目的:了解缺氧对人视网膜色素上皮(retinal pigmentepithelium,RPE)细胞Toll样受体4(Toll-like receptor4,TLR4)表达的影响。方法:采用200μmol/L CoCl2处理RPE细胞建立化学缺氧模型,以未处理细胞作对照,在缺氧处理后2,4,8,12和24h用免疫荧光法观察TLR4在细胞中的定位,用RT-PCR和Western blot检测细胞中TLR4表达水平。结果:共聚焦显微镜观察到正常对照组RPE细胞胞质内有微弱的TLR4荧光表达,缺氧后胞质内荧光表达明显增强,缺氧12h荧光强度达最强。RT-PCR和Western blot结果分别提示随缺氧时间延长,RPE细胞中TLR4mRNA和蛋白表达增强水平升高,缺氧至12h表达高峰最强(P<0.05),24h表达开始下降,各组差异有统计学意义。结论:缺氧可诱导RPE细胞中TLR4表达增强。

AIM:To observe the effects of hypoxia on the expression of Toll-like receptor 4 (TLR4) in cultured human retinal pigment epithelium (RPE) cells.METHODS:Chemical hypoxic model was established by treating the cultured human RPE cells with 200μmol/L CoCl2.The location of TLR4 in human RPE cells was observed by immunofluorescence assay,and RT-PCR together with Western blot were applied to detect the expression of TLR4 2,4,8,12,24 hours after the treatment.RESULTS:Immunofluorescence assay revealed that TLR4 was slightly expressed in human RPE cells under normoxic condition.After hypoxic treatment,there was a significant increase in green fluorescence within the cytoplasm.TLR4 expression tend to intensify over time and peaked at 12 hours,which was confirmed by RT-PCR and Western blot assay exhibiting TLR4 mRNA/protein levels (P<0.05).The expression levels began to decrease at 24 hours,and there was statistical significance among the groups.CONCLUSION:Hypoxia can strengthen TLR4 expression in cultured human RPE cells.

R774.1

中国国家自然科学基金资助项目(No.30371516,30672291)~~