视网膜神经节细胞无血清上清培养液诱导大鼠视网膜干细胞的分化

Retinal ganglion cell-conditioned medium alters the differentiation of rat retinal progenitor cells

目的探讨视网膜神经节细胞无血清上清培养液对视网膜干细胞分化的影响。方法分离大鼠视网膜干细胞和视网膜神经节细胞;采用免疫荧光法鉴定体外培养的大鼠视网膜干细胞与视网膜神经节细胞,视网膜干细胞以Nestin抗体进行鉴定,视网膜神经节细胞以Thy-1抗体进行鉴定;以视网膜神经节细胞无血清上清培养液培养视网膜干细胞,以不加入条件培养液培养的视网膜干细胞为对照组,收集分化细胞,采用q PCR法检测Nestin、Pax6、Thy-1及Brn-3的基因表达。结果培养的视网膜干细胞Nestin抗体染色阳性,视网膜神经节细胞Thy-1抗体染色阳性。培养视网膜干细胞72 h后,与对照组相比,无血清上清培养液组细胞Nestin和PAX6基因相对表达量降低,差异均有统计学意义(均为P<0.000 1);Thy-1和Brn-3基因相对表达量升高,差异均有统计学意义(均为P<0.05)。结论视网膜神经节细胞无血清上清培养液能够诱导视网膜干细胞分化为视网膜神经节样细胞。

Objective T o investigate the effects of retinal ganglion cell-conditioned medium on the differentiation of retinal stem cells. Methods Retinal stem cells and retinal ganglion cells w ere isolated from rats,and immunofluorescence staining w as applied to identify rat retinal stem cells and retinal ganglion cells w ith Nestin and T hy-1 antibody,respectively. Retinal stem cells w ere cultured in presence or absence of retinal ganglion cell-conditioned medium for 72 h,follow ed by detection of Nestin,PAX6,T hy-1 and Brn-3 gene expression in retinal stem cells by qPC R. Results Isolated retinal stem cells w ere Nestin positive,and retinal ganglion cells w ere T hy-1 positive, indicating the success of isolation. C ompared to retinal stem cells cultured w ithout ganglion cell-conditioned medium,ones cultured w ith ganglion-conditioned medium had significantly dow nregulated expression of Nestin and PAX6( both P < 0. 000 1),and markedly upregulated expression of T hy-1 and Brn-3( both P < 0. 05). Conclusion Retinal ganglion cell-conditioned medium can induce the differentiation of retinal progenitor cells into retinal ganglion-like cells.