Study On The Effect Of Resveratrol On Alleviating Emphysema And Airway Inflammation Induced By Elastase And Atmospheric Particulate Matte

Background and Objective:Chronic obstructive pulmonary disease(COPD)is a heterogeneous disease characterized by chronic airway inflammation and destruction of the lung parenchyma(emphysema)as the main pathological features,the pathogenesis of which has not been fully elucidated and progress in treatment has been limited.With the advancement of pulmonary rehabilitation with integrated traditional Chinese and Western medicine in China,new progress has been made in the research of Chinese herbal medicines related to COPD.Resveratrol(Res)is a polyphenolic compound found in Chinese herbs such as thuja,quinoa,cassia,etc.,and related studies have shown its potential application value in COPD.However,studies on the interventional effects and mechanisms of resveratrol in COPD have been mainly based on mouse model induced by long-term cigarette smoke exposure,whether it has protective effects in mouse models of COPD with different phenotypes based on different exposure factors and its mechanism remain unclear.The receptor for advanced glycation end products(RAGE)is one of the pattern recognition receptors(PRR)expressed by airway epithelial cells and plays a key role in the development of airway inflammation and emphysema in COPD.Several studies have confirmed that resveratrol can regulate RAGE expression and attenuate the inflammatory response of mouse macrophages by inhibiting RAGE-mediated TLR4/NF-κB/ROS and MAPK/NF-κB inflammasome pathway.Such studies are more focused on diabetes and oncology,and the effect of resveratrol on RAGE expression and the regulatory mechanism in the field of respiratory diseases are not yet clear.Therefore,this study will focus on two important pathological features of COPD,emphysema and airway inflammation,and investigate the protective effects and mechanisms of resveratrol in different phenotypes and periods of COPD through a mouse model induced by porcine pancreatic elastase(PPE)and atmospheric particulate matter(PM),so as to provide new ideas for the prevention and treatment of COPD.Material and Methods:(1)In vivo,C57BL/6N mice were exposed to PPE(single dose or 1 dose/week for 4 weeks)or PM suspension(2 doses/week for 4 and 8 weeks)via intratracheal instillation to construct mouse models of emphysema phenotype and airway inflammation phenotype of COPD,respectively.PPE-induced mouse model groups:blank control group(PBS+Vehicle),blank administration group(PBS+Resveratrol-H),model control group(PPE+Vehicle),model administration low dose group(PPE+Resveratrol-L),model administration medium dose group(PPE+Resveratrol-M),model administration high dose group(PPE+Resveratrol-H),with 5-8 mice in each group(long-term model:n=6-8,short-term model:n=5-6).The PM-induced mouse models were grouped into the blank control group(PBS+Vehicle),model control group(PM+Vehicle),and model administration group(PM+Resveratrol),with 4-6 mice in each group(long-term model:n=3-5,short-term model:n=46).Mice in the model administration group were given low,medium and high dose of resveratrol solution(25,50 and 75 mg/kg)or medium dose resveratrol solution(50 mg/kg)by oral gavage at 2h and 24 h before intratracheal PPE or PM and daily from day 2 after modeling;mice in the blank administration group were given the same dose as those in the model administration high dose group(75 mg/kg);mice in the blank and model control groups were given equal amounts of CMC-Na or DMSO(Vehicle).Mice were executed 24 h after the final intervention and compared in general status,body weight,lung function(IC,Rn,Rrs,Ers,and Crs),lung histopathology(HE staining and Congo red staining),airway inflammation(macrophage,neutrophil,and eosinophil),adaptive immune cell levels(CD3+,CD3+ CD4+,CD3+ CD8+and CD3-B220+)and the expression of RAGE.Meanwhile,transcriptome RNA sequencing was performed using lung tissue.In addition,according to the above modeling approach,FPS-ZM1,the RAGE-specific inhibitor,was injected intraperitoneally after intratracheal PPE(1 time/day for 28 days),and the remaining two groups were given equal amounts of DMSO(Vehicle).The mice were executed 24 h after the final intervention,and the observation and detection indexes were as described above.(2)Clinical specimens were collected,and the expression of RAGE in peripheral blood and lung tissue of COPD patients and healthy subjects(healthy donors)were detected.Correlation analysis was performed on the lung function parameters FEV1%,FEV1/FVC and RAGE mRNA expression levels in peripheral blood of COPD patients.Results:(1)The experiment showed that emphysema related or airway inflammatory phenotype of COPD related pathophysiological changes could be seen 4 days after a single intratracheal PPE or 2 months after continuous intratracheal PM,including changes in general status(positive symptoms such as wasting,mental depression,unresponsiveness,arching back curl,and slow movement),decreased lung function(IC,Rn,Rrs,and Ers elevated,Crs decreased),histopathological changes(collapse and fracture of alveolar wall,formation of large alveoli;or airway mucosal folds,mucociliary lodging,epithelial cell proliferation and shedding,airway smooth muscle thickening,etc.)and airway inflammation(total cells,eosinophil,macrophage,and neutrophil count in BALF significantly increased;a large number of eosinophil infiltration can be seen around airways and blood vessels;or macrophage activation and phagocytosis of PM).Resveratrol has protective effects on PPE-induced mouse models in different periods by improving lung function,reducing alveolar wall destruction and reducing airway inflammation(medium and high dose).Resveratrol has anti-and pro-inflammatory bidirectional regulatory effects,which the medium and high dose of resveratrol can inhibit the activation,recruitment,and infiltration of inflammatory cells in BALF and lung tissue,while low dose of resveratrol can further aggravate airway inflammation;Resveratrol may inhibit PPE-induced emphysema and airway inflammation in experimental mice by regulating expression of miRNA and the genes involved in airway inflammation and oxidative stress in COPD.Resveratrol showed a tendency to improve lung function and histomorphological destruction in mice with PM-induced airway inflammation phenotype,but had no effect on antiinflammation and regulation of adaptive immune cells level.(2)Clinical data showed that the expression of RAGE mRNA and RAGE protein were increased in COPD patients,and the level of RAGE mRNA was negatively correlated with pulmonary function parameters FEV1%and FEV1/FVC.RAGE-specific inhibitor(FPS-ZM1)was protective against PPE-induced emphysema phenotype in mice by reducing alveolar wall destruction,decreasing airway inflammation and increasing sRAGE protein expression.However,the effect of resveratrol on RAGE expression is unclear.Conclusions:(1)Resveratrol has protective effect on PPE-induced emphysema phenotype in a mouse model of COPD(COPD-like)in different periods by regulating the expression of miRNA and the genes(S100a9、Cxcl、Ccl、Mmp12)involved in airway inflammation and oxidative stress and miRNA-mRNA regulatory networks to reduce airway inflammation and improving lung function;Resveratrol has no protective effects on the PM-induced airway inflammation phenotype in a mouse model of COPD.(2)RAGE expression was increased in COPD patients and negatively correlated with lung function;Reducing RAGE expression could partially inhibit PPE-induced emphysema and airway inflammation;The effect of resveratrol on RAGE expression in a mouse model of COPD with PPE-induced emphysema phenotype needs further validation.