Caracterização molecular da região promotora do gene BAT1 em pacientes com Leishmaniose Cutânea causada por Leishmania guyanensis no estado do Amazonas

Abstract

Leishmaniasis is a globally neglected infectious-parasitic disease, affecting almost 110,000 individuals in Brazil in the last 5 years. In the region of the municipality of Manaus-AM, most cases of Cutaneous Leishmaniasis (LC) are recorded, with Leishmania guyanensis as the main etiological agent. HLA-B Associated Transcript 1 (BAT1) regulates inflammation by modulating the expression of pro-inflammatory cytokines by playing a protective role in various immunopathological disorders. Thus, the objective of this study was to determine the influence of existing polymorphisms in the promoter region of the BAT1 gene in the clinical setting of patients with LC with a case-control study model. In addition, serum concentrations of the cytokine Tumor Necrosis Factor (TNF)-alpha in each population were correlated with the polymorphisms found in the attempt to identify sub-phenotypes in patients with LC based on the study of molecular markers in the BAT1 gene. For the molecular study, amplifications were performed using the conventional Polymerase Chain Reaction (PCR) technique in the promoter region of the BAT1 gene and were purified by the ExoSAP-IT ™ PCR Product Cleanup Reagent-Thermofisher methodology, with subsequent sequencing by the described termination method by Sanger. Our results were based on a population of 516 individuals divided into 267 cases and 249 controls, of which 79.7% and 68.7%, respectively, were male. The most frequent age group was 19 to 39 years. SNPs -22 (rs2239527) and -348 (rs2239528) were the most frequent in the study population, including for the first time in the literature, SNP-277. The frequencies found for SNPs -22 and -348 were very similar between cases (31.15% / 4.28%) and controls (25.78% / 6.91%), respectively, with no association between these two populations . Serum TNF-alpha levels were higher in cases and gender independent (p <0.001). In addition, significant elevated levels involved the carriers of SNPs -22 and -348 only when they were cases rather than between healthy people. Serum TNF levels were elevated proportionally in the presence of polymorphisms, and almost twice as high in those homozygous cases (52.19 ± 18.43) than controls (32.15 ± 29.74) (p <0.001). Importantly, higher serum TNF levels were associated with SNP -348 (72.89 ± 49.33) in cases than in -22 (41.13 ± 22.16) in the same group. In addition, when concomitantly found in the same sample, the -22 and -348 SNPs further increased the expression of serum TNF levels. Our results reinforce the role of BAT1 from evidence that the presence of polymorphism indirectly affects TNF expression. Thus, we believe that this may influence the outcome and clinical manifestation of the patient with Leishmaniasis, especially in the presentation of cutaneous lesions, since there is a correlation between the diameter of the lesion in the LC and the frequency of production of inflammatory cytokines.