摘要
目的研究热休克蛋白(heat shock protein90,HSP90)抑制剂17-丙烯胺基-17-去甲氧基格尔德霉素(17-allylamino-17-demethoxygeldanamycin,17-AAG)对缺氧诱导的视网膜色素上皮(retinal pigment epithelial,RPE)细胞基质细胞衍生因子-1(stro-malcell-derivedfactor-1,SDF-1)表达的影响。方法将培养的第3代RPE细胞分为缺氧对照组、DMSO对照组和17-AAG预处理组。其中17-AAG预处理组根据浓度不同又分为6组:0.01μmol.L-1、0.10μmol.L-1、0.50μmol.L-1、1.0μmol.L-1、5.0μmol.L-1、10.0μmol.L-1。利用氯化钴建立体外RPE细胞的化学缺氧模型,不同浓度的HSP90抑制剂17-AAG预处理RPE细胞1h后给予12h缺氧处理,RT-PCR和Westernblotting方法检测SDF-1表达变化情况。结果缺氧对照组、DMSO对照组和不同浓度17-AAG预处理组SDF-1mRNA与内参β-actinmRNA光密度比值分别为0.89±0.04、0.93±0.07、0.62±0.06、0.52±0.06、0.43±0.06、0.28±0.04、0.21±0.04、0.17±0.03;SDF-1蛋白与内参β-actin蛋白光密度比值分别为0.76±0.04、0.80±0.06、0.65±0.04、0.34±0.03、0.20±0.02、0.16±0.02、0.07±0.02、0.05±0.01。与缺氧对照组比较,DMSO对照组SDF-1mRNA和蛋白的表达无明显变化,而不同浓度17-AAG预处理组SDF-1mRNA和蛋白的表达明显降低(P均<0.05),呈浓度依赖性。结论HSP90的特异抑制剂17-AAG能有效抑制缺氧条件下RPE细胞SDF-1表达。
Objective To investigate effects of heat shock protein 90（HSP90） inhibitor 17-allylamino-17-demethoxygeldanamycin（17-AAG） on expression of hypoxia induced stromal cell-derived factor-1（SDF-1） in retinal pigment epithelial（RPE） cells.Methods The cultured RPE cells at third passage were divided into hypoxic control group,DMSO control group and 17-AAG pretreatment groups,which were subdivided into six groups according to different concentrations（0.01 μmol·L-1,0.10 μmol·L-1,0.50 μmol·L-1,1.0 μmol·L-1,5.0 μmol·L-1,10.0 μmol·L-1）.Cobalt chloride was used to imitate chemical hypoxia to build RPE cell models in vitro.Before treated in hypoxia for 12 hours,RPE cells were pretreated with different concentrations of HSP90 inhibitor 17-AAG for 1 hour.RT-PCR and Western blotting analysis were used to examine the expression of SDF-1.Results The optical density ratios of SDF-1 mRNA and β-actin mRNA of hypoxic control group,DMSO control group and 17-AAG pretreatment groups were 0.89±0.04,0.93±0.07,0.62±0.06,0.52±0.06,0.43±0.06,0.28±0.04,0.21±0.04,0.17±0.03,respectively;and the optical density ratios of SDF-1 protein and β-actin protein were 0.76±0.04,0.80±0.06,0.65±0.04,0.34±0.03,0.20±0.02,0.16±0.02,0.07±0.02,0.05±0.01,respectively.Compared with hypoxic control group,the expression of SDF-1 mRNA and protein were without obvious changes in DMSO control group,but there were significant decrease for expression of SDF-1 mRNA and protein in 17-AAG pretreatment groups（both P〈0.05）,and the decrease was in concentration-dependent manner.Conclusion HSP90 specific inhibitor 17-AAG can effectively inhibit the expression of hypoxia induced SDF-1 in RPE cells.
出处
《眼科新进展》
CAS
北大核心
2009年第10期748-751,共4页
Recent Advances in Ophthalmology
