慢病毒介导骨形态发生蛋白2和血管内皮生长因子165双基因转染骨髓间充质干细胞复合脱钙松质骨治疗兔股骨头坏死  

Bone marrow mesenchymal stem cells co-transfected by lentivirus-mediated bone morphogenetic protein 2 and vascular endothelial growth factor 165 in combination with demineralized bone matrix for treating rabbit femoral head necrosis

作　　者：马跃刚 李强[1] 陶旋[1] 周桢杰 朱伦井 段江涛 Ma Yuegang;Li Qiang;Tao Xuan;Zhou Zhenjie;Zhu Lunjing;Duan Jiangtao(Affiliated Hospital of Guilin Medical University,Guilin 541000,Guangxi Zhuang Autonomous Region,China)

机构地区：[1]桂林医学院附属医院

出　　处：《中国组织工程研究》2019年第33期5275-5280,共6页Chinese Journal of Tissue Engineering Research

基　　金：国家自然科学基金项目(31160199),项目负责人:李强;广西壮族自治区自然科学基金项目(2014GXNSFAA118263),项目负责人:李强;广西省教育厅科研项目(LX2014254),项目负责人:李强~~

摘　　要：背景:骨形态发生蛋白2和血管内皮生长因子165均在骨修复过程中扮演着重要角色。利用慢病毒介导这2种基因转染骨髓间充质干细胞复合脱钙松质骨治疗骨损伤具有重要意义。目的:观察慢病毒介导骨形态发生蛋白2和血管内皮生长因子165基因共转染骨髓间充质干细胞复合脱钙松质骨治疗兔股骨头坏死的效果,以此来探讨双基因在未来骨损伤修复过程中的可行性。方法:采用液氮冰冻联合髓芯减压方法制备股骨头坏死模型,将造模成功的实验兔分为3组(n=12):(1)未转染组:植入骨髓间充质干细胞复合脱钙松质骨;(2)单基因转染组:植入骨形态发生蛋白2转染的骨髓间充质干细胞复合脱钙松质骨;(3)双基因转染组:植入骨形态发生蛋白2和血管内皮生长因子165共转染的骨髓间充质干细胞复合脱钙松质骨。术后3,6,9周各组分别处死4只兔子,Westernblot检测股骨头内骨形态发生蛋白2蛋白的表达量;苏木精-伊红染色观察股骨头坏死修复情况。实验方案经桂林医学院动物实验伦理委员会批准。结果与结论:(1)术后3,6,9周各组均有骨形态发生蛋白2阳性表达,单基因转染组、双基因转染组表达量均高于未转染组(P<0.05),且呈递增趋势;术后3周,单基因转染组、双基因转染组骨形态发生蛋白2表达差异无显著性意义(P>0.05);术后第6,9周,双基因转染组骨形态发生蛋白2表达量明显高于单基因转染组(P<0.05);(2)术后9周,双基因转染组新生骨痂可填满股骨头缺损处,未转染组、单基因转染组虽有骨痂形成,但均未填满缺损处;苏木精-伊红染色示未转染组有初级骨小梁形成,但骨细胞坏死较多,骨小梁较少;单基因转染组有大量骨小梁形成,形态较好,仅有部分中断,仍不及正常骨组织;双基因转染组形成大量成熟致密骨小梁,形态与正常骨组织无明显差异;(3)结果表明,慢病毒介导骨形态发生蛋白2和血管内�BACKGROUND: Bone morphogenetic protein 2 (BMP-2) and vascular endothelial growth factor 165 (VEGF-165) play important roles in bonerepair. It is of great significance to treat bone injury by lentivirus-transfected bone marrow mesenchymal stem cells combined withdemineralized bone matrix.OBJECTIVE: To observe the therapeutic effects of bone marrow mesenchymal stem cells with lentivirus-mediated BMP-2 and VEGF-165co-transfection and demineralized bone matrix in a rabbit model of osteonecrosis of the femoral head, so as to explore the feasibility of dualgenes in the bone repair.METHODS: Femoral head necrosis model was prepared by liquid nitrogen freezing combined with core decompression. Rabbit modelssuccessfully prepared were divided into three groups (n=12/group): non-transfection group, treated with bone marrow mesenchymal stemcells combined with demineralized bone matrix;single gene transfection group, treated with BMP-2-transfected bone marrow mesenchymalstem cells combined with demineralized bone matrix;dual-gene transfection group, treated with BMP-2 and VEGF-165 co-transfected bonemarrow mesenchymal stem cells combined with demineralized bone matrix. Four rabbits from each group were sacrificed at 3, 6, and 9 weekspostoperatively. Western blot was used to detect the expression of BMP-2 protein in the femoral head of each group. Hematoxylin-eosinstaining was used to observe the repair of femoral head necrosis in each group. The study protocol was approved by the Animal EthicsCommittee of Guilin Medical University.RESULTS AND CONCLUSION: BMP-2 positively expressed in all groups at 3, 6, and 9 weeks postoperatively, and the positive expression oftarget protein was significantly higher in the two transfection groups than the non-transfection group (P < 0.05). The positive expression ofBMP-2 in the two transfection groups showed an increasing trend at three time periods. At 3 weeks postoperatively, there was no difference inthe BMP-2 expression between the two transfection groups, while the BMP-2 expression was s

关 键 词：慢病毒 骨髓间充质干细胞 骨形态发生蛋白2 血管内皮生长因子165 脱钙松质骨 双基因转染 股骨头坏死 国家自然科学基金 

分 类 号：R459.9[医药卫生—治疗学] R394.2[医药卫生—临床医学]