体外诱导人骨髓间充质干细胞向色素上皮样细胞分化的研究

Induction of pigment epithelioid cells differentiated from mesenchymal stem cells in vitro

目的探讨人骨髓间充质干细胞(mesenchymal stem cells,MSC)向色素上皮样细胞诱导、分化的可行性,为视网膜移植提供种子细胞的来源。方法体外培养人视网膜色素上皮细胞(human retinal pigment epithelium,HRPE)和MSC,应用免疫细胞化学法检测CD34、CD44和角蛋白的表达情况,确定2种细胞的来源;通过Transwell6孔板非接触共培养的方式,建立HRPE和MSC共培养体系,显微镜下观察人MSC的形态学变化;对于诱导后的MSC进行免疫细胞化学鉴定,确定角蛋白的表达情况。结果MSC呈纤维样克隆样生长,人MSC的CD44(+),CD34(-);HRPE原代培养时细胞内布满色素颗粒,角蛋白表达(+)。HRPE和MSC共培养体系中,部分长梭形的MSC逐渐变成多边形,细胞内出现典型的色素颗粒;分化的色素上皮样细胞角蛋白表达阳性。结论MSC与HRPE在非接触共培养诱导方式下能分化成色素上皮样细胞;分化的细胞具有上皮细胞的特异性标志,诱导后的细胞是否具有HRPE的功能尚有待于进一步研究。

Objective To investigate the feasibility of induction and differentiation of mesenchymal stem cells （MSC） to pigment epithelioid cells, and to provide seed cells for retinal transplantation. Methods Human retinal pigment epithelium （HRPE） and MSC were cultured in vitro. Expressions of CD34, CD44 and keratin were detected with immunocytochemical method to determine the origins of these two different cells. Co-cultural system of HRPE and MSC was established by non-contact co-cultural way with Transwell 5 pore plate. Morphologic changes of human MSC were observed under microscopy. Induced MSC was identified with immunocytochemical method to determine the expression of keratin. Results MSC was with fiber-like and clone-like growth, and the expression of CD44 was positive, while CD34 was negative. Cells were filled with pigment granules at primary culture of HRPE, and the expression of keratin was positive. In the co-cultural system of HRPE and MSC, part of MSC of spindle- shaped appearance changed to polygon-shaped gradually ,and filled with typical pigment granules. The differentiated pigment epithelioid cells were positively stained for keratin. Conclusions MSC can be differentiated into pigment epithelioid cells with HRPE by non-contact co-cultural way. Differentiated cells show some specific characteristics of epithelial cells. The induced cells with characteristics of HRPE or not needs more researches.