机构地区:[1]新乡市中心医院呼吸与危重症医学科,河南新乡453000
出 处:《临床肺科杂志》2018年第10期1824-1828,共5页Journal of Clinical Pulmonary Medicine
摘 要:目的探究免疫球蛋白重链结合蛋白(GRP78)的表达量降低对急性肺损伤(ALI)上皮细胞凋亡、增殖以及核因子-κB(NF-κB)信号通路的影响。方法采用脂质体Lipofectamine 2000将siRNA-GRP78转入肺上皮MLE-12细胞中,使用10μg/m L脂多糖(LPS)作用于生长状态良好的细胞24 h,建立急性肺损伤模型。实验分为4组:对照组为未经处理的细胞,模型组为LPS干预的细胞,空载体组为先把siRNA-NC转入细胞中然后经LPS干预,低表达组为先把siRNA-GRP78转入细胞中后经LPS干预。CCK-8检测细胞的增殖情况,流式细胞术检测细胞的凋亡率,蛋白质印迹法(Western blot)检测细胞中GRP78、P65、核因子κB抑制蛋白(IκB)、细胞周期蛋白D1(Cyclin D1)的表达量。结果与对照组相比,模型组、空载体组、低表达组细胞中GRP78蛋白的表达量显著升高(P<0.05);与模型组相比,低表达组显著降低(P<0.05)。与对照组相比,模型组、空载体组、低表达组细胞的存活率显著下降(P<0.05),凋亡率显著升高(P<0.05);与模型组相比,低表达组细胞的存活率显著升高(P<0.05),凋亡率显著降低(P<0.05)。与对照组相比,模型组和空载体组细胞中P65(P<0.05)、IκB(P<0.05)、Cyclin D1(P<0.05)蛋白的表达量显著降低;与模型组相比,低表达组细胞中P65、IκB、Cyclin D1蛋白的表达量显著升高(P<0.05)。结论下调GRP78表达抑制急性肺损伤上皮细胞凋亡,促进其增殖,其作用机制可能是通过抑制NF-κB信号通路介导的下游靶基因的表达来发挥作用。Objective To investigate the effect of reduced expression of immunoglobulin heavy chain binding protein( GRP78) on apoptosis,proliferation and NF-κB signaling pathway of epithelial cells in acute lung injury(ALI).Methods siRNA-GRP78 were transfected into lung epithelial MLE-12 cells by lipofectamine 2000.An acute lung injury model was established by using 10 μg/m L of lipopolysaccharide( LPS) acting on a well growing cell for 24 h.The experiment were divided into four groups: the control group of untreated cells,the model group treated with LPS,the empty vector group transfected into siRNA-NC and then treated by LPS and,the low expression group transfected into siRNA-NC and then treated by LPS.The proliferation of cells were detected by CCK-8 and apoptosis rate was detected by flow cytometry.Western blot was used to detect the expression of GRP78,P65,nuclear factor kappa B inhibitor protein( IκB) and cyclin D1.Results Compared with the control group,the expression of GRP78 protein in the model group,the empty vector group and the low expression group increased significantly( P〈0.05),and it was lower in the low expression group than in the model group( P〈0.05).Compared with the control group,the survival rate decreased significantly( P〈0.05) and the apoptosis rate increased significantly in the model group,the empty vector group and the low expression group( P〈0.05),and the survival rate of the cells was significantly higher and the apoptosis was lower in the low expression group than in the model group( P〈0.05).Compared with the control group,the expression of P65,IκB and Cyclin D1 proteins in the model group and the empty vector group were significantly reduced,but they were significantly higher in the low expression group than in the model group( P〈0.05).Conclusion The down-regulation of GRP78 expression can inhibit apoptosis and promote proliferation of epithelial cells in acute lung injury.Its mechanism may be by inhibiting NF-κB signaling pathway med
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