激活态雪旺细胞源性神经营养因子对脐血间充质干细胞分化的影响  

Impact of activated schwann cells-derived neuronotrophins on differentiation of human umbilical cord blood mesenchymal stem cells into neurogenic cellsin vitro

作　　者：周先虎[1] 郝岩 冯世庆[1] 孟恒星 陈有[1] 宁广智[1] 

机构地区：[1]天津医科大学总医院骨科,300052 [2]天津市协和干细胞研究中心

出　　处：《中华实验外科杂志》2012年第4期719-722,共4页Chinese Journal of Experimental Surgery

基　　金：国家自然科学基金资助项目（81171714、81070982）

摘　　要：目的观察激活态雪旺细胞（ASCs）分泌神经营养因子对诱导人脐血间充质干细胞（HUCBMSCs）神经方向分化的作用。方法采用双酶消化组织块法结合机械分离法分离培养ASCs；Ficoll法分离得到人脐血单个核细胞（HCMNCs）并经贴壁培养、分离后获得HUCBMSCs。采用全反式微甲酸＋碱性成纤维细胞生长因子（bFGF）＋表皮生长因子（EGF）（对照组）、ASCs培养基半量换液法（实验组）。应用免疫组织化学、蛋白印迹法（Westernblot）半定量、荧光实时聚合酶链反应（Real—timePCR）[神经纤维200（NF200）、胶质纤维酸性蛋白（GFAP）、髓磷脂碱性蛋白（MBP）]定量分析等方法在诱导培养后1、2、3、4周进行综合评价HUCBMSCs诱导分化，对实验结果进行统计学分析。结果免疫组织化学染色结果证实HUCBMSCs在ASCs和全反式微甲酸及bFGF、EGF的作用下向神经源性细胞分化。不同诱导时间细胞阳性率、Westernblot半定量、Real—timePCR定量分析结果显示差异有统计学意义（P〈0．05），而同一时间内两种诱导方法比较差异无统计学意义（P〉0．05）。结论ASCs可以分泌多种神经营养因子并促进HUCBMSCs向神经源性细胞分化，与传统全反式微甲酸诱导方法比较更简单。Objective To observe the capability of human umbilical cord blood mesenehymal stem cells （HUCBMSCs） to differentiate into neurogenie cells in vitro under the impact of activated Sehwann cells （ASCs）-derived neurenotrephins. Methods ASCs were cultured by using the dual-enzyme digestion method combined with mechanical separation. HUCBMSCs were cultured through isolating HCMNCs by Ficoil＇ s way and then plastic-adherent culture. Two methods were used for HUCBMSCs culture ： all-trans retinoie acid ＋ bFGF ＋ EGF （ control group） , 1/2 DMEM ＋ ASCs culture （ medium after ASCs cultured for 2 days, experimental group ）. Immunohistochemistry, Western blotting, and semi-quantitative and real- time polymerase chain reaction （PCR, NF200, GFAP, MBP） quantitative analysis were used to evaluate neurogenie differentiation of HUCBMSCs. Results The results of immunohistoehemistry staining, such as NF200, GFAP and MBP, confirmed that HUCBMSCs could differentiate into neurogenie ceils under the effect of ASCs and RA ＋ bFGF ＋ EGF. The statistical analysis of Western blotting and RT-PCR （ NF200, GFAP, MBP） indicated that there existed significant difference between two groups. Conclusion HUCBMSCs can proliferate and differentiate into neuregenie cells under the effect of numerous neurotrephie factors from ASCs in vitro.

关 键 词：脐血间充质干细胞 雪旺细胞 神经修复 分化 

分 类 号：R329[医药卫生—人体解剖和组织胚胎学]