Association Between Classical Risk Factors, Long Non-coding RNA Gene Polymorphisms And Coronary Artery Disease Risk

Objective:1.Through the comparison between positive coronary angiography (CAG) cases and negative controls, we analyze the association between risk factors and CAD.2.To evaluate the relationship between ANRIL rs2069416 A>T, ANRIL rs2383206 A>G, ANRIL rs10757274 A>G, ANRIL rs4977574 A>G, H19 rs2839698 OT, DOCK9-AS2 rs913432 A>G polymorphisms and CAD. Haplotype analysis on the association between SNPs in ANRIL gene and the risk of CAD.3. To investigate the gene-gene interaction of SNPs in lncRNA gene on the risk of CAD.Methods:1. Study on the clinical risk factors:Samples were collected from the cardiology department of TEDA International Cardiovascular Hospital.895 patients with coronary artery disease (CAD) and 594 controls with negative coronary angiography (CAG) were interviewed. Samples were conducted an epidemiological survey and collected information on physical examination, medical history and biochemistry test.2. Genetic association studies:Samples were collected from the cardiology department of TEDA International Cardiovascular Hospital. A total of 811 persons, containing 611 CAG-positive cases and 200 CAG negative control, were recruited. SNPs genotyping were performed utilizing a custom-by-design 48-Plex SNPscanTM Kit.3. Data processing and analysis:Establish Epidata database based on the characteristics of data and data were analyzed by SPSS19.0 and STATA12.0 software. Statistical methods include frequency distribution and nonparametric test, t-test, Chi-square test and multivariate Logistic regression.Results:1. A total of 895 patients with coronary artery disease (CAD) and 594 control people with negative CAG were enrolled into our study. There were significant differences in demographic data such as gender, age, hypertension, diabetes mellitus, smoking, drinking, history of CAD and myocardial infarction (MI), and clinical biochemical indicators such as uric acid (UA), creatinine (Cre), fibrinogen (FIB), fasting blood-glucose (FBG), total cholesterol (TC), Triglycerides (TG), High-density lipoprotenin associated cholesterol (HDL-C), Low-density lipoprotenin associated cholesterol (LDL-C) between patients group and control group (P< 0.05). In our study, people with high levels of UA and Cre have 45% and 74% increased risk of CAD (Crude OR=1.45,95%CI:1.17-1.78; Crude OR=I.74, 95%CI:1.41-2.15) respectively, however, after adjustment by age and gender, no significant association was observed. High levels of FBG and FIB both can increase the risk of CAD by 110%(crude OR=2.10,95%CI:1.68-2.61) and 52%(crude OR=1.52,95% CI:1.23-1.88) respectively, while after adjustment by age and gender,127% (Adjusted OR=2.27,95%CI: 1.80-2.85) and 53% (Adjusted OR=1.53,95% CI:1.22-1.92) increased risk were found respectively. High levels of TC and LDL-C both can decrease the risk of CAD by 53% (crude OR=0.47,95%CI:0.38-0.59) and 54% (crude OR=0.46,95%CI:0.37-0.57) respectively, while after adjustment by age and gender,48% (Adjusted OR=0.52,95%CI:0.42-0.65) and 51% (Adjusted OR=0.49,95%CI:0.40-0.62) increased risk were observed respectively. After adjustment by age and gender, high levels of TG can increase the risk of CAD by 45% (Adjusted OR=1.45,95%CI:1.16-1.80), and low levels of TG can increase the risk of CAD by 31% (Adjusted OR=1.31,95%CI:1.05-1.63). Multivariate analysis suggests that high levels of TG, FBG and FIB, low levels of LDL-C, hypertension, diabetes mellitus, smoking, history of CAD and (MI), age≥60y and male are the risk factors of CAD (P<0.05).2. There were significant association between ANRIL rs10757274 A/G polymorphism and CAD (GG vs AA:OR=1.67,95%CI=1.00-2.80; AG+GG vs AA:OR=1.53,95%CI=1.05-2.23; G vs A:OR=1.29,95%C 1= 1.01-1.65). When stratified by age and gender, male subjects carrying AG, GG and AG+GG genotype had 101%,163% and 114% increased risk of CAD (OR=2.01,95%CI:1.14-3.53; OR=2.63,95%CI:1.16-5.96; OR=2.14,95%CI:1.25-3.67) respectively, compare those with the AA genotype. Older subjects (>60 years old) carrying AG and AG+GG genotype had 95% and 88% increased risk of CAD than those with the AA genotype (OR=1.95,95%CI:1.14-3.33; OR=1.88,95%CI:1.14-3.10) respectively. After adjustment by age, gender, hypertension, diabetes mellitus and smoking, significant association was also observed. Under the homozygous model, we have an 84.39% power to detect an OR of 1.87 (a= 0.05). There were also significant association between ANRIL rs4977574 A/G polymorphism and CAD (AG vs AA:OR=1.59,95%CI=1.03-2.45; GG vs AA:OR=1.94,95%CI=1.10-3.42; AG+GG vs AA:OR=1.67,95%CI=1.11-2.53; G vs A: OR=1.37,95%CI=1.05-1.80). When stratified by age and gender, compare with the AA genotype, male subjects carrying AG, GG and AG+GG genotype had 101%,191% and 121% increased risk of CAD (OR=2.01,95%CI:1.09-3.72; OR=2.91,95%CI:1.21-6.96; OR=2.21, 95%CI:1.23-3.95) respectively. Younger subjects (≤60 years old) carrying AG+GG genotype had a 113% increased risk of CAD than those with the AA genotype (OR=2.13,95%CI: 1.07-4.28), younger subjects (< 60 years old) carrying GG genotype had 320% and 183% increased risk of CAD than those with the AA and AA+AG genotype (OR=4.20,95%CI: 1.55-11.34; OR-2.83,95%CI:1.21-6.64) respectively. Older subjects (>60 years old) carrying AG and AG+GG genotype had 94% and 79% increased risk of CAD than those with the AA genotype (OR=1.94,95%CI:1.07-3.50; OR=1.79,95%CI:1.03-3.12) respectively. After adjustment by age, gender, hypertension, diabetes mellitus and smoking, significant association was also observed. Under the homozygous model, we have a 92.57% power to detect an OR of 2.18 (α= 0.05). However, no association between the other four SNPs and CAD risk was found.3. GMDR model showed that there was no statistical significance in two-locus, three-locus, four-locus and five-locus models (P>0.05).4. Results of haplotype analysis indicated that there was no association between haplotypes and CAD (P>0.05).Conclusion:1. Gender, age, hypertension, diabetes mellitus, smoking, drinking, history of CAD and MI, UA, Cre, FIB, FBG, TC, TG, HDL-C and LDL-C are the risk factors for CAD.2. There were significant association between ANRIL rs10757274 A/G and ANRIL rs4977574 A/G polymorphisms and CAD risk.3. Male subjects earring rs10757274 A/G and rs4977574 A/G polymorphisms have increased risk for CAD. Older subjects (>60 years old) carring rs 10757274 A/G polymorphism have increased risk for CAD.4. There were no association between the ANRIL rs2069416 A/T, ANRIL rs2383206 A/G, H19 rs2839698 C/T and DOCK9-AS2 rs913432 A/G polymorphisms and CAD.