人前脑啡肽基因修饰骨髓间充质干细胞系的生物学特性  

Human preproenkephalin gene-modified bone marrow mesenchymal stem cell line: its biological characteristics

作　　者：李新茂[1] 

机构地区：[1]南阳医学高等专科学校第一附属医院麻醉科,河南省南阳市473000

出　　处：《中国组织工程研究》2016年第19期2741-2747,共7页Chinese Journal of Tissue Engineering Research

摘　　要：背景:有研究发现修饰的细胞株移植能够有镇痛效果,但人前脑啡肽基因修饰骨髓间充质干细胞系的生物学特性的研究较少。目的:观察人前脑啡肽基因修饰骨髓间充质干细胞的生物学特性。方法:分离培养骨髓间充质干细胞,建立人前脑啡肽基因修饰的人骨髓间充质干细胞系。结果与结论:1锥虫蓝染色:冻存复苏后的P4代人骨髓间充质干细胞、人骨髓间充质干细胞-p BABE和人骨髓间充质干细胞-人前脑啡肽细胞活力差异无显著性意义(P>0.05);2流式细胞仪检测:P4代人骨髓间充质干细胞、人骨髓间充质干细胞-p BABE和人骨髓间充质干细胞-人前脑啡肽细胞的CD29和CD44的表达均呈阳性,CD34和CD45的表达均呈阴性;3成脂和成骨诱导:P4代人骨髓间充质干细胞、人骨髓间充质干细胞-p BABE和人骨髓间充质干细胞-人前脑啡肽细胞之间脂肪细胞占总细胞的比例比较差异无显著性意义(P>0.05),成骨诱导分化后,茜素红染色可见红色钙沉积;4PCR检测:人前脑啡肽基因在P4代骨髓间充质干细胞-p BABE细胞中高表达,在P4代人骨髓间充质干细胞中低表达;5免疫荧光染色:转染重组质粒p BABE-人前脑啡肽的人骨髓间充质干细胞中可见脑啡肽蛋白表达;6结果表明,人前脑啡肽基因修饰的人骨髓间充质干细胞系保持了骨髓间充质干细胞的多向分化潜能或分裂增殖能力,并可分泌脑啡肽蛋白。BACKGROUND： Previous studies have found that the transplantation of modified cell lines exhibit analgesic effect. But little is reported on the biological characteristics of human preproenkephalin gene-modified bone marrow mesenchymal stem cell lines. OBJECTIVE： To observe the biological characteristics of human preproenkephalin gene-modified bone marrow mesenchymal stem cells. METHODS： Bone marrow mesenchymal stem cells were isolated and cultured to establish human preproenkephalin gene-modified bone marrow mesenchymal stem cell lines. RESULTS AND CONCLUSION： After freezing-thawing, passage 4 human bone marrow mesenchymal stem cells, human bone marrow mesenchymal stem cells-pBABE and human bone marrow mesenchymal stem cells-human preproenkephalin exhibited no significant changes in the cell viability（P〉0.05）, as well as in the fat cell proportion after adipogenic induction（P〉0.05）. Moreover, red calcium deposition was presented in all these cells by alizarin red staining after osteogenic induction. Flow cytometry results showed that passage 4 human bone marrow mesenchymal stem cells, human bone marrow mesenchymal stem cells-pBABE and human bone marrow mesenchymal stem cells-human preproenkephalin could express CD29 and CD44, but not express CD34 and CD45. Human preproenkephalin genes were highly expressed in human bone marrow mesenchymal stem cells-pBABE, but lowly expressed in passage 4 human bone marrow mesenchymal stem cells. Additionally, recombinant plasmid pBABE-preproenkephalin-modified human bone marrow mesenchymal stem cells could express enkephalin protein. In conclusion, human preproenkephalin gene-modified human bone marrow mesenchymal stem cell lines can maintain the pluripotent differentiation or proliferation capacity of bone marrow mesenchymal stem cells, and secrete enkephalin protein.

关 键 词：干细胞 组织工程 脑啡肽类 骨髓干细胞 脑啡肽 疼痛 基因 镇痛 骨髓间充质干细胞 反转录 病毒 生物学特性 

分 类 号：R394.2[医药卫生—医学遗传学]