成年大鼠眼睫状体缘色素上皮组织的神经前体细胞的培养和鉴定

Analysis of neural precursor cells of retinal pigmented epithelium in ciliary margin of the adult rat eye in vitro

目的 探讨成年大鼠眼睫状体缘色素上皮产生神经前体细胞的潜力及其生物学特征。方法 取成年SD大鼠睫状体缘处的色素上皮组织块 ,置于含bFGF和B2 7的DMEM/F12 无血清培养液中进行神经前体细胞培养 ,免疫组化反应染色鉴定细胞的表型。结果 培养 5～ 8d ,组织块长出由众多无色素和色素细胞构成的细胞集落 ,集落的大多数细胞处于增殖状态(BrdU反应阳性 ) ,并表达神经前体细胞的标志物 (nestin)。撤掉bFGF和B2 7,加入胎牛血清培养 3～ 5d ,集落的细胞发生分化 ,分别表达神经元和星形胶质细胞的标志物 (NSE、GFAP) ,但不表达少突胶质细胞的标志物 (O4 )。结论 由成年大鼠睫状体缘色素上皮长出的细胞集落不仅含有增殖能力和多分化潜能的神经前体细胞 ,而且尚含有色素细胞 。

Objective To investigate the potentials of retinal pigmented epithelium in ciliary margin (CM) of the adult rat eye to generate neural precursor cells (NPCs). Methods Explants of retinal pigmented epithelium from CM of adult SD rat eye were in vitro cultured in serum free medium containing bFGF and B 27 . The phenotypes of cultured cells were identified by immunohistochemical analyses. Results After 5～8 days neural spheres were generated from the explants. All neural spheres consisted of pigmented and non pigmented cells. The majority of cells in the spheres were BrdU immunoreactive and expressed the marker of NPCs (nestin). When the spheres were subcultured in medium containing fetal bovine serum for 3～5 days, cells in the spheres differentiated and expressed neuronal marker (NSE), astrocytic marker (GFAP), respectively; but did not expressed oligodendrocytic marker (O4). Conclusions Neural spheres generated from the explants of CM not only contain NPCs with the characteristics of mitotic ability and multipotential, but also contain pigmented cells. The retinal pigmented epithelium in CM may be the common germinal zone of both nervous specific cells and pigmented cells in adult rodent retina.