Study of polymorphisms in the genes for TNF-α in pediatric patients in the ICU of Instituto Fernandes Figueira – Fundação Oswaldo Cruz

Abstract

Recent advances in molecular biology and genomics have led to increased understanding of the pathophysiological process directly relevant for pediatric intensive care, such as sepsis, ARDS and multiple system organ failure. Several point mutations, called single nucleotide polymorphisms (SNPs), have been identified. The gene of the inflammatory cytokine TNF-α has been indicated as an important candidate for such studies since it has an important function on immuno-inflammatory response, and important SNPs have already been identified. Among these, those that can be detached are those located at positions -863 (C→A) and -308 (G→A) in its promoter region, which can be related directly with the expression of this cytokine and, consequently, with the regulation of the circulating levels of the protein. The aim of this study was to evaluate the polymorphism of the TNF-α gene in positions -863 and -308 in the group of pediatric patients with sepsis/SIRS in the ICU of the Fernandes Figueira Institute – FIOCRUZ. The DNA was extracted from samples of whole blood and with swabs containing oral mucosal cells (in patients that had received blood transfusion) with a mixture of detergents. The molecular determination of the genotypes was carried out using the techniques of PCR-restriction fragment length polymorphism for position -308 and PCR-amplification refractory mutational system for position -863. Eighty-five patients of both sexes and with age varying between 0 and 12 years, with sepsis/SIRS, were admitted. The genotypic frequencies were GG (0.8) and GA (0.2) for -308, and CC (0.77), CA (0.22) and AA (0.01) for -863. The allelic frequency for -308 was G (0.9) and A (0.1), while for -863 it was C (0.88) and A (0.12). Through the analysis of the possible genotypic combinations, it was observed that the more frequent haplotype was CG (0.78) and, using analyses of maximum likelihood, it was verified that the locus did not meet in linkage disequilibrium. The population in this study was in Hardy-Weinberg equilibrium for both the studied polymorphisms. The present study evaluated the genetic characterization of locus TNF-α in this population of pediatric patients with sepsis/SIRS of the metropolitan region of Rio de Janeiro, being comparable with population data of studies in other countries. However, other studies using controls groups should be performed to verify the utility of these polymorphisms as molecular markers for sepsis severity or susceptibility.