Study On The Effects Of Modulation Of Wnt Signaling Pathway In Paraquat-induced Pulmonary Fibrosis In Rats

ObjectiveTo explore the role of Wnt signaling pathway in paraquat-induced pulmonary fibrosis in rats model,and further elucidate the pathogenesis mechanism of paraquat-induced pulmonary fibrosis,regulate the Wnt signaling pathway in pulmonary fibrosis in order to provide a new idea for new drug and treatment of PQ poisoning.Methodsexperiment one:A total of 36 male Sprague Dawley(SD)rats were randomly divided into two groups(n=18 for each group):Control group,PQ poisoning group.The PQ poisoning group received a single intraperitoneal injection of PQ(20 mg/kg),the Control group received an equivalent volume of sterile saline.Two groups of rats were respectively sacrificed six on 7 days,14 days,28 days,then taken out their lung tissue,the pathological changes were assessed by HE staining and Masson staining.Use Acid hydrolysis method to measure HYP content.Protein microarray was used to detect the expressions level of some protein.β-catenin and MMP-2,which are the key protein of Wnt signaling pathway,were detected by Western Blot.Occludin,E-cadherin and a-SMA,vimentin were detected by immunohistochemical staining.experiment two:A total of 54 male Sprague Dawley(SD)rats were randomly divided into three groups(n=18 for each group):Control group,PQ poisoning group and Inhibitor group.The PQ poisoning group received a single intraperitoneal injection of PQ(20 mg/kg),the Control group received an equivalent volume of sterile saline,while the Inhibitor group received a single intraperitoneal injection of PQ(20 mg/kg)and DKK1(2mg/kg,100p 1 PBS,Continuous dosing for a week),which is the specific inhibitors of Wnt signaling pathway.Three groups of rats were respectively taken six on 7 days,14 days,28 days,Puncture of femoral artery blood line for blood gas analysis,then sacrificed and taken out their lung tissue,the pathological changes were assessed by HE staining and Masson staining.Use Acid hydrolysis method to measure HYP content.β-catenin and MMP-2,which are the key protein of Wnt signaling pathway,were detected by Western Blot.Occludin,E-cadherin and a-SMA,vimentin were detected by immunohistochemical staining.Resultsexperiment one:1.PQ group rat lung gross specimens show point shape hemorrhage stove,a large number of white nodules,hard quality of material.HE staining results indicated that structural injury of the lung tissues,the alveolar interval was more broadening and bodiness.Masson staining results indicated that a lager of blue fiber cords deposition,but the control group were normal.2.Compared with control group,the HYP content of PQ-7d does not increase,but the HYP content of PQ-14d increased significantly,as the extension of infected time,HYP content increased gradually,there were statistically significant differences(P<0.05).3.RayBio Protein microarray results demonstrated that the expression level of IL-1β,PDGF,β-catenin,MMP-2 and TGF-β 1 increased significantly compared with Control group.4.Western Blot results indicated that the expression level of β-catenin and MMP-2 were significantly increased in PQ poisoning group compared with Control group.Immunohistochemical staining indicated that occluding,E-cadherin were down regulated and α-SMA,vimentin were up regulated.experiment two:1.The blood gas analysis indicated that PaO2、SpO2 were significant descended and PaCO2 was significant rised in PQ group.Compared with PQ group,PaO2、SpO2 were significant rised and PaCO2 was significant descended in Inhibitor group,there werestatistically significant differences(P<0.05).2.HE staining and Masson staining results indicated that structural of the lung tissues was less injury,and blue fiber cords was less deposition in Inhibitor group.3.Compared with PQ group,the HYP content of Inhibitor group descended significantly,Western Blot results indicated that the expression level of β-catenin and MMP-2 were significantly descended in Inhibitor group.4.Immunohistochemical staining indicated that,Compared with PQ group,occluding,E-cadherin were up regulated and α-SMA,vimentin were down regulated in Inhibitor group.Conclusion1.A single intraperitoneal injection of PQ can establish a stable model of pulmonary fibrosis,Wnt signaling pathways involved in the process of PQ induced pulmonary fibrosis in rats.2.The PQ may occur through activation of Wnt signaling pathways to induce lung epithelial cells of EMT and to promote the formation of fibroblast,Inhibit of Wnt signaling pathway can reduce the degree of PQ poisoning induced pulmonary fibrosis in rats,in order to provide a new idea for new drug and treatment of PQ poisoning.