摘要
目的研究细胞因子信号抑制因子(suppressor of cytokine signaling,SOCS3)对JAK2/STAT3信号通路的调控作用及对急性肺损伤的影响。方法将A549细胞分为control组、model组、LPS+SOCS3组、LPS+SOCS3 NC组、LPS+AG490组、LPS+SOCS3+AG490组和LPS+SOCS3 NC+AG490组。control组A549细胞正常培养;model组细胞用LPS处理;LPS+SOCS3组和LPS+SOCS3 NC组细胞在用LPS处理后,分别转染SOCS3过表达质粒和NC质粒;LPS+SOCS3+AG490组和LPS+SOCS3 NC+AG490组细胞分别在model组和LPS+SOCS3 NC组基础上用JAK2特异性抑制剂处理。显微镜观察各组细胞形态学改变;CCK-8测定各组细胞活力;ELISA检测促炎因子IL-6和TNF-α含量;qPCR检测各组细胞SOCS3、JAK2、STAT3基因表达;Western blot法检测细胞中SOCS3、JAK2、STAT3、p-JAK2、p-STAT3蛋白表达变化。结果与control组相比,model组细胞皱缩,连接松散,细胞增殖能力下降(P<0.01),IL-6和TNF-α浓度均上升(P<0.01),SOCS3 mRNA表达下调(P<0.01),JAK2、STAT3 mRNA表达均上调(P<0.01),SOCS3蛋白表达下调(P<0.01),p-JAK2、p-STAT3蛋白表达上调(P<0.01)。与model组相比,LPS+SOCS3组和LPS+AG490组均细胞形态恢复,连接紧密,细胞增殖能力上升(P<0.01),IL-6和TNF-α浓度均下降(P<0.01),SOCS3 mRNA表达上调(P<0.01),JAK2、STAT3 mRNA表达下调(P<0.01),SOCS3蛋白表达上调(P<0.01),p-JAK2、p-STAT3蛋白表达均下调(P<0.01)。与LPS+AG490组及LPS+SOCS3 NC+AG490组相比,LPS+SOCS3+AG490组细胞形态进一步恢复,连接更加紧密,细胞增殖能力上升(P<0.01),IL-6和TNF-α浓度均下调(P<0.01),SOCS3 mRNA表达上调(P<0.01),JAK2、STAT3 mRNA表达均下调(P<0.01),SOCS3蛋白表达上调(P<0.01),p-JAK2、p-STAT3蛋白表达下调(P<0.01)。结论SOCS3过表达能够调控JAK2/STAT3信号通路,抑制脂多糖引起的急性肺损伤,与JAK2/STAT3通路抑制剂联用效果更佳。
Objective To explore the effect of suppressor of cytokine signaling(SOCS3)on JAK2/STAT3 signaling pathway and acute lung injury.Methods The cells were divided into control group,model group,LPS+SOCS3 group,LPS+SOCS3 NC group,LPS+AG490 group,LPS+SOCS3+AG490 group and LPS+SOCS3 NC+AG490 group.A549 cells in control group were cultured normally.A549 cells were treated with LPS to establish the acute lung injury model in model group.The cells in LPS+SOCS3 group and LPS+SOCS3 NC group were treated with LPS,and then transfected with SOCS3 overexpression plasmid and NC plasmid,respectively.A549 cells in LPS+SOCS3+AG490 group and LPS+SOCS3 NC+AG490 group were treated with JAK2 specific inhibitors on the basis of treatment in model group and LPS+SOCS3 NC group,respectively.The morphological changes of cells in each group were observed under a microscope.Cell viability was determined by CCK-8.The contents of pro-inflammatory cytokines IL-6 and TNF-αwere determined by ELISA.The expression levels of SOCS3,JAK2 and STAT3 genes were detected by qPCR.The protein expression levels of SOCS3,JAK2,STAT3,p-JAK2 and p-STAT3 were detected by Western blot.Results Compared with control group,the cell crinkling and loosening of junctions were found in model group,the cell proliferation ability was decreased(P<0.01),both IL-6 and TNF-αconcentrations were increased(P<0.01),SOCS3 mRNA expression was down-regulated(P<0.01),JAK2 and STAT3 mRNA levels were up-regulated(P<0.01),SOCS3 protein expression was down-regulated(P<0.01),and p-JAK2 and p-STAT3 protein expression levels were up-regulated(P<0.01).Compared with model group,the morphology of cells recovered and the connections became tighter in LPS+SOCS3 group and LPS+AG490 group,the cell proliferation capacity was increased(P<0.01),IL-6 and TNF-αcontents were decreased(P<0.01),SOCS3 mRNA level was upregulated(P<0.01),JAK2 and STAT3 mRNA levels were down-regulated(P<0.01),SOCS3 protein expression was up-regulated(P<0.01),and p-JAK2 and p-STAT3 protein expression levels were down-regulated
作者
周斌
万少兵
王瑛
余平
典万康
周莹
周琴
ZHOU Bin;WAN Shaobing;WANG Ying;YU Ping;DIAN Wankang;ZHOU Ying;ZHOU Qin(Department of Emergency Medicine,Wuhan Third Hospital,Wuhan 430060,China)
出处
《山西医科大学学报》
CAS
2023年第6期778-784,共7页
Journal of Shanxi Medical University
基金
武汉市卫健委医学科研项目面上项目(WX21B32)。
