Avaliação funcional de polimorfismos nos genes EBI3 e IL12A associados ao desenvolvimento de asma em uma coorte de crianças no município de Salvador-Ba

Abstract

Asthma is a chronic inflammatory disease of the airways, non-transmissible, influenced by environmental and genetic factors. Allergic asthma is the most common phenotype of the disease and is associated with an exacerbation of the Th2 immune response. Several cell types are involved in the regulation of the immune system in the lung, with emphasis on regulatory cells Treg and Breg. Several inhibitory controls included by these cells have already been admitted, among which are the release of suppressive cytokines such as IL-10, TGF-β and IL-35. IL-35 is a heterodimeric cytokine, composed of the EBI3 and IL12p35 subunits, which are encoded by the EBI3 and IL12A genes, respectively. Changes in the levels of this cytokine have been associated with asthma and atopy. The consternation of the genetic component in the development of asthma is widely reported in the literature. In this context, given the importance of Treg cells and genetic susceptibility, this study set out to investigate the functional impact of polymorphisms in the regulatory genes EBI3 and IL12A in a population of Brazilian children. DNA from 1.218 children was genotyped using the Illumina 2.5 Human Chip Omni Bead. Logistic regression analyzes were performed using PLINK 1.9 software to verify the association between polymorphisms in EBI3 and IL12A, asthma and atopic markers, adjusted for sex, age, helminth survivors and ancestry markers. mRNA expression was performed using real-time qPCR. A total of 4 markers for IL12A and 5 for EBI3 were found. The surprising results that the C allele of rs2243131 in IL12A was positively associated with asthma (OR 1.35, CI 1.06–1.71), asthma severity (OR 1.36, CI 1.02–1.81), positive skin test for Blatella germanica (OR 1.59, CI 1.09–2.22), and also positively associated with the spontaneous production of IL-5 (OR: 1.71; CI: 1.11–2.62). The A allele of rs568408 in IL12A was also positively associated with a positive skin test for B. germanica (OR 1.65, CI 1.10-2, 37). rs582537 in IL12A was associated with a positive skin test for B. germanica (OR 0.64, CI 0.42-0.98) and Dermatophaoides pteronyssinus (OR 0.77, CI 0.60-0.98), in addition to being associated with the natural production of INF-y (OR : 0.52; CI: 0.52–0.99). With regard to EBI3, all the variants found were incorporated into atopy markers: the C allele of rs78749916 (OR 0.61, CI 0.40-0.93), the G allele of rs77145509 (OR 0.66, CI 0.46-0.94), and the A allele of rs76353132 (OR 0.68, CI 0.43-0.96), were associated with a positive skin test for Periplaneta americana. The rs76353132 variant was also associated with spontaneous INF-y production (OR: 0.72; CI: 0.52-0.99). The rs428253 C allele was associated with a positive skin test for at least one allergen (OR: 0.64; CI: 0.44–0.92), and the rs4905 G allele was associated with a positive IgE for at least one allergen (OR 0.62, CI 0 .40-0.95). IL12A mRNA expression levels were reduced in atopic asthmatic subjects when compared to controls. EBI3 mRNA expression levels were decreased in atopic asthmatic subjects compared to non-asthmatic and atopic subjects, and when compared to controls. In this study, we were able, for the first time, to describe new variants in the IL-35 regulatory pathway linked to asthma and atopy, highlighting the importance of immune regulation in the pathogenesis of asthma.