槲皮素对高糖诱导的脉络膜和视网膜细胞血管生成的影响及其与细胞自噬的关系

Effect of quercetin on high-glucose-induced choroidal and retinal angiogenesis and its relationship with autophagy

目的观察槲皮素对高糖诱导的脉络膜和视网膜细胞血管生成的影响及其与细胞自噬的关系。方法将恒河猴脉络膜-视网膜内皮细胞(RF/6A)随机分为三组:对照组(DMEM培养基中培养)、高糖组(DMEM培养基中加入25 mmol/L D-葡萄糖)、高糖+槲皮素组(在DMEM培养基中加入25 mmol/L D-葡萄糖及100μmol/L槲皮素),培养24 h时,Western blot法检测细胞中自噬标志蛋白微管相关蛋白1轻链3(LC3)及Beclin-1的表达,荧光显微镜观察GFP-LC3融合蛋白的表达,划痕实验检测细胞迁移,Matrigel实验检测细胞管腔形成。结果与对照组相比,高糖组RF/6A细胞中LC3及Beclin-1蛋白表达增强(P<0.05),GFP-LC3阳性斑点细胞比例增多(P<0.05);与高糖组相比,高糖+槲皮素组LC3及Beclin-1蛋白表达减少(P<0.05),GFP-LC3阳性斑点细胞比例降低(P<0.05)。与对照组相比,高糖组RF/6A细胞迁移距离明显增大(P<0.05)、管腔形成数明显增多(P<0.05);高糖+槲皮素组细胞迁移和管腔形成明显减弱(P<0.05)。结论槲皮素可抑制高糖诱导的脉络膜和视网膜血管生成,该抑制作用可能与其抑制细胞自噬有关。

Objective To observe the effect of quercetin on high-glucose-induced choroidal and retinal cellular angiogenesis and its relationship with autophagy.Methods Rhesus monkey choroid-retinal endothelial cells(RF/6A)were randomly divided into three groups:control group(DMEM medium),high-glucose group(25 mmol/L D-glucose in DMEM medium),high-glucose+quercetin group(25 mmol/L D-glucose and 100μmol/L quercetin in DMEM medium).After 24 h culture,Western blot was used to detect the expression of cellular autophagy marker proteins microtubule associated protein 1 light chain 3(LC3)and Beclin-1,and fluorescence microscopy was used to observe the expression of GFP-LC3 fusion protein.Cell migration and tube formation were detected by scratch assay and matrigel assay,respectively.Results Compared with control group,the protein expression of LC3 and Beclin-1 in RF/6A cells significantly increased in high-glucose group(P<0.05),and the proportion of GFP-LC3 positive speckled cells also increased(P<0.05).Compared with high-glucose group,the expression of LC3and Beclin-1 decreased in high-glucose+quercetin group(P<0.05),and the proportion of GFP-LC3 positive speckled cells also decreased(P<0.05).Compared with control group,the RF/6A cell migration distance and tube formation number were significantly increased in high-glucose group(P<0.05).Cell migration and tube formation were significantly reduced in high-glucose+quercetin group(P<0.05).Conclusion Quercetin can inhibit choroidal and retinal angiogenesis induced by high glucose,which may be related to its inhibitory role in autophagy.