黄芪汤对高糖诱导足细胞损伤的保护作用及机制研究

目的观察黄芪汤对高糖诱导足细胞损伤的影响及作用机制。方法实验分为5组:对照组足细胞采用5.5 mmol/L葡萄糖培养液培养,模型组足细胞采用30 mmol/L葡萄糖培养液培养,黄芪汤30μg/mL组、黄芪汤100μg/mL组、黄芪汤300μg/mL组分别采用30 mmol/L葡萄糖培养液和相应浓度黄芪汤进行培养。细胞培养48 h后,采用CCK-8法检测各组细胞存活率,采用流式细胞术检测各组细胞凋亡情况,采用免疫荧光法检测各组细胞标记蛋白podocin表达情况,采用Western blot检测各组细胞内质网应激通路蛋白葡萄糖调节蛋白78(GRP78)、磷酸化蛋白激酶R样内质网激酶(p-PERK)、蛋白激酶R样内质网激酶(PERK)、磷酸化真核起始因子2α(p-eIF2α)、真核起始因子2α(eIF2α)]及其介导的凋亡相关蛋白增强子结合蛋白同源蛋白(CHOP)、B细胞淋巴瘤因子2(Bcl-2)]表达情况。结果模型组细胞存活率明显低于对照组(P<0.05),细胞凋亡率明显高于对照组(P均<0.05);黄芪汤各组细胞存活率均明显高于模型组(P均<0.05),细胞凋亡率均明显低于模型组(P均<0.05),且呈浓度依赖性。模型组podocin表达荧光强度明显弱于对照组,黄芪汤各组随着药物浓度升高podocin表达荧光强度明显增强。模型组GRP78、p-PERK、p-eIF2α、CHOP表达水平均明显高于对照组(P均<0.05);黄芪汤各组GRP78、p-PERK、p-eIF2α、CHOP表达水平均明显低于模型组(P均<0.05),且黄芪汤各组随药物浓度的升高GRP78、p-PERK、p-eIF2α表达水平均逐渐降低(P均<0.05)。模型组Bcl-2表达水平明显低于对照组(P<0.05);黄芪汤100μg/mL组、黄芪汤300μg/mL组Bcl-2表达水平均明显高于模型组(P均<0.05);黄芪汤100μg/mL组与黄芪汤300μg/mL组Bcl-2和CHOP表达水平比较差异均无统计学意义(P均>0.05)。结论黄芪汤可呈浓度依赖性的抑制高糖诱导的足细胞损伤,其作用可能与抑制PERK介导的内质网应激途径有关。

Protective effect of Huangqi decoction on high glucose-induced podocyte injury and its mechanism

Objective It is to observe the effect and mechanism of Huangqi decoction on high glucose-induced podocyte injury.Methods The experiment was divided into 5 groups:podocytes in the control group were cultured with 5.5 mmol/L glucose medium,podocytes in the model group were cultured with 30 mmol/L glucose medium,Huangqi decoction 30μg/mL group,Huangqi decoction 100μg/mL group and Huangqi decoction 300μg/mL group were cultured with 30 mmol/L glucose medium and the corresponding concentration of Huangqi decoction.After culturing for 48 h,the cell survival rate of each group was detected by CCK-8 method,the apoptosis of each group was detected by flow cytometry,the expression of the cell marker protein podocin was detected by immunofluorescence,and the Western blot was used to detect the expression of endoplasmic reticulum stress pathway proteinsglucose regulatory protein 78(GRP78),phosphorylated protein kinase R-like ER kinase(p-PERK),protein kinase R-like ER kinase(PERK),phosphorylated eukaryotic initiation factor 2α(p-eIF2α),eukaryotic initiation factor 2α(eIF2α)]and their apoptosis-related proteinsenhancer-binding protein homologous protein(CHOP),B-cell lymphoma factor 2(Bcl-2)].Results The cell survival rate of the model group was significantly lower and the apoptosis rate was significantly higher than that of the control group(P<0.05);the cell survival rate of each group of Huangqi decoction was significantly higher and the rate of apoptosis was significantly lower than that of the model group(P<0.05),the results were concentration-dependent.The fluorescence intensity of podocin expression in the model group was significantly weaker than that of the control group,and the fluorescence intensity of podocin expression in the Huangqi decoction groups increased with the increase of drug concentration.The expression levels of GRP78,p-PERK,p-eIF2α,and CHOP in the model group were significantly higher than those in the control group(P<0.05);the expression levels of GRP78,p-PERK,p-eIF2α,and CHOP in the Huangqi decoction groups were significantly lower than those in the model group(P<0.05),and the expression levels of GRP78,p-PERK,p-eIF2αgradually decreased with the increase of drug concentration in each group of Huangqi decoction(P<0.05).The expression level of Bcl-2 in the model group was significantly lower than that in the control group(P<0.05);the Bcl-2 expression levels in the Huangqi decoction 100μg/mL group and Huangqi decoction 300μg/mL group were significantly higher than the model group(P<0.05).There was no significant difference in the expression levels of Bcl-2 and CHOP between Huangqi decoction 100μg/mL group and Huangqi decoction 300μg/mL group(P>0.05).Conclusion Huangqi decoction can inhibit podocyte injury induced by high glucose in a concentration-dependent manner,and its effect may be related to the inhibition of the ERK-mediated endoplasmic reticulum stress pathway.