人眼虹膜色素上皮细胞的培养与鉴定

Cultivation of human iris pigment epithelia and identification of its characteristics

目的 :观察体外培养的人眼虹膜色素上皮细胞(irispigmentepithelium ,IPE)的生长状况 ,用透射电镜、细胞免疫化学实验鉴定其性质 ,为进一步研究IPE细胞生理、病理作用提供模型。方法 :对来源于不同个体的健康人眼采用酶消化加显微分离法 ,分离IPE细胞 ,对其进行培养、传代、冻存和复苏。用透射电镜、细胞免疫化学法鉴定细胞性质。建立IPE细胞的有限细胞系。结果 :原代培养的IPE呈棕黑色多角形或不规则形 ,2 4h内 73%细胞贴壁。贴壁细胞体积变大呈多边形或方形 ,胞核轮廓变清 ,胞浆内充满黑色素颗粒。分裂增殖生长的细胞色素颗粒逐渐减少 ,4～ 6代后细胞内色素随着不断分裂逐渐减少至消失 ,部分形态变为成纤维细胞状。电镜下可见细胞表面有丰富的微绒毛 ,胞浆内含少量色素颗粒。细胞免疫化学抗角蛋白与抗S 10 0染色显示IPE细胞胞质呈鲜红色着色。结论 :采用酶消化加显微分离法可以建立人IPE的有限细胞系。

Objective:To establish a method to culture a human iris pigment epithelium(IPE) cell line.Methods:Enzyme assisted microdissection was used to isolate IPE cells derived from healthy donor eyes. Separated IPE was then cultured,purified,passaged,freeze preserved and resuscitated. Transmission electron microscope immunochemistry was used to identify and study the characteristics of human IPE.Results:Separated IPE showed as granular or irregular with brown color. Seventy three percent of IPE anchored in 24 h. Anchored IPE spread in multiangular or dental form. After 4～6 circles,the pigment of IPE decreased and even disappeared. Some cells became fibroblast like. Transmission electron microscopy displayed rich microvillus on the surface of IPE with little pigmented grain. Cultured IPE showed as cytokine and S 100 protein positive under immunochemistry study.Conclusion:Enzyme assisted microdissection is a reliable and quick method to establish an IPE cell line.