Role Of STAT3 In IL-6-Induced Epithelial-Mesenchymal Transition In Human Peritoneal Mesothelial Cells

Background It was found that with the prolongation of peritoneal dialysis(PD)time in patients who have end-stage renal disease(ESRD),the morphology and function of peritoneum will change,which leading to peritoneal fibrosis(PF)and ultrafiltration failure(UFF).At present,UFF has become the main reason why PD patients are forced to withdraw from PD,so it is particularly important to explore and find ways to alleviate peritoneal fibrosis and delay the occurrence of UFF in peritoneal dialysis.Objective In this study,human peritoneal mesothelial cells(HPMCs)were stimulated by IL-6 to induce epithelial-mesenchymal transformation(EMT),and the effect of signal transducer and activator of transcription 3(STAT3)on epithelial-mesenchymaltransformation was investigated.Methods Human peritoneal mesothelial cells((HPMCs)were cultured in vitro and stimulated with different concentrations of IL-6 for different times.The cells were grouped as follows:(1)HPMCs were stimulated with 50 ug/L IL-6 at different times,and divided into groups according to stimulation time of 0 h,24 h,48 h and 72 h;(2)HPMCs were stimulated with different concentrations of IL-6 for 24 hours,and grouped with 0 ug/L,50 ug/L and 100 ug/L.HPMCs cell lines which stably silenced STAT3 gene were cultured by lenti-virus vector mediating RNA interference technique and grouped into:(1)control group;(2)IL-6 group;(3)empty vector group;(4)empty vector + IL-6 group;(5)lentivirus infection group;(6)lentivirus infection + IL-6 group.The expression and distribution of E-cadherin and alpha-smooth muscle actin(alpha-SMA)which are EMT-related proteins were detected by immunofluorescence.Western blot and real-time polymerase chain reaction(qRT-PCR)were used to detect Janus kinase 2(JAK2)/STAT3 signal transduction pathway proteins and EMT-related proteins and genes expression.SPSS 22.0 software was used to analyze the experimental data.Results Compared with the control group,with the prolongation of IL-6 stimulation time and the increase of IL-6 concentration,the expression of E-cadherin decreased(P < 0.05),the expression of alpha-SMA,vascular endothelial growth factor protein(VEGF)increased(P < 0.05),and the expression of pathway protein p-JAK2/JAK2 increased(P < 0.05).The ratio of p-STAT3/STAT3 increased(P <0.05).When STAT3 gene was silenced by lentivirus,the expression of alpha-SMA and VEGF in lentivirus infection + IL-6 group decreased(P < 0.05),while the expression of E-cadherin increased(P < 0.05).The results of cellular immunofluorescence showed that in the control group,a small amount of alpha-SMA was expressed in the cytoplasm,while E-cadherin was expressed in the cell membrane.Compared with the control group,the expression of alpha-SMA in the cytoplasm increased and the expression of E-cadherin on the cell membrane decreased after stimulating HPMCs 24 hours by 50 ug/L IL-6.Conclusion IL-6 can activate JAK2/STAT3 signaling pathway and participate in the process of EMT and stimulate the secretion of vascular endothelial growth factor(VEGF)in HPMCs.The silencing of STAT3 gene by lentivirus can inhibit the occurrence of EMT in human peritoneal mesothelial cells.