Phlegm Syndrome Of Metabolic Syndrome And CELSR2,PPARG Study On The Relationship Of Gene Promoter Region Methylation

Objective: The DNA methylation levels at Cp G sites of CELSR2 and PPARG genes in peripheral blood of patients with phlegm syndrome and non-phlegm syndrome and healthy people were detected by pyrosequencing method.Meanwhile,the gene modification effects of environmental factors such as smoking,drinking,staying up late and physical activity were integrated to more comprehensively and accurately identify the epigenetic mechanism of MS phlegm syndrome formation.To elucidate the etiology and pathogenesis of phlegm syndrome of MS,and to provide the basis for the early diagnosis and treatment of the disease with traditional Chinese medicine.Methods: 1.Collect MS patients and healthy people meeting the diagnostic criteria,and collect their basic data and four diagnosis information.Using syndrome differentiation method,27 patients with phlegm syndrome and 27 patients with non-phlegm syndrome were screened,and 27 healthy people were selected as controls.Blood samples were collected and biochemical indexes were detected.2.DNA methylation levels at Cp G sites of CELSR2 and PPARG genes were detected to analyze the differences in gene methylation levels among all groups.3.Pearson correlation analysis and Spearman correlation analysis were used to study the correlation of phlegm syndrome element score,physicochemical index and differential Cp G site methylation level in each group.4.Chi-square test was used to analyze the environmental factors related to MS phlegm syndrome,and binary logistic regression model was used to evaluate the association between the DNA methylation level of Cp G sites and the risk of MS phlegm syndrome.Results:1.General data of the healthy group and MS group with phlegm syndrome and non-phlegm syndrome(1)There was no significant difference in gender and age composition among all groups(P>0.05).2.Comparison of clinical physicochemical indexes between healthy group and MS group with phlegm syndrome and non-phlegm syndrome(1)Compared with healthy group,SBP,DBP,BMI,WC,ALT,GGT,FPG and TG in MS non-phlegm syndrome group and MS phlegm syndrome group were increased,and the differences were statistically significant(P<0.05,P<0.01);AST/ALT and HDL-C decreased,and the difference was statistically significant(P<0.01).AST and ALP increased in MS phlegm syndrome group,and the difference was statistically significant(P<0.01).(2)Compared with the MS non-phlegm syndrome group,DBP,BMI,WC,GGT,FPG and TG in the MS phlegm syndrome group were increased,and the differences were statistically significant(P<0.05).3.Comparison of DNA methylation levels of CELSR2 and PPARG genes between healthy group and MS group with phlegm syndrome and non-phlegm syndrome(1)There were no significant differences in DNA methylation levels of CELSR2 and PPARG genes among all groups(P>0.05).(2)Compared with the healthy group,the DNA methylation levels of CELSR2 gene Cp G28 and Cp G34 in the MS non-phlegm syndrome group were decreased,and the difference was statistically significant(P<0.05,P<0.01);The methylation levels of CELSR2 gene Cp G24 and Cp G34 in MS phlegm syndrome group were decreased,and the difference was statistically significant(P<0.05,P<0.01).(3)Compared with the non-phlegm syndrome group,DNA methylation levels of CELSR2 gene Cp G20,Cp G24 and Cp G34 in the MS phlegm syndrome group were decreased,and the difference was statistically significant(P<0.01).(4)Compared with the healthy group,the DNA methylation levels of PPARG gene Cp G19 and Cp G24 in the MS non-phlegm syndrome group were decreased,and the difference was statistically significant(P<0.05);The methylation level of PPARG gene Cp G19 in MS phlegm syndrome group was decreased,and the difference was statistically significant(P<0.05).(5)Compared with the non-phlegm syndrome group,the methylation level of PPARG gene Cp G10 DNA in the MS phlegm syndrome group was decreased,and the difference was statistically significant(P<0.01).4.Correlation between phlegm syndrome element score,clinical physicochemical indexes and CELSR2 gene Cp G34 DNA methylation level in healthy group,MS phlegm syndrome group and non-phlegm syndrome group(1)The phlegm syndrome element score in MS phlegm syndrome group was positively correlated with BMI,WC and TG(P<0.05,P<0.01);The phlegm syndrome element score in MS non-phlegm syndrome group was positively correlated with DBP,BMI and WC(P<0.05),was negatively correlated with AST(P<0.05);Sputum syndrome element score in healthy group was positively correlated with DBP,WC and TG(P<0.05,P<0.01).(2)The methylation level of Cp G34 DNA of CELSR2 gene was positively correlated with HDL-C in MS non-phlegm group(P<0.05).(3)There was no statistically significant correlation between phlegm syndrome score and CELSR2 Cp G34 DNA methylation level in each group(P>0.05).5.Comparison of environmental influencing factors between healthy group and MS group(phlegm syndrome,non-phlegm syndrome)(1)There were statistically significant differences in smoking and drinking between the healthy group and the MS group(P<0.01);There was no statistical significance in staying up late and physical activity(P>0.05).(2)There was statistically significant difference in alcohol consumption between the MS non-phlegm syndrome group and the MS phlegm syndrome group(P<0.05);There was no significant difference in smoking,staying up late and physical activity(P>0.05).6.Correlation between methylation levels at different Cp G sites and environmental risk factors and phlegm syndrome of MS(1)Compared with the normal methylation rate of CELSR2 gene Cp G34,the risk of phlegm syndrome with reduced methylation rate of CELSR2 gene Cp G34 increased to 4.14times(P<0.05);(2)Compared with no alcohol consumption,the risk of MS phlegm syndrome was 3.55 times higher with alcohol consumption(P<0.05).Conclusions:1.The decreased DNA methylation level at Cp G34 site of CELSR2 gene may be the epigenetic basis for the formation of phlegm syndrome in MS.Reduced DNA methylation at Cp G19 of PPARG gene may be the epigenetic basis of MS formation.2.Smoking and drinking are risk factors for the formation of MS,and drinking is risk factors for the formation of phlegm syndrome of MS;The formation of phlegm syndrome of MS may be caused by the combination of decreased methylation level of Cp G34 gene in CELSR2 and alcohol consumption.3.The increase of DBP,BMI,WC,GGT,FPG and TG may increase the risk of phlegm syndrome formation in MS;The formation of phlegm syndrome in MS may be positively correlated with BMI,WC and TG.