Engineering Research

基　　金：2012年广东省医学科研基金项目(A2012456)~~

摘　　要：背景:基因转染技术正积极地应用于组织再生治疗之中,研究表明骨形态发生蛋白7具有骨诱导特性,可以有效促进成骨细胞的发育以及新骨的形成。目的:探讨骨形态发生蛋白7腺病毒基因转染对骨髓基质干细胞生物学功能的影响。方法:分离、培养羊骨髓基质干细胞,利用重组骨形态发生蛋白7腺病毒(adenovirus bone morphogenetic protein 7,Adeno-BMP7)对第2代骨髓基质干细胞进行基因转染,透射电镜观察转染后细胞超微结构。流式细胞仪检测转染后的细胞周期,Western blot检测骨形态发生蛋白7的表达,Von Kossa染色观察钙结节形态情况。取转染后第3天及相应未转染的骨髓基质干细胞制备珊瑚-细胞复合物,于裸鼠脊柱两侧背部皮下注射4周和8周后进行大体观察和组织学检测。结果与结论:体外细胞超微结构观察可见Adeno-BMP7基因转染骨髓基质干细胞存在活跃的物质合成代谢;流式细胞仪检测发现转染不会对骨髓基质干细胞的细胞周期产生明显影响;Western blot检测转染后的骨髓基质干细胞存在骨形态发生蛋白7蛋白的表达;Von Kossa染色可见转染Adeno-BMP7后的骨髓基质干细胞形成较大的钙结节。经裸鼠皮下回植实验发现,Adeno-BMP7转染后骨髓基质干细胞的成骨能力和成骨质量明显提高。以上结果表明经Adeno-BMP7转染可以有效促进骨髓基质干细胞成骨分化。BACKGROUND: Gene transfer techniques have been actively used in tissue regeneration therapy. Bone morphogenetic protein 7 with osteoinductive properties can effectively promote osteoblast growth and new bone formation.OBJECTIVE: To explore the influence of bone morphogenetic protein 7 adenovirus gene transfection on the biological function of bone marrow stromal stem cells.METHODS: Goat bone marrow stromal stem cells were isolated, cultured and transfected by recombinant adenovirus containing bone morphogenetic protein 7(Adeno-BMP7). Cell ultrastructure was observed by transmission electronic microscope, cell cycle was detected by flow cytometry, expression of bone morphogenetic protein 7 was measure by western blot assay, and Von Kossa staining was used to observe the formation of calcium nodules. Bone marrow stromal stem cells transfected for 3 days and untransfected were used to prepare coral-cell complexes that were injected subcutaneously into the back of nude mice for 4 weeks and 8 weeks, followed by gross observation and histological examination.RESULTS AND CONCLUSION: Adeno-BMP7-transfected bone marrow stromal stem cells appeared to have active substance synthesis and metabolism. Adeno-BMP7 transfection played no effect on the cell cycle of bone marrow stromal stem cells. Bone morphogenetic protein 7 was expressed in the transfected bone marrow stromal stem cells.Larger calcium nodules were visible after Adeno-BMP7 transfection. Adeno-BMP7-transfected bone marrow stromal stem cells showed stronger osteogenic capability and higher bone quality. These results demonstrate that Adeno-BMP7 transfection can effectively promote the osteogenic differentiation of bone marrow stromal stem cells.

关 键 词：骨髓 间质干细胞 骨形态发生蛋白7 基因 干细胞 骨髓干细胞 骨形态发生蛋白7腺病毒基因 转染 骨髓基质干细胞 生物学功能 细胞周期 钙结节 成骨分化 超微结构 

分 类 号：R329.2[医药卫生—人体解剖和组织胚胎学]