Effects Of Short-term Smoking Cessation Intervention On DNA Methylation Of COPD Related Genes In Smokers

Object:As a common and preventable environmental risk factor contributes to several diseases,smoking is a major risk factor of chronic obstructive pulmonary disease(COPD).COPD is a chronic inflammatory disease characterized by continuous airflow limitation.A number of studies at home and abroad have shown that COPD is related to abnormal expression of DNA methylation.However,there are few studies on the differential expression of DNA methylation by smoking cessation intervention,and no prospective studies have been reported on the effect of short-term smoking cessation intervention on DNA methylation of genes related to COPD in smokers.In this project,Illumina Infinium MethylationEPIC BeadChip(850K)was used to study the differential expression of DNA methylation of smokers'COPD related genes before and after short-term smoking cessation intervention,so as to provide a theoretical basis for the prevention and treatment of COPD.Methods:1.86 cases of smokers who visited the smoking cessation clinic of Henan Provincial People's Hospital from 2018.5-2019.12 and 16 cases of non-smokers who visited the physical examination center were included.General information of the subjects was collected,including age,gender and weight,and were examined by exhaled carbon monoxide.2.According to the willingness of the subjects,93 subjects were examined for lung function,including 77 smokers and 16 non-smokers.Blood samples were collected from 68 patients,including 52 smokers and 16 non-smokers.3.The smoking cessation status of smokers were followed up for half a year.8 smokers who had not smoked for more than 3 months and had no significant lifestyle changes were selected Blood samples were collected,pulmonary function and exhaled carbon monoxide test were performed on the 8 smokers.4.Serum levels of IL-6,IL-8,IL-10,CRP,IL-17 and SP-D were detected by ELISA.5.Whole blood DNA was isolated from 8 subjects before and after smoking cessation intervention,and the samples were analyzed using Illumina 850K.6.SPSS22.0 software was used for data collation and analysis,and R package Limma was used for statistical analysis of DNA methylation data P<0.05 was statistically significant.Results:1.There was no significant difference in age,gender and weight between the smoking group and the non-smoking group.FEV1(3.24±0.69)and FEV1/FVC%(77.22±8.77)were lower in the smoking group than in the non-smoking group(P<0.05).There was no significant difference in FEV1%pred,FVC and FVC%pred(P>0.05).The cumulative smoking amount(package)of smokers was negatively correlated with FVC,FEV1 and FEV1/FVC%(r=-0.39,P<0.001;r=-0.52,P<0.001;r=-0.29,P<0.05).2.After smoking cessation intervention,subjects'expiratory CO and tobacco dependence scores decreased significantly compared with those before,showing statistical differences(P<0.05,P<0.01).There was no statistical difference in FEV1,FVC,FEV1%pred and FVC%pred of the subjects' lung function after smoking cessation intervention.3.Serum IL-6,IL-8 and IL-17 in the smoking group were higher than that in the non-smoking group,but there was no statistical significance in IL-6 and IL-17(P>0.05).Serum IL-10 and SP-D in the smoking group were lower than those in the non-smoking group,with no statistical difference(P>0.05).4.There was a negative correlation between IL-10 and nicotine dependence(r=-0.272,P=0.025).IL-8 was positively correlated with the degree of nicotine dependence and daily smoking(r=0.411,P=0.001;r=0.301,P=0.013);CRP was negatively correlated with FEV1/FVC%(r=-0.297,P=0.022),and SP-D was positively correlated with FVC%pred(r=0.269,P=0.041).5.Compared with before smoking cessation intervention,about 25,444 genes showed differential expression of methylation sites after smoking cessation intervention(p.value<0.05).6.Before and after smoking cessation intervention,CRP,BCL3,AQP3,SOCS3 and other inflammation-related genes were differentially expressed in DNA methylation sites.7.Short-term smoking cessation intervention can increase or decrease the methylation level of COPD related Notch signaling pathway,PI3K/AKT signaling pathway,Nrf2 pathway,EPAS1,BID,GCLC and other genes towards the direction of improving COPD.Conclusion:Smoking leads to decreased lung function of smokers,which is related to the degree of tobacco dependence and the cumulative smoking amount(bags).Smoking causes the body to be in a state of chronic inflammatory response,which is positively correlated with the degree of tobacco dependence and daily smoking.Short-term smoking cessation interventions may improve the occurrence and development of COPD by altering the level of DNA methylation of COPD related genes.