Evaluation of the TLR 4 and TLR 5 genes methylation and mRNA expression in patients with Inflammatory Bowel Disease

Abstract

Background Immune system dysfunction, especially Toll-Like Receptors (TLRs), is a major factor in the pathogenesis of IBD. The functional roles of TLR4 and TLR5 as mediators of innate immune responses to pathogenic microbes in the gastrointestinal tract and their association with epigenetic alterations in the pathogenesis of this disease are very important. We aimed to investigate the aberrant methylation changes in TLR4 and TLR5 gene promoter DNA sequences and evaluate the association of TLR4 and TLR5 gene expression with IBD patient subtypes. Methods This case-control study was performed on the tissues of 33 patients with inflammatory bowel disease and 20 healthy controls. After RNA extraction and cDNA synthesis, the expression of TLR4 and TLR5 was examined using qRT-PCR. In addition, following DNA extraction and treatment with sodium bisulfite, the variation in the methylation rate of TLR4 and TLR5 gene promoters was evaluated using High Resolution Melting (HRM) method. Results According to the findings, TLR4 expression increased in the Ulcerative colitis patients compared to the control group (P = 0.0385) but the expression of TLR5 gene in the patients with Ulcerative colitis (UC) and Crohn disease (CD) did not change compared to the control group (P = 0.2710). In addition, no significant correlation was found between the expression of TLR4 and TLR5 and sex. In addition, a significant different was observed in the TLR4 promoter methylation pattern between, UC and normal tissues (P = 0.028), also, a significant different was observed in the TLR4 promoter methylation between, CD and normal tissues (P = 0.006), but in TLR5 this difference was not significant. Conclusion These data suggest that the increased expression of TLR4 and aberrant methylation changes indicate the importance of this gene in the pathogenesis of IBD and its role as a biomarker in IBD patients.