The Mechanism Of Wnt/β-catenin In Exercise Improving COPD Diaphragm Dysfunction

ObjectiveChronic obstructive pulmonary disease(COPD)is characterized by progressive airflow limitation with low systemic inflammation and continued deterioration results in dyspnea.Diaphragm dysfunction is an important cause of dyspnea and is closely related to inflammation.Exercise can inhibit systemic inflammation in COPD and effectively improve diaphragm dysfunction,but the mechanism is unclear.Wnt/β-catenin is a target for exercise,which inhibits inflammation and promotes muscle repair.In addition,chemerin/CMKLR1 is a potential downstream signaling pathway for Wnt/β-catenin.Therefore,on the premise of exercise regulating inflammation to improve COPD diaphragm dysfunction,this study investigated the role of Wnt/β-catenin in this process and explored whether chemerin/CMKLR1 is regulated by Wnt/β-catenin,further revealing the mechanism of exercise improving COPD diaphragm dysfunction.MethodsThirty-six 8-week-old C57BL/6J male mice were randomly divided into control group(CG),model group(MG),agonist group(AG),agonist exercise group(AEG),inhibitor group(IG)and inhibitor exercise group(IEG),with six mice in each group.Except for CG,all groups of mice underwent 25 weeks of incremental whole-body cigarette smoke exposure to establish the COPD model.After model establishment,AEG and IEG perform the adaptive exercise for a week.Subsequently,AG was injected intraperitoneally with Li Cl(a Wnt agonist);IG was injected intraperitoneally with XAV939(a Wnt inhibitor);AEG was injected intraperitoneally with Li Cl followed by exercise intervention;IEG was injected intraperitoneally with XAV939 followed by exercise intervention,and the above drug and exercise interventions lasted for eight weeks.MG did not make any intervention after the completion of modeling,and CG did not make any intervention throughout the experiment.After the intervention,the diaphragm of mice in each group was divided into three parts:(1)to observe the diaphragm morphological changes of mice by HE staining,(2)for isolated contraction muscle strength test,(3)to detect the expression of Wnt1,β-catenin,chemerin,and CMKLR1 in the diaphragm by Western blot;detection of inflammatory markers(TNF-α,IL-1β,and IL-8)in serum by ELISA.Results(1)Diaphragm HE staining.Compared to CG,the MG mice diaphragm showed pathological manifestations such as disturbed fiber arrangement,partial breakage,and increased fiber gaps,while the fiber cross-sectional area decreased but was not significant.Compared with MG,the diaphragm of AG mice showed significantly improved fiber disorder and damage,and the fiber cross-sectional area was increased significantly(p = 0.008);the diaphragm of IG mice had similar histology as MG,and the fiber cross-sectional area decreased but did not reach statistical difference.Compared with AG,AEG mice showed less diaphragm fiber damage and abnormal tissue proliferation,more orderly and tight fiber arrangement,and a significant increase in diaphragm cross-sectional area(p= 0.01).Compared with IG,IEG mice had narrower diaphragmatic fiber gaps,significantly more nuclei,and a significant increase in fiber cross-sectional area(p < 0.001).(2)Diaphragm isolated contraction muscle strength test.Compared with CG,MG mice showed a decrease in diaphragm contractile muscle strength;compared with MG,AG mice showed an increase in diaphragm muscle strength;when comparing AEG with AG and IEG with IG,diaphragm contractile muscle strength was increased,but none of these changes were statistically significant.Only when AEG was compared with MG was a significant increase in diaphragmatic contractile muscle force(p = 0.0198).(3)Expression of Wnt1,β-catenin,chemerin,and CMKLR1 proteins in the diaphragm.Compared with CG,the diaphragm of MG mice showed a decreasing trend in Wnt1 protein expression and a significant increase in chemerin protein expression(p < 0.001).Compared with MG,the diaphragm of AG mice showed a relative increase in Wnt1 protein expression and a significant decrease in chemerin protein(p < 0.001);the diaphragm of IG mice showed a relative decrease in Wnt1 protein expression.When comparing AEG with AG and IEG with IG,the diaphragm both showed an increasing trend in Wnt1 protein expression.When IEG was compared with IG and MG separately,the diaphragm of mice showed a significant decrease in chemerin protein(p < 0.001).Wnt1 protein expression in the diaphragm was only significantly different when comparing AEG with IG(p = 0.0198),and no statistical differences were seen between the other two groups.No statistical differences in β-catenin and CMKLR1 protein expression were observed when compared among the six groups or between the two groups separately.(4)Serum levels of inflammatory markers(TNF-α,IL-1β,and IL-8).Compared with CG,serum levels of TNF-α(p = 0.0206)and IL-1β(p = 0.0139)were significantly increased,while serum levels of IL-8 were relatively higher in MG mice.Compared with MG,serum levels of TNF-α,IL-1β,and IL-8 were reduced in both AG and IG mice,and the reduction was more pronounced in AG mice,but none reached statistical differences.Compared with AG,AEG showed reduced but non-significant levels of TNF-α,IL-1β,and IL-8,and AEG showed a significant reduction in IL-1β levels only when compared with MG(p = 0.0147).ConclusionExercise may improve COPD diaphragm dysfunction by activating Wnt/β-catenin to reduce expression levels of chemerin and circulating levels of inflammatory factors.