Influence of NOD-like receptor 2 gene polymorphisms on muramyl dipeptide induced pro-inflammatory response in patients with active pulmonary tuberculosis and household contacts

Abstract

Purpose: The immune response induced by nucleotide-binding oligomerization domain-2(NOD2) is associated with the production of cytokines affected by the host's genetic background. The present study aimed to examine the effects of NOD2; 802C > T, 2105G > A polymorphisms associated with altered cytokine levels in patients with active pulmonary tuberculosis disease, Latent TB subjects (household contacts(HHC) and healthy controls(HC).

Methods: Genetic polymorphisms were analyzed by Restriction Fragment Length Polymorphism(RFLP) in 102-PTB patients, 102-HHC, and 132-HC. QuantiFERON-TB Gold In-Tube test was performed to identify latent TB infection in 60-HHC. Estimated their cytokine levels by ELISA in MDP (muramyl dipeptide) stimulated culture supernatants of all the groups. Further, we studied pre-mRNA structures by insilico analysis and relative gene expression by RT-PCR.

Results: Recessive genetic models of NOD2 802C > T SNP with TT genotype and AA genotype of NOD2 2105G > A SNP were significantly associated with increased TB risk in PTB patients and HHC compared with HC. In vitro stimulations were performed with NOD2 ligand MDP in PTB patients and latent TB subjects: QuantiFERON positive household contacts (QFT + ve HHC)and QuantiFERON negative household contacts(QFT-ve HHC). The results showed that reduced TNF-α and enhanced IL-12, IL-1β indicate that these cytokines may play an essential role in the initial maintenance of cell-mediated immunity. Our study demonstrated the correlation between NOD2 polymorphism with IL-1β, TNF-α, IL-12 levels. Insilico analysis represents the pre-mRNA secondary structures affected by NOD2 SNPs. We also observed the difference in m RNA levels in variant and wild genotypes.

Conclusion: This finding may lead to the forthcoming development of immunotherapy and may be used as predictive markers to identify high-risk individuals for TB disease.

Keywords: Innate immune response; Interleukin-12; Interleukin-1β; Latent TB; MDP stimulation; Mycobacterium tuberculosis; NOD-like receptor2 polymorphisms; Tumor necrosis factor-α.