骨髓干细胞诱导形成的内皮细胞在体内外环境中的稳定性  

Phenotypical Stability of Endothelial Cells Differentiated from Bone Marrow Mesenchymal Stem Cell in Vitro and in Vivo

作　　者：董悦[1] 张雷[1] 邵素霞[1] 尹青[1] 陈炜[1] 赵春芳[1] 

机构地区：[1]河北医科大学组织胚胎教研室,石家庄050017

出　　处：《细胞生物学杂志》2009年第6期823-830,共8页Chinese Journal of Cell Biology

基　　金：河北省自然科学基金资助项目(No.C2008001030)~~

摘　　要：采用异硫氰酸荧光素(fluorescein isothiocyanate,FITC)标记的CD34(FITC-CD34)、免疫细胞化学及透射电镜鉴定血管内皮生长因子(vascular endothelial growth factor,VEGF)诱导Sprague-Dawley(SD)大鼠骨髓间充质干细胞(bone marrow mesenchymal stem cells,BM-MSCs)分化形成的内皮细胞(诱导组)。应用三维脱细胞真皮基质(acellular dermal matrix,ADM)支架组织,接种诱导形成的内皮细胞,种植于SD大鼠体内生长14天。同时采用不含VEGF的DMEM(DMEM组)和EGM-2培养基(EGM-2组),体外培养诱导形成的内皮细胞14天,酶联免疫吸附测定法(enzyme-linkedimmunosorbent assay,ELISA)检测各组细胞上清液中内皮素(endothelin-1,ET-1)和Ⅷ因子相关抗原(Ⅷfactor related antigen,F-ⅧAg)含量,进行统计学处理。实验结果表明,VEGF诱导BM-MSCs培养14天,细胞呈"铺路石"样排列,FITC-CD34呈现绿色荧光,免疫细胞化学抗CD3l和CD34表达阳性,细胞质中含Weibel-Palade小体,具有内皮细胞的特征。在SD大鼠体内生长7天和14天,内皮细胞保留其特征,对抗CD31和CD34表达阳性。内皮细胞在体外不含VEGF的DMEM培养基和EGM-2培养基中各培养14天后,细胞回复到诱导前状态,FITC-CD34反应阴性。ELISA检测DMEM组和EGM-2组中ET-1和F-ⅧAg含量明显低于诱导组(P<0.05)。研究结果显示,BM-MSCs诱导形成的内皮细胞,在体内环境中生存能保留其特征;但在缺乏VEGF诱导的体外培养基中生长,不能维持其特征。The endothelial cells which were differentiated from bone marrow mesenchymal stem cells （BM-MSCs） of Sprague-Dawley （SD） rats induced by vascular endothelial growth factor （VEGF） （induced group） were identified by fluorescein isothiocyanate （FITC） labelled CD34 （FITC-CD34）, and assayed by immunocytochemistry against CD31 and CD34. The ultrastructures of the endothelial cells were observed by transmission electron microscope （TEM）. After then the endothelial cells were seeded onto a three-dimensional acellular dermal matrix （ADM） scaffold, and transplanted into skeletal muscles of SD rats in vivo for 7 d and 14 d. DMEM without VEGF medium （DMEM group） and EGM-2 medium （EGM-2 group） was used respectively to culture the endothelial cells for 14 d in vitro, and the contents of both endothelin-1 （ET-1） and VIII factor related antigen （F-ⅧAg） in different groups were detected by the method of enzyme-linked immunosorbent assay （ELISA）. The results showed that after being induced by VEGF for 14 d, BM-MSCs were arranged like ＂paver＂, which showed green fluorescence of TITC-CD34 and were positive immunocytochemical reactions for CD31 and CD34. TEM observation found that there was typical Weibel-Palade body in the cytoplasm of these ceils showed the endothelial features. The endothelial cells survived and kept the endothelial features in vivo of SD rat for 7 d and 14 d after transplantation, these cells were positive reactions for CD31 and CD34. In contrast, after being cultured respectively in DMEM without VEGF medium and EGM-2 medium for 14 d, the endothelial cells did not show green fluorescence of TITC-CD34. ELISA results displayed the significantly lower contents of both ET-1 and F-ⅧAg of DMEM group and EGM-2 group than those of induced group （P〈0.05）. This study demonstrated that the endothelial cells which derived from BM-MSCs induced by VEGF could keep the phenotypical stability in vivo, but could not maintain the stability in the culture medium witho

关 键 词：骨髓干细胞 内皮细胞 诱导 血管内皮生长因子 

分 类 号：Q813.1[生物学—生物工程]