【摘要】  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（0.187vs 0.151，0.236vs 0.176，0.245vs 0.215，0.261 vs 0.238， 0.279vs 0.254；P <0.05）；高浓度（10mg/L）明显抑制细胞增殖（0.121vs 0.151，0.081vs 0.176，0.094vs 0.215，0.065vs 0.238，0.078vs 0.254；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。

【关键词】  层黏连蛋白；虹膜色素上皮细胞；培养，体外

     Effects of laminin on human iris pigment epithelial cells

    Xin-Ling Wang1, Tao Yu2, Guo-Gang Zhang3, Qi-Chang Yan1, Jin-Song Zhang1

    Foundation items: The Thirty-Three Science Fund Projection for Post-Doctor of the State Council (No.2003033282); Senior School Science Research Projection of Education Office of Liaoning Province of China (No.2004C041) and (No.05L576); Natural Science Foundation of Scientific Office of Liaoning Province of China (No.20062091)

    1Department of Ophthalmology, the Forth Affiliated Hospital, China Medical University, Shenyang 110005, Liaoning Province, China; 2Department of Radiology, Liaoning Cancer Hospital, Shenyang 110042, Liaoning Province, China; 3School of Traditional Chinese Medical, Shenyang Pharmaceutical University, Shenyang 110016,Liaoning Province, China

     Abstract AIM: To observe the effects of laminin on human iris pigment epithelial (IPE) cells in vitro . METHODS: Human IPE 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 MTT 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 vs 0.151, 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.

    · KEYWORDS: laminin; iris pigment epithelial cells; culture; in vitro

     0引言

    年龄相关性黄斑变性（age-related macular degeneration，ARMD）因存在视网膜色素上皮（retinal pigment epithelium，RPE）细胞功能异常而导致脉络膜新生血管形成，感光细胞变性，视力下降[1]。目前认为，RPE细胞移植是治疗ARMD的有效方法，但自体RPE细胞很难获取，而同种异体RPE移植，免疫排斥反应的发生往往难以避免。由于虹膜色素上皮(iridal pigment epithilium，IPE)细胞和RPE细胞胚胎发育的同源性及生物学功能的相似性[2]，使IPE替代RPE自体移植成为防治眼底病的新途径。

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