层黏连蛋白体外对人虹膜色素上皮细胞的影响

Effects of laminin on human iris pigment epithelial cells

目的:研究不同浓度的层黏连蛋白对体外培养的人虹膜色素上皮细胞(IPE)的作用。方法:采用酶辅助的显微分离法对成人眼进行IPE的原代和传代培养,观察其形态;以不同浓度的层黏连蛋白(0,1,5,10mg/L)处理第3代IPE细胞,MTT法作出生长曲线,免疫荧光法检测细胞角蛋白1(CK1)表达的变化。结果:原代培养的IPE细胞内色素丰富,而传代培养后色素显著减少;低浓度层黏连蛋白(5mg/L)明显促进IPE细胞增殖(0.187vs0.151,0.236vs0.176,0.245vs0.215,0.261vs0.238,0.279vs0.254;P<0.05);高浓度(10mg/L)明显抑制细胞增殖(0.121vs0.151,0.081vs0.176,0.094vs0.215,0.065vs0.238,0.078vs0.254;P<0.05);CK1染色均为阳性,高浓度层黏连蛋白(10mg/L)使IPE细胞聚集,CK1表达明显增强。结论:层黏连蛋白对IPE细胞具有双重作用,低浓度的层黏连蛋白适于IPE细胞的体外培养。

AIM： To observe the effects of laminin on human iris pigment epithelial （IPE） cells in vitro. HETHODS： Human I PE cells were isolated by enzyme-assisted microdissection method and cultured in plates. The cultivation of IPE cells was observed and the morphological features were recorded with digital camera. The third passage subcultured IPE cells were treated with Laminin of varies concentration （0, 1, 5 and 10mg/L）. The cells proliferation was detected with HTT assay and the growth curve were made to show the different trend. Positive expression of cytokeratin-1 （CK1） was recorded with immunofluorescent microscopy. RESULTS： There were a great number of pigmented granules within primary IPE cells and decreased gradually with the subculture passages. The IPE cells proliferations were significantly stimulated by Laminin of 5mg/L （0.187 vs0.151, 0.236 vs 0.176, 0.245 vs 0.215, 0.261 vs 0.238, 0.279 vs 0.254; P〈0.05）, and inhibited at the concentration of 10mg/L （0.121 vs 0.151, 0.081 vs 0.176, 0.094 vs 0.215, 0.065 vs 0.238, 0.078 vs 0.254; P 〈0.05）. All the cells expressed positive CK1, while the cells were accumulated to increase CK1 expression when treated with 10mg/L Laminin. CONCLUSION： Laminin show double effects to human IPE cells, the concentration relatively low may be much suitable for human IPE in vitro culture.