IL-27 Inhibit Pulmonary Fibrosis By Activating Autophagy Via The ERK/p38 Pathway

Objectives:Previous studies have confirmed that IL-27 can inhibit the formation of pulmonary fibrosis,and this experiment intends to further explore the mechanism of IL-27 in the formation of pulmonary fibrosis from the perspective of ERK/p38 pathway and autophagy regulation.Methods:60 healthy male adult C57/BL mice(7 to 8 weeks)to give free food and drink,and conventional breeding to observe three days(normal group)were randomly divided into NC group,the BLM group(bleomycin),BLM+IL-27 group(bleomycin+IL-27),the BLM+ RAPA group(bleomycin+rapamycin),the BLM+SP600125 group(bleomycin+ERK pathway inhibitor)and BLM+SB203580(bleomycin+p38 lightning pathway inhibitor),each group of 10,corresponding drug intervention was given according to the grouping,28 days after death in mice.1.NC group,the BLM group,BLM+IL-27 group and BLM+RAPA,Masson ’s staining method are used to observe the lung tissue pathology structure change,rt-pcr detection COL-Is COL-III collagen expression level,Western blot detection LC3B-I,LC3B-II and p62 autophagy related factors,to study role of IL-27 in regulating autophagy and pulmonary fibrosis.2.Expression levels of p-erk,ERK,p-p38 and p38 in lung tissues were detected by Western blot in NC group,BLM group,BLM+IL-27 group,BLM+RAPA group,BLM+SP600125 group and BLM+SB203580 group.To detect whether the ERK/p38 signaling pathway is involved in pulmonary fibrosis and the regulatory effect of IL-27 for the ERK/p38 signaling pathway.3.The NC group,the BLM group,the BLM+IL-27 group,BLM+SP600125 group and BLM+SB203580,Masson ’s staining method are used to observe the lung tissue pathology structure change,rt-pcr detection COL-I,COL-III collagen expression level,Western blot detection LC3B-Ⅰ,LC3B-II autophagy related factors,to study role of ERK/p38 signaling pathway in regulating autophagy and pulmonary fibrosis.Results:1.The effect of IL-27 on autophagy and pulmonary fibrosis in mice:Masson staining results showed that BLM-induced lung tissue sections of mice showed typical pulmonary fibrosis-like changes,with a large amount of damage to alveolar structure and significant thickening of residual alveolar septum.Multiple large areas of collagen deposition and fibrosis-like changes were observed in lung tissue sections.The alveolar structure of the IL-27 intervention group and the RAPA intervention group was basically intact,with partial collagen deposition,and the degree of pulmonary fibrosis was significantly reduced compared with that of the BLM group.Rt-pcr results confirmed that the content of COL-Ⅰ,Ⅲ in the lung tissues of BLM-treated mice was significantly increased(P<0.001),and the COL-1,111 expression was decreased after the action of IL-27 or RAPA compared with that of BLM group(P<0.001).Western blotting test results show that compared with the control group,the BLM processingin lung tissue of mice LC3B-II with LC3B-I ratio significantly decreased and the expression of p62 levels increased significantly(P<0.001),while the IL-27 or RAPA after treatment compared with BLM group LC3B-Ⅱ and LC3B-Ⅰ ratio significantly increased,the expression level of P62 was significantly decreased(P<0.001).The results showed that in blm-induced pulmonary fibrosis,autophagy was inhibited,and IL-27 and rapamycin could activate the expression of autophagy.2.The regulatory effect of IL-27 on ERK/p38 signaling pathway:Western blotting test results showed that,compared with the control group,the p-erk to ERK ratio and p-p38 to p38 ratio were significantly increased after BLM treatment(p<0.001).The phosphorylation levels of p38 and ERK in BLM group were significantly decreased after the combination of ERK pathway inhibitor(SP600125)or p38 pathway inhibitor(SB203580)with BLM(P<0.001).Phosphorylation levels of p38 and ERK were significantly decreased after IL-27 or RAPA treatment compared with BLM group(P<0.001).3.The effect of IL-27 regulation of ERK/p38 pathway on autophagy and pulmonary fibrosis:Masson’s results showed that the alveolar structure of the IL-27 intervention group was basically intact,and partial collagen deposition was observed.The degree of pulmonary fibrosis was significantly reduced compared with that of the BLM group.The alveolar structure was basically preserved in the ERK pathway inhibitor(SP600125)group or the p38 pathway inhibitor(SB203580)group,and the degree of pulmonary fibrosis was reduced compared with that of the BLM group.Rt-pcr results confirmed that COL-I,III expression in the IL-27 or ERK/p38 pathway inhibitor treatment group was lower than that in the BLM group(P<0.001).Western blotting test results show that compared with BLM group,IL-27 or p38 lightning/ERK pathway inhibitor treatment in mice after LC3B-II with LC3B-I ratio significantly increased(P<0.001).Conclusions:1.IL-27 significantly up-regulates the level of autophagy and inhibits BLM-induced pulmonary fibrosis.2.IL-27 can inhibit the activation of ERK/p38 signaling pathway in BLM-induced pulmonary fibrosis.3.IL-27 up-regulates the level of autophagy by down-regulating the ERK/p38 signaling pathway,and alleviates the progression of BLM-induced pulmonary fibrosis.