Effects Of ABCA1 Gene Polymorphism And Methylation Modification On Premature Coronary Heart Disease

Objectives: ATP-binding cassette transporter A1(ABCA1)plays an crucial role in anti-atherosclerosis by regulating HDL synthesis,reversing cholesterol transport(RCT)and inflammation inhibition.The purpose of this study was to analyze the relationship between ABCA1 gene polymorphism(R219K,-565 C/T)and ABCA1 promoter methylation level with age,sex,lipid levels,inflammatory mediators and other clinical indicators,and to determine the effects on the risk of p CAD(including premature acute coronary syndrome),and to explore the main factors affecting the methylation level of ABCA1 promoter,to further provide a new direction for the early prediction and clinical diagnosis and treatment of p CAD.Methods: A total of 120 patients with p CAD hospitalized in the Cardiology Department of our center from May 2019 to December 2019 were enrolled.Meanwhile,88 age and gender matched asymptomatic individuals from the health examination center of our hospital were selected as the control group.Basic clinical data of patients were routinely collected.Serum TC,TG,LDL-C,HDL-C,FPG,Hcy,Hb A1 c,ABCA1 and CPR,cf DNA/NETs and IL-1β were measured with fasting venous blood.Then the whole blood DNA was extracted and the ABCA1(R219K,-565C/T)polymorphism were detected by polymerase chain reaction-restriction fragment length polymorphism(PCR-RFLP).At the same time,the patients with premature ACS in p CAD(90 cases)and the control group(88 cases)were classified as subgroup.DNA extracted from subgroup samples were evaluated for ABCA1 DNA promoter methylation using pyrosequencing method.SPSS 20.0 was used for statistical analysis of the above dates.Part I Results: Effect of gene polymorphism on p CAD(1)The BMI,plasma LDL,Hb A1 c,CRP,cf DNA/NETs and IL-1β levels were signifcantly higher in the p CAD group than those in the control group(P<0.05),whereas the HDL level and ABCA1 content were significantly lower(P<0.01);Patients with p CAD were also more likely to have diabetes and family history of p CAD than the control group.(2)Compared with the control group,the ratio of KK+RK genotype and the K allele in p CAD group were significantly decreased,while the frequency of R allele was higher(P<0.05).The distribution frequency of 565C/T genotype has higher CT+TT genotype and T allele in the p CAD group,while the frequency of C allele is lower than that in the control group(P<0.05).(3)Plasma HDL-C level of R219 K RK+KK genotype was higher than that of RR genotype patients,but no similar results were found in the control group.And there were no significant differences in plasma LDL,TC,TG,ABCA1 levels and Inflammatory mediators among different R219 K genotypes in p CAD group,control group and the whole study population(P>0.05).The 565C/T gene polymorphism had no significant effect on the serum lipid levels,Inflammatory mediators and ABCA1content(P>0.05).(4)Logistic regression analysis showed that the risk of p CAD in KK and RK genotypes was 70.7%(OR=0.294,P=0.007)and 62.1%(OR=0.379,P=0.014)lower than that in 219 RR genotypes,respectively.However,ABCA1-565C/T polymorphism had no significant effect on the risk of p CAD(P>0.05).Part II Results: Relationship between ABCA1 promoter methylation and p ACS(1)Compared with the control group,the level of HDL-C in p ACS group was significantly lower(P<0.01),while BMI,Hb A1 c,LDL,waist circumference,IL-1β,cf DNA/NETs,CRP levels and the proportion of patients with family history of p CAD and diabetes were significantly higher(P<0.05).(2)The methylation levels of 8 different sites(Cp G1-8)in the ABCA1 promoter of the p ACS group and their average values were significantly higher than those of the control group(P<0.01),while the serum ABCA1 content was lower than that of the control group(P<0.05).(3)Binary Logistic regression analysis showed that the mean methylation level of ABCA1 promoter,Hb A1 c,LDL,cf DNA/NETs and IL-1β levels were independent risk factors for the p ACS,while HDL was a protective factor for p ACS.(4)Correlation analysis showed that the mean methylation level of ABCA1 DNA promoter was negatively correlated with serum HDL-C level(r=-0.488,P<0.001)and positively correlated with Gensini Score,CRP,cf DNA/NETs and IL-1βlevels(P<0.001).Multiple linear regression analysis showed that plasma CRP,cf DNA,IL-1β,Hcy and Hb A1 c were independent factors affecting the methylation level of ABCA1 promoter after excluding factors such as age,gender,smoking history and so on.Conclusion:1.Regression analysis showed that the risk of p CAD in KK and RK genotypes were significantly reduced,and R219 K gene polymorphism has an effect on plasma HDL-C level,but has no relationship with TG,TC,LDL levels and inflammatory mediators,suggesting that the 219 K allele is a protective factor for p CAD and may play a role by regulating HDL-C levels.2.Increased ABCA1 promoter methylation level was associated with decreased HDL-C level and increased the risk of p ACS independently of lipid levels.Therefore,ABCA1-A promoter methylation level could be used as a potential biomarker to predict the risk of p ACS and assist in the early diagnosis and prevention of p CAD in a non-invasive way.3.Serum inflammatory mediators(CRP,cf DNA/NETs,and IL-1β)were positively correlated with the mean methylation level of ABCA1 promoter,and were independent factors influencing the methylation level of ABCA1 promoter.Therefore,we hypothesized that by regulating the inflammatory response,DNA methylation status could be changed,and the expression of ABCA1 can be adjusted indirectly or directly to intervene early in CAD progression.