IL-37 Inhibits Invasion And Metastasis In Non-small Cell Lung Cancer Through Suppression Of The IL-6/STAT3 Signaling Pathway

Objective:To investigate the effects of interleukin-37(IL-37)on the invasion and metastasis of non-small cell lung cancer(NSCLC)and its possible molecular mechanisms.Methods:Forty NSCLC patients with no history of surgery,radiothrapy,chemotherapy and biological therapy were enrolled as NSCLC group,and forty health physical volunteers were enrolled as healthy controls.Enzyme-linked immunosorbent assay(ELISA)was used to detect plasma IL-37 expression in two groups and the correlation of IL-37 level and clinical characteristics in NSCLC patients was analyzed.In vitro,the NSCLC cell,A549 cells were cultured and was treated with recombinant human IL-37protein(rhIL-37)in different concentrations(0ng/ml,10ng/ml,100ng/ml and 500ng/mL),the invasion and metastasis of A549 cells were detected using Transwell ~TMM invasion and scratch wound healing assays and the appropriate concentration of IL-37 was selected;A549 cells were cultured and was treated with recombinant human IL-6 protein(rhIL-6)in different concentrations(0ng/ml,10ng/ml,50ng/ml and 100ng/mL),the invasion and metastasis of A549 cells were detected using Transwell ~TMM invasion and scratch wound healing assays and the appropriate concentration of IL-6 was selected;A549 cells were randomly divided into fort groups(control group,IL-6group,IL-37 group and IL-6+IL-37group).A549 cells in IL-6 group and IL-6+IL-37 group were cultured with IL-6 in appropriate concentration,and A549 cells in IL-37 group were cultured with IL-37 in appropriate concentration for 24 hours,then IL-37 in appropriate concentration was added into IL-6+IL-37 group and A549 cells in all groups were cultured for 24 hours.The invasion and metastasis of A549 cells in four groups were detected using Transwell~TMM invasion and scratch wound healing assays,protein expression of STAT3,pSTAT3,E-cadherin,Vimentin and N-cadherin were detected using Western blotting,and mRNA expression of STAT3,E-cadherin,vimentin and N-cadherin was assessed in each group using RT-PCR.Results:IL-37 plasma expression was decreased in NSCLC patients compared with the healthy controls(P<0.05),and the down-regulation of IL-37 was correlated with tumor stage(P<0.05),and has no correlation with age,sex and smoking history in patiens and pathological types of NSCLC.In vitro,the results of Transwell~TMM invasion and scratch wound healing assays showed that compared with control group(IL-37 in 0ng/ml),IL-37in 100ng/ml and 500ng/mL inhibited the invasion and migration of A549 cells(P<0.05).Compared with control group(IL-6 in 0ng/ml),IL-6 in 500ng/ml and 100ng/mL promoted the invasion and migration of A549 cells(P<0.05).The concentration of50ng/ml and 100ng/ml were selected as the appropriate concentration of IL-6 and IL-37,respectively.Infourgroups(controlgroup,IL-6group,IL-37groupand IL-6+IL-37group),the results of Transwell ~TMM invasion and scratch wound healing assays showed that compared with IL-6 group,the invasion and migration of A549 cells in IL-6+IL-37 group were inhibited(P<0.05).The results of RT-PCR showed that vimentin and N-cadherin mRNA expression were increased,STAT3 and E-cadherin mRNA expression were decreased in IL-6 group compared with control group(P<0.05);while vimentin and N-cadherin mRNA expression were decreased,STAT3 and E-cadherin m RNA expression were increased in IL-6+IL-37 group compared with IL-6 group(P<0.05).The results of western blot showed that pSTAT3,vimentin and N-cadherin protein expression were increased,STAT3 and E-cadherin protein expression were decreased in IL-6 group.However,the opposite pattern was observed in the IL-37+IL-6group,inhibiting the activation of STAT3 and EMT induced by IL-6.Conclusions:Our results showed that IL-37 plays an inhibitory role in NSCLC progression,possiblybysuppressingSTAT3activationanddecreasing epithelial-mesenchymal transition(EMT)by inhibiting IL-6 expression.IL-37 could serve as a potential tumor suppressor in NSCLC.