Effects Of Air Pollutants On Hospitalization Of Chronic Obstructive Pulmonary Disease And IL37 Effect And Mechanism On Fine Particulate Matter Induced Extracellular Matrix Deposition In Bronchial Epithelium

Chronic obstructive pulmonary disease(COPD)is a common disease that seriously endangers human health.COPD have been an increasingly important global public health problem.It is estimated that there are about 70 million COPD patients world wild.According to Wang Chen's epidemiological survey of chronic obstructive pulmonary disease in China between 2012 and 2015,the number of COPD in China is about 100 million.The prevalence rate in adults over 20 years old was 8.6%,and the prevalence rate of COPD is 13.7%among people over 40 years old in China.It has become a chronic disease equal to hypertension and diabetes,constituting a major disease burden.A large proportion of deaths from chronic obstructive pulmonary disease occur during acute exacerbation and in subsequent weeks,especially in hospitalized patients with severe acute exacerbation.Exacerbation is one of the most important causes for mortality associated with COPD.AECOPD accounts for the largest proportion of the total COPD burden in the healthcare system and it is projected to be the 3rd leading cause of death by 2020.Bacterial and viral infections,exposure to tobacco,some workplace dusts and fumes,weather and air quality may be key factors contributing to deterioration of COPD symptoms.Recent epidemiological investigations also reported that environmental factors such as ambient PM2.5 and PM10 contribute to COPD onsets and mortality.PM2.5,PM10,NO2,SO2 and O3 are closely related to the occurrence and development of acute exacerbation of COPD.But the specific mechanism remain unclear.Repeated or sustained exposure of these substances to airway epithelium causes epithelial damage,activating various signaling pathways for repair of the damage to maintain its integrity.WNT(Wingless/Int)signaling activation may lead to abnormal production of TGF-? and induction of epithelial-mesenchymal transition in epithelial cells,regulating tissue repair and extracellular matrix deposition,such as fibronectin,matrix metalloproteinase and so on.At present,the effect of air pollutants on acute exacerbations of chronic obstructive pulmonary disease is mainly focused on epidemiological investigation.Most previous studies have commonly used the moving average method,accounting for the lag effects of pollutants on Acute Exacerbations of Chronic obstructive pulmonary disease(AECOPD)-associated hospitalizations;however,this method underestimated the cumulative effect.Therefore,we adopted an advanced method,the poison generalized linear regression model combined with the distributed lag nonlinear model,to estimate the relative cumulative risk and to calculate the attribution of air pollutants leading to acute exacerbation of hospitalization for chronic obstructive pulmonary disease.To further analyze the effects of air pollutants from coal burning on AECOPD and the cumulative relative risk of air pollutants to AECOPD hospitalization and the difference of hospitalization burden in winter and spring,we divided the whole time series into heating season and non-heating season.In basic research,we study whether PM2.5 can cause extracellular matrix deposition of bronchial epithelial cells and how its pathogenesis needs further study.WNT-5A are widely studied nonclassical Wnt signaling ligands.Besides promoting inflammatory response,WNT-5A is also involved in extracellular matrix deposition,airway remodeling.WNT-5A can induce extracellular matrix deposition in asthma,pulmonary interstitial fibrosis have been reported,the pathogenesis in COPD is still unclear,and there are few literature reports on WNT-5A and PM2.5 induced extracellular matrix deposition.Moreover,what pathways WNT-5A regulate PM2.5 induced extracellular matrix deposition remains unclear and further studies are needed to clarify.Our research is divided into three parts:part I,we use the Poisson generalized linear regression model and the distributed lag nonlinear model to analyze the main relative cumulative risk factors of air pollutants on hospitalization for chronic obstructive pulmonary disease at high air pollution levels in Shijiazhuang city,and calculate the burden of hospital attribution.To find susceptible populations,we stratified analysis by age and sex.Reduce the risk of acute exacerbation of chronic obstructive pulmonary disease and reduce the economic burden in hospital by controlling pollutant levels.The second part mainly studies the difference between air pollutants in heating and non-heating season,and analyzes the relative cumulative risk factors and attribution burden of air pollutants on hospitalization for chronic obstructive pulmonary disease in different seasons.Part ? is the basic research.We proposed to use suspended particulate matter PM2.5 with aerodynamic diameter<2.5?m as a study object to observe the effect of PM2.5 on extracellular matrix deposition in human bronchial epithelial cells to further analyze the changes of WNT-5A expression levels in extracellular matrix deposition.Effect of blocking WNT-5A function on PM2.5 induced extracellular matrix deposition in bronchial epithelial cells and explore the relationship between WNT-5A and oxidative stress,inflammatory response signaling pathway.Part One Air Pollutants association with hospitalization and attribution burden for chronic obstructive pulmonary disease in Shijia zhuangObjective:To analyze the main relative cumulative risk factors of chronic obstructive pulmonary disease hospitalization caused by air pollutants at high air pollution levels in Shijiazhuang city,and calculate the burden of hospital attribution.We investigated susceptible people based on sex and retirement status.It can reduce acute exacerbation of hospital risk and economic burden in chronic obstructive pulmonary disease by controlling levels of air pollutants.Methods:Data on hospitalization for AECOPD,air pollution and meteorological factors from 1 January2013 to 31 December 2016 were collected in Shijiazhuang,China.We used a poisson generalized linear regression model combined with a distributed lag non-linear model(DLNMs)to evaluate the relative cumulative risk for lag 0-7 days and examined the potential effect modification by age,gender via stratification analyses,controlling for long-term trends,seasonal patterns,meteorological factors and other possible confounders.Then,we computed hospitalizations attributable percentage for air pollutants.Results:1.Between 2013 and 2016,a total of 9358 patients(male:female,2.61:1)with AECOPD lived in the Shijiazhuang urban area.In this study,there were approximately 6 cases of AECOPD every day.2.The AECOPD-associated relative cumulative risks for PM2.5,PM10,NO2,SO2,and CO for lag0-7 days were significantly positively correlated with hospitalization.However,for 03,a negative linear correlation was observed.3 Increases of 10?g/m3 in PM2.5,PM10,SO2,NO2 and CO levels were associated with 2.4%(95%CI(1.015-1.032))?1.4%(95%CI(1.008-1.02))?3.4%(95%CI(1.013-1.055))?6.7%(95%CI(1.04-1.096))?1.8%(95%CI(1.011-1.024))increases COPD hospitalizations in the retired patients,Which were larger than unretired patients.The NO2 Cum RR of AECOPD admission was the greatest at 6.7%.4.Increases of 10 ?g/m3 in PM2.5,PM10,SO2,NO2,CO and O3 levels were associated with 2.1%(95%CI(1.006-1.035))?1.3%(95%CI(1.003-1.023))?2.6%(95%CI(0.993-1.06))?5.7%(95%CI(1.013-1.103))?1.6%(95%CI(1.005-1.028))?1.6%(95%CI(0.987,1.046))increases COPD hospitalizations in the female patients,Which were larger than male patients.The NO2Cum RR of AECOPD admission was the greatest at 5.7%.5.The NO2Cum RR of AECOPD admission was the greatest.10?g m3 increases in daily NO2 concentration was associated with a 6.7%and 5.7%increase in COPD hospitalizations in the retired and female group,respectively.6.13%(95%Cl(5.5-20.4%or 1220 admissions)),9.4%(95%CI(-1.3-18.4%or 879 admissions)),1.7%(95%Cl(-10.9-12.6%or 156 admissions)),9.7%(95%Cl(-3.4-21.2%or 905 admissions)),and 8.8%(95%Cl(-0.6%-17.3%or 828 admissions))of the COPD hospitalizationswere attributable to PM2.5,PM10,SO2,NO2,and CO exposure,respectively.7.If the air pollutant levels decreased to the 24-h average grade ? levels of NAAQS of China,the reductions in the AECOPD-associated attributable percentages for PM2.5,PM10,SO2,NO2,and CO would be approximately 11.2%(95%CI(0.047,0.202),7.5%(95%CI(-0.010,0.182),0.5%(95%CI(-0.131,0.126),3.9%(95%CI(-0.035,0.222)and 3.2%(95%CI(0.000,0.177)respectively.Summary:1.The AECOPD-associated relative cumulative risks for PM2.5,PM10,NO2,SO2,and CO for lag0-7 days were significantly positively correlated with hospitalization.However,for O3,a negative linear correlation was observed.2.The associations were differed by individual characteristics.The retired and female population were highly vulnerable.3.13%,9.4%,1.7%,9.7%,and 8.8%of the COPD hospitalizations were attributable to PM2.5,PM10,SO2,NO2,and CO exposure,respectively.4.If the air pollution concentration was reduced to the grade ? levels of NAAQS,the AECOPD attributable percentage for PM2.5 and PM10 would decrease by 80%.Part Two To compare the effects of air pollutants on hospitalization relevance and burden of chronic obstructive pulmonary disease between heating and non-heating seasonObjective:To study the difference between air pollutants in heating season and non-heating season,and to analyze the relative cumulative risk factors and attribution burden of air pollutants on chronic obstructive pulmonary disease hospitalization in different seasons.Methods:The whole time series was split into heating season and non-heating season.We used a quasi-poisson generalized linear regression model combined with distributed lag non-linear models(DLNMs)to estimate the relative cumulative risk and calculate the air pollutant hospitalization burden of AECOPD for lag 0-7 days in heating season and non-heating season controlling for long-term trends,seasonal patterns,meteorological factors and other possible confounders.Results:1.There were higher PM2.5,PM10,NO2,SO2,and CO concentrations in heating seasons(daily average162.22+120.63?g/m3?263.16±164.32?g/m3?73.29±31.5?g/m3?106.49±74.65?g/m3?2.49±1.75 mg/m3)than non-heating season(84.13±60?g/m3?164.54±74.65?g/m3?46±20.6?g/m3?34.19±25.97?g/m3?1.0±0.5mg/m3)in Shijiazhuang;However,O3 was higher in non-heating season(the maximum 8-h moving average concentrations 109.54+50.2?g/m3)than heating season(39.79+27.83 ?g/m3).2.The AECOPD-associated relative cumulative risks for PM2.5,PM10,NO2,and SO2 for lag 0-7 days were significantly positively association with hospitalization in heating and non-heating season;however,that for O3 exhibited negative correlation in heating season and positive correlation in non-heating season and that for CO exhibited positive correlation in heating season and negative correlation in non-heating season.3.A 10 ?g/m3 increase of PM2.5,PM10,SO2,and NO2 levels was associated with a 1.0%(95%CI(0.997,1.022),0.5%(95%CI(0.995,1.014),0.2%(95%CI(0.973,1.031),and 2.2%(95%CI(0.982,1.064)increase COPD hospitalizations in heating season,respectively,over a lag 0-7 days,but O3 with a 1.3%(95%CI(0.916,1.062)decrease COPD hospitalizations.An increase of 0.1 mg/m3 in CO levels was associated with a 0.3%(95%CI(0.994,1.012)increase COPD hospitalizations in heating season;However,we found that 10?g/m3 increase of PM2.5,PM10,SO2,NO2,and O3 levels was associated with a 2.6%(95%CI(1.008,1.045),1.6%(95%CI(1.004,1.028),5.3%(95%CI(0.991,1.118),1.9%(95%CI(0.963,1.079),and 1.0%(95%CI(0.989,1.032)increase COPD hospitalizations in non-heating season,respectively,over a lag 0-7 days.An increase of 0.1 mg/m3 in CO levels was associated with 0.3%(0.974,1.022)decrease COPD hospitalizations in non-heating season.4.17.8%(95%CI(-0.34-34%or 743 admissions)),12.9%(95%CI(-15.6-32%or 538 admissions)),1.7%(95%Cl(-44.7-28.7%or 72 admissions)),16.7%(95%CI(-13.8-38.3%or 698 admissions)),10.5%(95%CI(-13.8-28%or 441 admissions))and-4%(95%Cl(-42-19.1%or-168 admissions))of AECOPD hospitalizations could be attributable to PM2.5,PM10,SO2,NO2,CO and O3 exposure in heating season,respectively;the results showed that 19.5%(95%Cl(5.6-30.7%or 979 admissions)),22.4%(95%CI(6.1-36%or 1127admissions)),15%(95%Cl(-4.2-29.8%or 754admissions)),8.3%(95%Cl(-20-28.1%or 418admissions)),10.4%(95%Cl(-15.3-30.1%or 522 admissions))and-2.5%(95%Cl(-27.5-18.2%or-125 admissions))of AECOPD hospitalizations could be attributable to PM2.5,PM10,SO2,NO2,O3 and CO exposure in non-heating season,respectively.Summary:1.There were higher PM2.5,PM10,NO2,SO2,and CO concentrations in heating seasons than non-heating season in Shijiazhuang;However,O3 was higher in non-heating season than heating season.2.The cumulative relative risk of NO2 was the greatest in every one unit of air pollutants during the heating season and the cumulative relative risk of SO2 was the greatest during the non-heating season.3.The attributable burden of AECOPD hospitalization in heating season and non-heating season are different.PM2.5,PM10,NO2 and CO are the main factors of heating season,while PM10,PM2.5,SO2 and O3 are the main factors of non-heating season.Conclusions:The main air pollutants causing chronic obstructive pulmonary disease hospitalization in heating and non-heating seasons are different,and when government departments develop interventions to reduce the risk of chronic obstructive pulmonary disease hospitalization,the effects of these factors on the burden of disease should be considered.Part Three Effects and mechanisms of IL37 on fine particulate matter induced extracellular matrix deposition in bronchial epitheliumObjective:The aim of this study was to observe the effect of PM2.5 on extracellular matrix deposition in human bronchial epithelial cells and to further analyze the levels of WNT-5A expression in extracellular matrix deposition.whether IL-37?WNT-5A siRNA can partially inhibit WNT-5A expression,attenuate the effect of PM2.5 on extracellular matrix deposition in bronchial epithelial cells,and explore the relationship between WNT-5A and oxidative stress,inflammatory response signaling pathway.Methods:In the study of cell model,the expression of the WNT-5A was detected by Western Blot by using different concentrations of PM2.5 on human epithelial cells for 24 hours.According to WNT-5A expression,we select the best concentration of PM2.5.16HBE cells were divided into normal group and PM2.5 group.The cells received PM2.5 stimulation for 24 hours,and use Western Blot to detect the expression of fibronectin,Nrf2,WNT-5A.The cellular immune fluorescence method was used to further verify the expression of WNT-5A.The reactive oxides produced and released by mitochondria were detected by fluorescence method.The ELASA method was used to observe whether could up-regulate the expression of IL6,IL-1?.Then,WNT-5A deficiency in HBE cells was achieved using WNT-5A related gene siRNA and IL-37 intervention to observe the expression of the WNT-5A and to observe whether it can attenuate the expression of fibronectin,Nrf2 and inflammatory factor IL6,IL-1? in response to PM2.5.Results:1.WNT-5A expression stimulating 16HBE with different concentrations of PM2.5:The expression of WNT-5A by Western Blot was significantly increased than the control group at different concentrations of PM2.5 stimulation for 24 hours(P<0.05);WNT-5A expression was significantly increased than the control group and other concentration groups at PM2.5(12.5?g/mL)stimulation(P<0.001).2.Fibronectin,Nrf2 expression stimulating 16HBE cell at PM2.5(12.5?g/mL):The expression of fibronectin,Nrf2 by Western Blot were significantly increased than the control group at PM2.5 12.5?g/ml for 24 hours(P<0.001).3.WNT-5A immunofluorescence expression:Much bright green fluorescence in the cytoplasm of HBE cells further confirm that PM2.5 can increase WNT-5A expression after exposure to PM2.5 for 24h,as indicated by inverted microscope analysis.This is consistent with Western Blot.4.Release of inflammatory factors IL6?IL-1?:The release of IL6?IL-1? by ELISA were significantly increased than the control group at different concentrations of PM2.5 stimulation for 24 hours(P<0.001).The IL6?IL-1? release were significantly increased than the control group and other concentration groups at PM2.5(12.5?g/mL)stimulation(P<0.001).5.PM2.5 stimulates reactive oxygen species(ROS)production:The production of reactive oxygen species stimulated by different concentrations of PM2.5 for 24 hours was significantly higher than that of the control group by DCFH-DA fluorescence intensity assay(P<0.01);The ROS production was significantly increased than the control group and other concentration groups at PM2.5(12.5?g/mL)stimulation(P<0.01).6.WNT-5A Immunofluorescence expression after IL37 intervention:PM2.5 stimulation was given after IL37 intervention,and the green fluorescence decreased in IL37 intervention group compared with PM2.5 group.7.The expression of WNT-5A?fibronectin?Nrf2 after IL37 intervention:PM2.5 stimulation was given after IL37 intervention,and the expression of WNT-5A?fibronectin?Nrf2 in IL37 intervention group was decreased compared with that PM2.5 group(P<0.05).8.The release of IL6?IL-1? after IL37 intervention:PM2.5 stimulation was given after IL37 intervention,and the release of IL6 in IL37 intervention group was decreased compared with that PM2.5 group(P<0.01);IL-1? was also decreased(P<0.001).9.The production of ROS after IL37 intervention:PM2.5 stimulation was given after IL37 intervention,and the production of ROS in IL37 intervention group was decreased compared with that PM2.5 group(P<0.001).10.WNT-5A Immunofluorescence expression after WNT-5A siRNA transfection:The green fluorescence of WNT-5A siRNA group decreased compared with Scrambled siRNA group;the green fluorescence decreased in the PM2.5 stimulated WNT-5A siRNA group compared with Scrambled siRNA group.11.The expression of WNT-5A?fibronectin?Nrf2 after transfection WNT-5A siRNA:The expression of WNT-5A?fibronectin?Nrf2 by Western Blot in WNT-5A siRNA group were lower than that Scrambled siRNA group(P<0.001);The expression of fibronectin,Nrf2 in the WNT-5A siRNA group with PM2.5 stimulation were lower than the Scrambled siRNA group(P<0.001).12.The release of IL6?IL-1? after transfection WNT-5A siRNA:The release of IL6?IL-1? in WNT-5A siRNA group were lower than that in Scrambled siRNA group(P<0.001);The release of IL6?IL-1? in the WNT-5A siRNA group with PM2.5 stimulation lower than the Scrambled siRNA group(P<0.001).13.The production of ROS after transfection WNT-5A siRNA:The production of ROS in WNT-5A siRNA group was lower than that in Scrambled siRNA group(P<0.001);the production of ROS in the WNT-5A siRNA group with PM2.5 stimulation lower than the Scrambled siRNA group(P<0.001).Summary:In vitro cell models,PM2.5 can induce elevated WNT-5A expression and aggravate extracellular matrix deposition,possibly through inflammatory,oxidative stress pathway to regulate extracellular matrix.WNT-5A siRNA and IL37 can inhibit the inflammatory response,oxidative stress and down regulate WNT-5A expression,which provides therapeutic targets for the treatment PM2.5 induced extracellular matrix deposition.