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目的观察炎症状态下高脂介导的内质网应激在人肾脏系膜细胞(HMCs)损伤中的作用,并探讨高脂在肾脏损伤中的作用机制。方法体外培养HMCs,分为对照组、高脂组、炎症组、高脂+炎症组、高脂+炎症+4-苯基丁酸(4-PBA)干预组(4-PBA干预组),分别培养24、48和72h。用油红O染色观察HMCs胞内脂质沉积水平;四甲基偶氮唑盐(MTT)法检测细胞增殖变化;实时荧光定量PCR检测GRP78、纤连蛋白(FN)mRNA水平;免疫细胞化学半定量分析糖调节蛋白78(GRP78)蛋白水平。结果 (1)与对照组比较,高脂组、炎症组、高脂+炎症组HMCs胞内脂质沉积增多(P均<0.01),细胞增殖增快,且呈时间依赖性(P均<0.01),同时FN mRNA表达升高(P均<0.01);与高脂+炎症组比较,4-PBA干预组胞内脂质沉积减少(P均<0.01),细胞增殖减慢(P均<0.01),且FN mRNA表达降低(P<0.05,P<0.01)。(2)与对照组比较,高脂组、炎症组、高脂+炎症组GRP78mRNA和蛋白表达升高(P<0.05,P<0.01);而4-PBA干预组GRP78mR-NA和蛋白表达较高脂+炎症组降低(P均<0.01)。(3)在24、48及72h,细胞脂质沉积水平与其增殖、GRP78蛋白及FN mR-NA水平均呈正相关(P均<0.01),GRP78蛋白表达水平与细胞增殖和FN mRNA水平呈正相关(P均<0.05)。结论炎症状态下,高脂可通过启动HMCs内质网应激致细胞损伤。
OBJECTIVE: To investigate the role of hypercholesterol-mediated endoplasmic reticulum stress in the injury of human renal mesangial cells (HMCs) under inflammatory conditions and to explore the mechanism of hyperlipidemia in renal injury. Methods HMCs were cultured in vitro and divided into 4 groups: control group, hyperlipidemia group, inflammation group, hyperlipidemia + inflammation group, hyperlipidemia + inflammation group and 4-PBA intervention group Cultures were 24,48 and 72 h. The intracellular lipid deposition of HMCs was observed by oil red O staining. The proliferation of HMCs was detected by MTT assay. The levels of GRP78 and fibronectin (FN) mRNA were detected by real-time fluorescence quantitative PCR. Quantitative analysis of glycoprotein 78 (GRP78) protein levels. Results (1) Compared with the control group, the intracellular lipid deposition in HMCs of hyperlipidemia group, inflammation group and hyperlipidemia + inflammation group increased (all P <0.01), and the proliferation of HMCs increased in a time-dependent manner (P <0.01). Compared with the hyperlipemia + inflammation group, the intracellular lipid deposition in the 4-PBA intervention group was decreased (P <0.01), and the proliferation of the cells slowed down (P <0.01) ), And the expression of FN mRNA decreased (P <0.05, P <0.01). (2) Compared with the control group, the expression of GRP78mRNA and protein in hyperlipidemia group, inflammation group and hyperlipidemia + inflammation group were significantly increased (P <0.05, P <0.01), while GRP78mRNA and protein were higher in 4-PBA intervention group Lipid + inflammation decreased (all P <0.01). (3) At 24, 48 and 72h, there was a positive correlation between cell lipid deposition and GRP78protein and FN mR-NA (all P <0.01), GRP78protein expression was positively correlated with cell proliferation and FN mRNA level P <0.05). Conclusion In the inflammatory state, hyperlipidemia can induce cell injury by activating the endoplasmic reticulum of HMCs.