二甲双胍通过抑制内质网应激诱导的细胞凋亡改善结肠炎黏膜上皮屏障损伤

Metformin alleviates intestinal epithelial barrier damage by inhibiting endoplasmic reticulum stress-induced cell apoptosis in colitis cell model

目的:探讨二甲双胍对溃疡性结肠炎黏膜上皮屏障损伤的作用及具体机制。方法:用人结肠癌细胞系Caco-2与人单核细胞系THP-1构建结肠炎体外细胞共培养模型,用1 mmol/L二甲双胍作用24 h后,运用流式细胞术检测肠上皮细胞凋亡情况,采用蛋白质印迹法检测紧密连接蛋白和内质网应激相关蛋白的表达水平。结果:与模型对照组比较,二甲双胍组细胞凋亡率从(14.22±2.34)%下降至(9.88±0.61)%(t=3.119,P<0.05),紧密连接蛋白1和密封蛋白1相对表达量增加(t=5.172和3.546,均P<0.05),内质网分子伴侣葡萄糖调节蛋白(GRP)78和内质网应激诱导的凋亡相关分子C/EBP同源蛋白(CHOP)、胱天蛋白酶(caspase)-12的蛋白表达水平下降(均P<0.05),蛋白激酶R样内质网激酶(PERK)和真核生物起始因子2α(eIF2α)的磷酸化水平下降(均P<0.05)。结论:二甲双胍可以通过减轻结肠炎肠上皮细胞的细胞凋亡和增加紧密连接蛋白的表达改善结肠炎肠黏膜上皮屏障损伤,其分子机制可能与抑制内质网应激诱导的细胞凋亡途径有关。

Objective:To investigate the effect and mechanism of metformin on intestinal epithelial barrier injury in ulcerative colitis.Methods:A cell model of colitis was established by co-culture of human colon cancer cell line Caco-2 and human monocyte cell line THP-1.The colitis model cells were treated with metformin at concentration of 1 mmol/L for 24 h.Flow cytometry was used to detect Caco-2 cell apoptosis,and Western blotting was used to detect the protein expression of tight junction proteins and endoplasmic reticulum stress-related proteins.Results:After metformin treatment,the apoptosis rate of Caco-2 cells was decreased from(14.22±2.34)%to(9.88±0.61)%(t=3.119,P<0.05),and the expression levels of tight junction protein-1 and claudin-1 increased(t=5.172 and 3.546,both P<0.05).In addition,the expression levels of endoplasmic reticulum-related proteins glucose regulated protein(GRP)78,C/EBP homologous protein(CHOP)and caspase-12,as well as the phosphorylation level of PRKR-like endoplasmic reticulum kinase(PERK)and eukaryotic translation initiation factor 2α(eIF2α)decreased(all P<0.05).Conclusion:Metformin may alleviate the intestinal epithelial barrier damage in colitis by reducing intestinal epithelial cell apoptosis and increasing the expression of tight junction proteins,which may be associated with the inhibition of endoplasmic reticulum stress-induced apoptotic pathway.