Understanding the expression of Toll-like receptors in Asian Indians predisposed to coronary artery disease Table I. Primer sequences used for TLR genes and GAPDH2 with PCR conditions Genes TLR2 TLR4 TLR9 GAPDH2 Primers 1642F 1713R 1951F 2020R 1533F 1626R 3210F 3268R Sequence CCAGGCTCTGGTGCTGACAT AGACTGCCCAGGGAAGAAAAA GGCATGCCTGTGCTGAGTT TGAGGACCGACACACCAATGA AGTCAATGGCTCCCAGTTCCT CGTGAATGAGTGCTCGTGGTA CAAGGCTGTGGGCAAGGT GGAAGGCCATGCCAGTGA adjustment for traditional risk factors, namely, age, gender, history of smoking, hypertension and diabetes. The correlation amongst the expression levels of TLR-2, TLR-4 and TLR-9 genes was assessed by multivariate statistics and Pearson’s correlation test. Logistic regression analysis was performed to assess the association of TLRs with CAD after eliminating the effect of age and gender. SPSS version 15 was employed for statistical analysis and a nominal p value of 0.05 or less was considered as statistically significant. Results The clinical characteristics of the study participants are provided in Table II. There was no significant difference between cases and controls with respect to the mean age, gender, systolic and diastolic blood pressure, body mass index, waist- hip-ratio, and smoking. Frequency of diabetes and hypertension was, however, significantly higher in the cases than in the controls (p < 0.001). Of the 60 CAD patients, 21 of them had suffered a myocardial infarction (MI) event, while 30 subjects had stable Table II. Clinical characteristics of study participants PCR condition Initial denaturation – 95°C for 10', 40 cycles of denaturation at 95°C for 15 s, annealing/extension at 60°C for 1', final dissociation step angina disease. Clinical details were missing for 9 individuals who had proven CAD based on a cardiologist’s diagnosis. The mean age at onset of CAD was 50 ±7.4 years for males and 56.7 ±4.2 years for females. Gene expression analysis The expression levels of the TLR-2 gene was found to be significantly elevated in the cases (1.295 ±0.09) as compared to the controls (1.033 ±0.08) (p = 0.015). On the other hand, the expression levels for the TLR-9 gene was significantly lower in the cases (1.522 ±0.18) than the controls (2.165 ±0.16) (p = 0.032) while there was no significant differences in the expression levels for the TLR-4 gene (p = 0.174) (Figure 1). This pattern did not change following covariate adjustment for age, gender, history of smoking, hypertension or diabetes (Figure 2). We observed a significant correlation of TLR-2 with TLR-4 (r = 0.803, p < 0.0001) and TLR-9 (r = 0.264, p = 0.003) as well as between TLR-4 and TLR-9 (r = 0.303, p = 0.001), respectively, after Controls (n = 79) Value of p Parameter Age [years] Gender (M/F) Body mass index [kg/m2] Waist/hip ratio Systolic blood pressure [mmHg] Diastolic blood pressure [mmHg] Height [cm] Weight [kg] Diabetes [%] Hypertension [%] Statin [%] Smoking (ever) Arch Med Sci 5, October / 2011 Cases (n = 60) 52.19 ±7.39 53/7 25.6 ±0.49 0.97 ±0.01 121.34 ±1.77 77.93 ±1.12 166.12 ±1.11 70.44 ±1.2 27 (47.4) 31 (55.4) 30 (50.0) 27 (46.6) 51.76 ±8.27 72/7 24.97 ±0.48 0.96 ±0.01 127.03 ±2.21 81.0 ±1.17 165.52 ±0.83 68.28 ±1.4 5 (6.4) 6 (7.7) 0 31 (39.7) 0.935 0.586 0.369 0.302 0.06 0.069 0.66 0.265 < 0.0001 < 0.0001 – 0.427 783