| Background Rheumatoid arthritis(RA) is a common autoimmune disease, characterized by T lymphovytes infiltration, systemic inflammation, persistent synovitis and severe impairment of joints, which leads to progressive destruction of cartilage and bone. And systemic lupus erythematosus(SLE) is a complex and systemic autoimmune disease, which affect various organ of the body, characterized by producing a variety of autoantibodies, complement activation and immune complexes deposition, and causing tissue and organ damage. Although the aetiology and pathogenesis of RA and SLE have not been completely demonstrated, the occurrence of the diseases are both believed to result from the combination of multiple factors, and genetic factors have been suggested to play an important role in the development and progression of the diseases. Recent many studies discover many susceptibility genes/loci for RA and SLE. However, the heritability of RA and SLE could not be completely explained by all these disclosed genetic factors. The genetic interaction might be one important explanation for the “missing heritability”.Numerous studies have proved that the T helper cell 17(Th17)–driven autoimmune response and its related inflammatory cytokines play an important role in the pathogenesis of RA and SLE. And many studies found that the genes involved in the regulation of Th17 cells which response variability and strongly associated with RA and SLE risk. These findings highlight Th17 cells response as important contributor to the onset of RA and SLE. The latest evidence suggests dipeptidylpeptidase 4(DPP4),chemokine(C-C motif) receptor 6(CCR6),interleukin 6 receptor(IL6R) and interleukin 23 receptor(IL23R) are susceptibility genes to RA. CD26 and CCR6 were recognized as molecular markers of Th17 cells, IL6 R and IL23 R were important cytokine receptor which involved Th17 signaling pathway.Recent studies have discovered that autoimmune diseases shared genetic background, and maybe have common molecular pathogenesis and biological pathways. Besides, current data indicate that the genetic interaction contribute to RA and SLE susceptibility. And our study have found the epistatic interaction between IL21(interleukin-21) and ETS1(E26 transformation-specific-1) in SLE. Thus our study was to discover the gene-gene interaction among Th17 cells related genes(DPP4, CCR6, IL6 R and IL23R) in RA and SLE.Objective This study was undertaken to examine the association of single nucleotide polymorphisms( DPP4 rs12617656, CCR6 rs1854853, IL6 R rs2228145, IL23 R rs7517847) within genes involved in Th17 signaling pathway with genetic susceptibility to RA and SLE, and further test for gene-gene interaction among these genes in RA and SLE in a Chinese population.Methods Independent case-control studies were conducted in unrelated ethnic Han Chinese in two stages. In the first stage, four DPP4 rs12617656, CCR6 rs1854853, IL6 R rs2228145, IL23 R rs7517847 were genotyped in RA, SLE and healthy controls by using Fluidigm 192.24 Dynamic Array and Taq Man probe genotyping detection technology. To examine the association of single nucleotide polymorphisms(DPP4 rs12617656, CCR6 rs1854853, IL6 R rs2228145, IL23 R rs7517847) with RA and SLE risk, and further test for gene-gene interaction among these genes in the diseases. In the second stage, we further recruited an additional independent cohort to validate significant interaction which was found in the first stage. Moreover, we detected the association of gene combination with the plasma levels of the Th17-related cytokines. All patients with RA and SLE were recruited from the Department of Rheumatology and Immunology of the First Affiliated Hospital of Anhui Medical University and Anhui Provincial Hospital. The diagnosis of RA was established according to the criteria of 1987 American College of Rheumatology(ACR) revised criteria for the classification of RA, and the diagnosis of SLE was based on the criteria of 1997 ACR revised criteria for the classification of SLE. The healthy controls were come from volunteer blood donors and medical center of hospital.Statistical analysis was performed using SPSS13.0 software. Estimated odds ratio(OR) and 95% confidence interval(95%CI) were calculated. Chi-square test and logistic regression analysis were applied to compare allele and genotype frequencies between patients and controls. Logistic regression analysis was used to estimate the multiplicative interaction. The additive interaction was analyzed by 2×2 factorial design, attributable proportion due to interaction(AP) and the relative excess risk due to interaction(RERI). The plasma levels of Th17-related cytokines was detected by enzyme linked immunosorbent assay(ELISA), the statistically difference between different groups was compared using the Kruskal-Wallis H test. A two-sided P value of less than 0.05 was accepted as statistically significant.Results(1) In the first stage, a total of 2452 samples(386 RA, 1058 SLE and 1008 controls) were included. The association of DPP4 rs12617656 with RA susceptibility was replicated in our study(T vs. C: P = 0.004, OR = 1.290, 95%CI: 1.087-1.530). We discovered the association of DPP4 rs12617656 with anti-citrullinated protein antibodies(ACPA)-positive RA(TT vs. TC+CC: P = 0.006, OR = 1.697, 95% CI:1.167-2.467; TT vs. TC vs. CC:P = 0.012, OR = 1.285, 95%CI = 1.056-1.563) and with rheumatoid factor(RF)-positive RA(TT vs. TC+CC: P = 0.011, OR = 1.649, 95%CI = 1.124-2.419; TT vs. TC vs. CC:P = 0.028, OR = 1.253, 95%CI = 1.025-1.532). The associations of IL6 R rs2228145 with RA and RF-positive RA(CCvs. CA+AA: P = 0.049, OR = 1.376, 95%CI = 1.002-1.891; TT vs. TC+CC: P = 0.030, OR = 1.476, 95%CI = 1.038-2.097) were found. We also examined the association between DPP4 rs12617656 and SLE in the Chinese Han population(T vs. C:P = 0.039, OR = 1.144, 95%CI = 1.007-1.299).(2) In the first study, the genetic interactions study in RA, we found a significant multiplicative interaction between DPP4 rs12617656 and CCR6 rs1854853(codominant model, P = 0.014). Additive interaction analysis revealed significant interaction DPP4 rs12617656 and CCR6 rs1854853(AP = 0.425, 95%CI = 0.026-0.824). For the gene-gene interaction between DPP4 rs12617656 and CCR6 rs1854853, we further observed the significant multiplicative interaction(dominant model, P = 0.030, OR = 2.054, 95%CI = 1.071-3.943; codominant model, P = 0.002, OR = 1.555, 95%CI = 1.176-2.054) and significant additive interaction(AP = 0.534, 95%CI = 0.185-0.882) in ACPA-positive RA; the significant multiplicative interaction(codominant model, P = 0.020,OR = 1.405, 95%CI = 1.056-1.870) and additive interaction(AP = 0.459, 95%CI = 0.045-0.874) in RF-positive RA. Moreover, the significant multiplicative interaction between DPP4 rs12617656 and IL6 R rs2228145 was identified in RA(codominant model, P = 0.031), and in RF-positive RA(dominant model, P = 0.034; odominant model, P = 0.010).(3) In the first study, the genetic interactions study in SLE, we discovered the significant multiplicative interaction between DPP4 rs12617656 and IL6 R rs2228145(recessive model, P = 0.003; codominant model, P = 0.002), further additive interaction analysis significant interaction(AP = 0.686, 95%CI = 0.436-0.936) was showed.(4) In the second stage for the additional independent cohort, we confirmed the significant multiplicative interaction(recessive model, P = 0.001) and significant additive interaction(AP = 1.088, 95%CI = 0.859-1.318) between DPP4 rs12617656 and CCR6 rs1854853 in RA. However we did not found any genetic interaction between DPP4 rs12617656 and IL6 R rs2228145 in RA and SLE.(5) In the pooled study for the genetic interactions research, the epistatic effects between DPP4 rs12617656 and CCR6 rs1854853 were validated in patients with RA, ACPA-positive RA and RF-positive RA. The genetic interactions between DPP4 rs12617656 and IL6 R rs2228145 were consistent in patients with RF- positive RA and SLE.(6) For the results of ELISA, the association of genes combination with the plasma level of the Th17-related cytokines(IL-17, IL-21, IL-22) or the disease activity score in patients with RA and SLE was not detected.Conclusions Our study identified novel genetic interaction between DPP4 rs12617656 and CCR6 rs1854853 in RA. Moreover, we also discovered epistatic interaction between DPP4 rs12617656 and IL6 R rs2228145 in RA and SLE. The results indicated that genetic interaction may jointly affect the susceptibility and progress of the disease. These findings highlight Th17 cells response as important contributor to the pathogenesis of RA and SLE. |
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