The Role And Mechanism Of CFTR In Small Airway Mucus Obstruction Of Chronic Obstructive Pulmonary Disease

【Background】Chronic obstructive pulmonary disease（COPD）is a preventable and treatable disease characterized by continuous airflow limitation,due to the airway and/or alveolar abnormality,which is mainly caused by exposure to noxious particles or gases such as cigarette smoke.The spirometry-defined prevalence of COPD was 8.6%,accounting for almost 100 million Chinese adults,which seriously harms public health and brings a huge threat to the economy.The airflow obstruction of COPD mainly exists in the small conducting airways.Histologic data suggest that small airway abnormality may precede emphysema.In patients with COPD,the small airways are often tortuous and narrowed,and occluded by mucus plugs.Mucus plugs are a significant cause of obstruction of the small airways in some COPD patients.A failure of mucus transport,with mucus hypersecretion and mucus accumulation,contributes to sputum production,airflow obstruction,and exacerbations in COPD.Cystic fibrosis transmembrane conductance regulator（CFTR）is an anion channel located in the apical membrane of epithelial cells,mainly transports Cl^-and HCO₃^-.The acquired CFTR dysfunction of large airway results in a reduced capacity of the airway epithelium to secrete fluid,which leads to ASL dehydration,and together with mucin hypersecretion,is predicted to aggravate mucus hyperconcentration and plugging in CF and COPD.However,the role and mechanism of CFTR in small airway mucus obstruction of COPD remain unknown.【Objective】To investigate the role and potential mechanism of CFTR in small airway mucus obstruction of COPD.It could help us to better understand the pathogenesis of COPD and may also provide new therapeutic target for treating small airway mucus obstruction in COPD patients.【Methods】A COPD rat model was established by cigarette smoke exposure combined with intratracheal instillation of lipopolysaccharide（LPS）and the characteristics of this model were evaluated.Rats in the model groups were intratracheally dripped with LPS on Day1,Day14,Day28,and exposed to cigarette smoke 50 minutes per time,4 times per day,6 days per week for 4 months.Then,lung function of the rats was measured by Buxco PFT.Lung tissue sections were stained with HE,PAS and examined by light microscopy for histological sections.The mRNA levels of CFTR,MUC5AC,MUC5B in the small airway tissue samples were examined by Quantitative Real-time PCR.The pH of rat airway surface fluid（ASL）was measured by a fiber optic pH transmitter with micro pH sensor.An ex vivo culture model of rat small airway tissue was also established.Rat small airway was dissected under a stereomicroscope at a magnification of 10～20 with iris scissors and watchmaker forceps,and was exposured to cigarette smoke extract（CSE）.The mRNA levels of CFTR,MUC5AC,MUC5B in the small airway tissue samples were measured by Quantitative Real-time PCR.COPD rats were orally administered with curcumin to enhance CFTR expression once a day for 60 days from Day61 to Day120.At the end of the experiment,lung function of the rats was evaluated by PFT.The pathological morphology of lung tissue and goblet cell metaplasia of small airway were examined by HE and PAS staining.The mRNA expression of CFTR,MUC5B,NF-κB in the small airway tissue samples were measured by qPCR.And the ASL pH was measured by a fiber optic pH transmitter with micro pH sensor.【Results】1.Establishment of COPD rat model.1)Two groups of rats showed a similar initial body weight.At 4 months,compared with the normal control group,the rats of cigarette smoke combined with LPS（Smoke&LPS）group have lower body weights（P<0.0001）.2)Compared with the normal control group,FEV100/FVC and Mean Mid Expiratory Flow（MMEF）were decreased significantly in the Smoke&LPS group（P<0.01 or p<0.05）.The Inspiratory Resistance（RI）and RV/TLC in Smoke&LPS group were higher than the control group（P<0.01）.3)The pathological morphology of lung tissue was examined by HE and PAS staining.Histological analysis of the Smoke&LPS group revealed the presence of the COPD-like pathological morphological changes,such as emphysematous lesions,small airway wall thickening,loss of ciliated cells,increased small airway mucus secretion and goblet cell metaplasia.4)The total leucocyte number in BALF from Smoke&LPS group was significantly increased as compared to that of the control group（P<0.05）.2.The role of CFTR in small airway mucus obstruction of COPD.1)Compared with the control group,the MUC5AC and MUC5B mRNA levels of small airway tissue samples in COPD group were significantly increased（P<0.05）.2)Compared with the control group,the CFTR mRNA level of small airway tissue samples in COPD group was significantly decreased（P<0.01）.3)The ASL pH of COPD group was lower than the control group（P<0.001）.4)CFTR mRNA level was reduced in CSE-treated small airway tissues ex vivo as compared to the untreated controls（P<0.05）.MUC5AC and MUC5B mRNA levels were increased in the CSE-treated small airway tissues compared to the untreated controls as assessed by qPCR（P<0.05）.3.Curcumin enhances CFTR expression and ameliorates small airway mucus obstruction in COPD rats.1)Orally adminstrated curcumin effectively attenuates lung function decline of COPD rats,with FEV100/FVC and MMEF both significantly increased（P<0.01）,RI and RV/TLC significantly decreased（P<0.01）.Curcumin partially inhibited the small airway thickening and goblet cell metaplasia in COPD rats（P<0.001）.The MUC5B and NF-κB mRNA levels of small airway samples were also partially suppressed by curcumin（P<0.01 and P<0.05）.And the ASL pH were partially reversed by curcumin as well（P<0.05）.【Conclusions】1.A COPD rat model was successfully established by cigarette smoke exposure combined with intratracheal instillation of lipopolysaccharide.2.CFTR expression was decreased in COPD small airway tissues.3.Dysfunction of CFTR in COPD small airway promotes mucus hypersecretion and abnormal viscosity,likely through activation of NF-κB pathway and down-regulation of ASL pH,eventually contribute to small airway mucus obstruction.4.Curcumin enhances CFTR expression and ameliorates small airway mucus obstruction in COPD rats.