A Mendelian Randomization Analysis Of The Association Between Inflammasomes And Coronary Artery Disease Risk

BackgroundCoronary Artery Disease（CAD）,short for Coronary Artery Diseaseor Ischemic Heart Disease,is an organic disease of heart dysfunction caused by coronary artery stenosis and/or insufficient blood supply.CAD has been a worldwide public health problem.In the United States,the total prevalence of CAD among adults≥20 years old is 6.7%,among which the prevalence of CAD is 7.4%in males and 6.2%in females.According to China cardiovascular disease report 2018,the prevalence of cardiovascular disease in China is on the rise,with CAD now exceeding 11 million.CAD is an acute clinical event caused by the instability/rupture of atherosclerotic plaque,and the chronic inflammatory environment may be the direct cause of plaque instability/rupture and thrombosis.The release and accumulation of inflammatory factors can promote the thinning of the fibrous cap of plaque and increase its instability.NLRP3 regulates the body’s chronic inflammatory response and promotes the conversion of proinflammatory precursors such as IL-1βand IL-18 into active forms.Il-18 is a"multipotent"inflammatory factor that plays an important role in the immune and inflammatory response process.It participates in a series of inflammatory responses by inducing the generation of INF-γ,the proliferation and migration of vascular smooth muscle cells,and the apoptosis of vascular endothelial cells.NLRP3 gene rs10754558（C/G）,IL-18 gene 187238（G/C）,and rs1946518（C/A）are all located in the promoter region of the corresponding genes.Mendelian randomization analysis using these SNPs as instrumental variables can avoid confounding factors and anti-reverse effects in traditional observational studies.The interference of causality to the association effect makes it possible to infer the association between the cytokine phenotype associated with the inflammatory body activation pathway and the occurrence of CAD.Objectives1.Using mendelian randomization method,on the basis of the association between genotype-phenotype and genotype-disease outcome,infer the association between the NLRP3 level and the risk of CAD.2.Based on data from systematic reviews and original research,using mendelian randomization method,on the basis of the association between genotype-phenotype and genotype-disease outcome,infer whether IL-18 levels are associated with increased risk of CAD.Methods1.Case-control study design:The study object was from the Department of Cardiology,Zhongda Hospital Affiliated to Southeast University.The patients with CAD confirmed by coronary angiography were taken as the case group,and the non-CAD individuals were taken as the control group.2.Laboratory studies:serum NLRP3,IL-18 levels were determined by ELISA.Genomic DNA was extracted and genotyping method of RFLP-PCR was used to detect the polymorphism of NLRP3 rs10754558（C/G）.3.Mendelian randomization study:NLRP3 gene rs10754558（C/G）,IL-18 gene187238（G/C）,and rs1946518（C/A）as instrumental variables,combining serologically related phenotypes NLRP3 and IL-18 levels,verify the association between genotype-phenotype,genotype-disease outcome,and then infer the causal association between related cytokines（phenotypes）and the occurrence of CAD.4.Data processing and analysis:Epidata was used for data entry,and SPSS22.0 and STATA 14.0 were used for statistical analysis.Means±standard deviation（）are used to represent the measurement data that conforms to the normal distribution.The measurement data is tested by t test,and the count data、Hardy–Weinberg equilibrium test were tested byχ² test.Logistic regression and linear models are used to construct regression models,and the correlation regression coefficientsβ_ZY of SNP（Z）and CAD（Y）,andβ_ZX of SNP（X）and cytokine level（X）are extracted.β_XY=β_ZY/β_ZX.All test results were statistically significant with P<0.05 on both sides.Results1.In the case-control study,321 patients with CAD diagnosed by CAG and 293non-CAD individuals in the control group were included,totaling 614.There were statistically significant differences in the levels of NLRP3,IL-18 between the coronary artery disease group and the control group.The CAD group was higher than the control group（P<0.05）.2.The association between the NLRP3 gene variant rs10754558（C/G）and the risk of CAD and the level of NLRP3 were statistically significant.The regression coefficientsβ_ZY=0.45 andβ_ZX=2.34.The association between NLRP3 level and the risk of CAD was inferred by Mendelian randomization.The correlation regression coefficientβ_XY=β_ZY/β_ZX=0.45/2.34=0.19.Correlation regression coefficientβ_XY=0.03 of NLRP3 level and CAD risk obtained through case-control study design.The association between NLRP3 levels and CAD risk was statistically significant.3.The association between IL-18 rs1946518（C/A）and the risk of CAD was statistically significant,and the association between IL-18 rs187238（G/C）and IL-18 level was statistically significant.But the association between IL-18 rs187238（G/C）and the risk of CAD was not statistically significant,and the association between IL-18 rs1946518（C/A）and the risk of IL-18 level were not statistically significant.Furthermore,the association between the level of IL-18 and the risk of CAD was not statistically significant.Therefore,the available evidence of association was insufficient to prove a causal association between IL-18 levels and CAD risk.Conclusions1.The inflammasome NLRP3 level was associated with the increased risk of CAD in a mendelian randomization study.2.The existing evidence of association was insufficient to prove a causal relationship between IL-18 levels and the risk of CAD.