嘌呤受体P2X7在人眼小梁网细胞上的表达及分布

目的　探讨嘌呤受体Ｐ２Ｘ７在体外培养正常人眼小梁网细胞上的表达及分布。方法　体外培养、鉴定正常人眼小梁网细胞，利用ＲＴ－ＰＣＲ、Ｗｅｓｔｅｒｎ－ｂｌｏｔ和免疫荧光法分别检测正常人眼小梁网细胞上Ｐ２Ｘ７受体ｍＲＮＡ和蛋白的表达情况及受体蛋白的主要分布部位。结果　光镜下观察原代培养的细胞具有人眼小梁网细胞的典型形态；免疫组织化学方法检测结果表明，培养的细胞纤维连接蛋白、层粘连蛋白、神经元特异性烯醇化酶及波形蛋白染色阳性，第Ⅷ因子染色阴性，鉴定为人眼小梁网细胞；ＲＴ－ＰＣＲ扩增到了Ｐ２Ｘ７ｍＲＮＡ１５２ｂｐ大小的基因片段，Ｗｅｓｔｅｒｎ－ｂｌｏｔ得到了相对分子质量约为７４０００的Ｐ２Ｘ７的蛋白条带，免疫荧光法证实Ｐ２Ｘ７受体在小梁网细胞膜、细胞质及细胞核内均有广泛分布。结论　人眼小梁网细胞上有Ｐ２Ｘ７受体表达，其在细胞内分布广泛。

Expression and distribution of purinergic receptor P2X7 in human trabecular meshwork cells

Objective To discuss the expression and distribution of the purinergic receptor P2X7 in human trabecular meshwork cells ( HTMC ) cultured in vitro. Methods HTMC ( obtained from donated bodies ’ eyeball) were cultured in vitro and identified. The expression and distribution of purinergic receptor P2X7 were detected by RT-PCR.Western-blot analysis and immunofluorescence. Results The typical morphological of HTMC were observed by optical microscope. Immunohistochenucaol assays confirmed that fibronectin , lanunin . neuron-specific enolase . vimentin were positive.and factor vm were negative, which indicated that the primary cultures in vitro were indeed HTMC. RT-PCR amplified a 152 bp gene segment of P2X7 mRNA. and Western-blot analysis showed a protein band of P2X7 with relative molecular mass of 74 000. Immunofluorescence confirmed a wide range of distribution of P2X7 receptor in the cell membrane, cytoplasm and nucleus of HTMC. Conclusion The purinergic receptor P2X7 is expressed in HTMC . and it is widely distributed in HTMC.