The Role And Mechanism Of E3 Ubiquitin Ligase Cbl-b In COPD With Depression

【Background】Depression is one of the most common complications of COPD.Depression in COPD can lead to the decline of quality of life,aggravation of dyspnea,increase of acute exacerbation,increased risk of hospitalization and decreased compliance with treatment,which seriously affect the quality of life of patients and cause huge economic burden to the family and society.At present,more and more attention has been paid to the clinical significance of COPD with depression,but its related basic research is less,and the pathobiological mechanism of COPD with depression is not clear.Cbl-b is an E3 ubiquitin ligase,which plays an important role in regulating anti-inflammatory.Inhibition of Cbl-b expression leads to production of proinflammatory cytokines.Previous studies of our group have shown that,during the occurrence and development of COPD,Cbl-b can inhibit CS-induced macrophage infiltration in lung tissue,which has a significant anti-inflammatory effect.Since neuroinflammation is one of the main mechanisms of depression,it is inferred that Cbl-b plays an important role in the pathogenesis of COPD complicated with depression induced by CS exposure.【Objectives】In this study,Cbl-b^-/-mice and cell model combined with molecular biology techniques were used to study whether chronic tobacco smoke exposure could induce COPD with depression in mice,and to study the role and mechanism of Cbl-b in COPD with depression induced by tobacco smoke exposure.【Methods】1.Whether chronic exposure to tobacco smoke can induce COPD combined with depression in mice:1)A stable COPD mouse model was established by continuous exposure to tobacco smoke for 24 weeks.2)The changes of FRC,FVC,FEV20/FVC,FEV50/FVC and FEV100/FVC were detected by pulmonary function test.3)The occurrence of depression was judged by behavioral evaluation and Elisa to detect the changes of 5-HT and corticosterone in plasma.4)The morphological changes of alveoli and airways and the degree of inflammatory infiltration were observed by hematoxylin-eosin staining in lung tissue of mice,and analyzed the mean alveolar intercept.5)The contents of IL-1β,IL-6 and KC in BALF were detected by Elisa.2.The role and mechanism of Cbl-b in COPD with depression caused by tobacco smoke exposure:1)The changes of morphology and number of nerve cells were observed and compared by hematoxylin-eosin staining and nissl staining in mouse brain tissue.2)Immunohistochemical detection of astrocyte marker GFAP activation in mouse brain.3)Immunofluorescence detection of microglia marker Iba1 activation in mouse brain.4)The expression of Cbl-b in mouse brain was determined by Western blotting,and the changes of p-ERK,p-JNK and other proteins were detected.3.Cigarette smoke extract stimulated mouse cell line BV-2 to construct neuroinflammatory cell model Mouse microglial cell line BV-2 was stimulated by different concentrations of CSE for 24 hours to establish an acute inflammatory cell model.Western blotting was used to detect the expression of i NOS,HO-1,NQO1,p-ERK and other proteins in BV-2 cells.【Results】1.Chronic exposure to tobacco smoke induces COPD with depression in mice:1)Tobacco smoke exposure induced an increase in pulmonary function parameters FRC and FVC and a significant decrease in respiratory velocity parameters FEV20/FVC,FEV50/FVC and FEV100/FVC in Cbl-b^-/-mice.2)Tobacco smoke exposure induced more obvious depression-like behavior in Cbl-b^-/-mice,and the content of 5-HT in plasma decreased significantly and corticosterone in plasma increased significantly.3)Tobacco smoke exposure induced significant emphysema and airway inflammation in Cbl-b^-/-mice,and the average intercept of alveoli increased.4)Tobacco smoke exposure induced increase of cytokines IL-1β,IL-6 and KC in BALF of Cbl-b^-/-mice.2.The role and mechanism of Cbl-b in COPD complicated with depression caused by tobacco smoke exposure:1)Significant morphological changes in Cbl-b^-/-mice induced by smoke exposure.2)Activation of astrocytes in hippocampal CA3 region and dentate gyrus.3)Increased expression of microglia marker Iba1 in hippocampal CA3 region and cerebral cortex.4)Tobacco smoke exposure increased the expression of Cbl-b protein in the brain of mice,and smoke exposure increased the expression of p-ERK and p-JNK in the brain of Cbl-b^-/-mice.3.The neuroinflammatory cell model was successfully established by stimulating BV-2 cells with 0.5%and 0.75%CSE for 24 hours,and the protein expression of i NOS,HO-1,NQO1 and p-ERK increased.【Conclusion】Chronic exposure to tobacco smoke could induce COPD with depression in mice,and the phenotype of Cbl-b^-/-mice was more significant.The mechanism may be related to the knockout of Cbl-b which regulates the expression of p-ERK and p-JNK proteins and aggravates the morphological changes of nerve cells and the activation of microglia in the brain.