Human leukocyte antigen (HLA)-F and -G gene polymorphisms and haplotypes are associated with malaria susceptibility in the Beninese Toffin children.

BACKGROUND: Little attention has been devoted to the role of the immunoregulatory HLA-E/-F/-G genes in malaria. We evaluated the entire HLA-E/-F/-G variability in Beninese children highly exposed to Plasmodium falciparum (P.f.) malaria. METHODS: 154 unrelated children were followed-up for six months and evaluated for the presence and number of malaria episodes. HLA-E/-F/-G genes were genotyped using massively parallel sequencing. Anti P.f. antibodies were evaluated using ELISA. RESULTS: Children carrying the G allele at HLA-F (-1499,rs183540921) showed increased P.f. asymptomatic/symptomatic ratio, suggesting that these children experienced more asymptomatic P.f. episodes than symptomatic one. Children carrying HLA-G-UTR-03 haplotype exhibited increased risk for symptomatic P.f. episodes and showed lower IgG2 response against P.f. GLURP-R2 when compared to the non-carriers. No associations were observed for the HLA-E gene. CONCLUSION: HLA-F associations may be related to the differential expression profiles of the encoded immunomodulatory molecules, and the regulatory sites at the HLA-G 3'UTR may be associated to posttranscriptional regulation of HLA-G and to host humoral response against P.f.

Hla-C genetic diversity and evolutionary insights in two samples from Brazil and Benin.

Human leukocyte antigen-C (HLA-C) is a classical HLA class I molecule that binds and presents peptides to cytotoxic T lymphocytes in the cell surface. HLA-C has a dual function because it also interacts with Killer-cell immunoglobulin-like receptors (KIR) receptors expressed in natural killer and T cells, modulating their activity. The structure and diversity of the HLA-C regulatory regions, as well as the relationship among variants along the HLA-C locus, are poorly addressed, and few population-based studies explored the HLA-C variability in the entire gene in different population samples. Here we present a molecular and bioinformatics method to evaluate the entire HLA-C diversity, including regulatory sequences. Then, we applied this method to survey the HLA-C diversity in two population samples with different demographic histories, one highly admixed from Brazil with major European contribution, and one from Benin with major African contribution. The HLA-C promoter and 3'UTR were very polymorphic with the presence of few, but highly divergent haplotypes. These segments also present conserved sequences that are shared among different primate species. Nucleotide diversity was higher in other segments rather than exons 2 and 3, particularly around exon 5 and the second half of the 3'UTR region. We detected evidence of balancing selection on the entire HLA-C locus and positive selection in the HLA-C leader peptide, for both populations. HLA-C motifs previously associated with KIR interaction and expression regulation are similar between both populations. Each allele group is associated with specific regulatory sequences, reflecting the high linkage disequilibrium along the entire HLA-C locus in both populations.

Comprehensive analysis of HFE gene in hereditary hemochromatosis and in diseases associated with acquired iron overload.

BACKGROUND: Patients with hepatitis C virus (HCV) and hepatocellular carcinoma (HCC) may or not develop iron overload (IO), which is associated with worst prognosis, because can cause serious damage to organs. HFE gene controls the iron uptake from gut, particularly in patients with hereditary hemochromatosis (HH). AIM: To identify associations between HFE coding region in patients exhibiting hereditary hemochromatosis and in diseases associated with acquired IO. METHODS: We sequenced exons 2 to 5 and boundary introns of HFE gene, evaluating all polymorphic sites in patients presenting hereditary (hemochromatosis) or acquired iron overload HCV and HCC) and in healthy controls, using Sanger sequencing. We also determined the ensemble of extended haplotype in healthy control individuals, including several major histocompatibility complex loci, using sequence specific probes. Haplotype reconstruction was performed using the Arlequin and Phase softwares, and linkage disequilibrium (LD) between histocompatibility loci and HFE gene was performed using the Haploview software. RESULTS: The HFE*003 allele was overrepresented (f = 71%) and HFE*001 allele was underrepresented (f = 14%) in HH patients compared to all groups. A strong linkage disequilibrium was observed among the H63D-G, IVS2(+4)-C and C282Y-G gene variants, particularly in HH; however, the mutation IVS2(+4)T>C was not directly associated with HH susceptibility. The HFE*001/HFE*002 genotype conferred susceptibility to HCC in HCV patients exhibiting IO (P = 0.02, OR = 14.14). Although HFE is telomeric to other histocompatibility genes, the H63D-G/IVS2(+4)-C (P ≤ 0.00001/P ≤ 0.0057) combination was in LD with HLA-B*44 allele group in healthy controls. No LD was observed between HFE alleles and other major histocompatibility loci. CONCLUSION: A differential HFE association was observed for HH and for diseases associated with acquired IO (HCV, HCC). Since HFE is very distant from other histocompatibility loci, only weak associations were observed with these alleles.

Post-transcriptional markers associated with clinical complications in Type 1 and Type 2 diabetes mellitus.

The delayed diagnosis and the inadequate treatment of diabetes increase the risk of chronic complications. The study of regulatory molecules such as miRNAs can provide expression profiles of diabetes and diabetes complications. We evaluated the mononuclear cell miRNA profiles of 63 Type 1 and Type 2 diabetes patients presenting or not microvascular complications, and 40 healthy controls, using massive parallel sequencing. Gene targets, enriched pathways, dendograms and miRNA-mRNA networks were performed for the differentially expressed miRNAs. Six more relevant miRNAs were validated by RT-qPCR and data mining analysis. MiRNAs associated with specific complications included: i) neuropathy (miR-873-5p, miR-125a-5p, miR-145-3p and miR-99b-5p); ii) nephropathy (miR-1249-3p, miR-193a-5p, miR-409-5p, miR-1271-5p, miR-501-3p, miR-148b-3p and miR-9-5p); and iii) retinopathy (miR-143-3p, miR-1271-5p, miR-409-5p and miR-199a-5p). These miRNAs mainly targeted gene families and specific genes associated with advanced glycation end products and their receptors. Sets of miRNAs were also defined as potential targets for diabetes/diabetes complication pathogenesis.

MicroRNA expression profiles discriminate childhood T- from B-acute lymphoblastic leukemia.

MicroRNAs (miRNAs) play a critical role on biological and cellular processes; the search for functional markers may be of importance for differential diagnosis, prognosis, and development of new therapeutic regimens. In this context, we evaluated the bone marrow miRNA profile of Brazilian children exhibiting T- or B-cell acute lymphoblastic leukemia (T-ALL or B-ALL), using massive parallel sequencing, using the HiSeq 2500 platform (Illumina). The differential expression analysis was conducted considering a leave-one-out approach and FDR ≤ 0.05. Machine learning algorithms were applied to search for the disease subset biomarkers. Target prediction, functional enrichment, and classification of biological categories were also performed. Sixteen miRNAs were differentially expressed between T- and B-ALL, of which 10 (miR-708-5p, miR-497-5p, miR-151a-5p, miR-151b, miR-371b-5p, miR-455-5p, miR-195-5p, miR-1266-5p, miR-574-5p, and miR-425-5p) were downregulated and six (miR-450b-5p, miR-450a-5p, miR-542-5p, miR-424-5p, miR-629-5p, and miR-29c-5p) were upregulated in childhood T-ALL. These miRNAs may be used for distinguishing childhood lymphoblastic leukemia subtypes, since it provided the clear separation of patients in these two distinct groups. Six relevant biological pathways were identified according to their role in leukemia, namely, viral carcinogenesis, cell cycle, and B-cell receptor signaling pathways for induced miRNAs and TGF-beta signaling, apoptosis, and NF-kappa B signaling for the repressed miRNAs, of which several miRNA gene targets participate in cell differentiation and hematopoiesis processes. Machine learning analysis pointed out miR-29c-5p expression as the best discriminator between childhood T- and B-ALL, which is involved in calcium signaling, critical for B-cell lymphocyte fate. Further studies are needed to assure the role of the 16 miRNAs and miR-29c-5p on acute lymphoblastic leukemia subtypes and on disease prognosis.

HLA-G, -E and -F regulatory and coding region variability and haplotypes in the Beninese Toffin population sample.

HLA-G/E/F genes exhibit immunomodulatory properties and are expressed in placenta. Little attention has been devoted to the study of these genes in sub-Saharan African populations, which are yet the most diverse. To fill this gap, we evaluated the complete gene variability, approximately 5.1 kb for HLA-G (n = 149), 7.7 kb for HLA-E (n = 150) and 6.2 kb for HLA-F (n = 152) in the remote Beninese Toffin population, using massive parallel sequencing. Overall, 96, 37 and 68 variable sites were detected along the entire HLA-G, -E and -F, respectively, arranged into region-specific haplotypes; i.e., promoter haplotypes (16, 19, and 15 respectively), coding haplotypes (19, 15, and 29 respectively), 3' untranslated region (3'UTR) haplotypes (12, 7 and 2, respectively) and extended haplotypes (33, 31 and 32 respectively). All promoter/coding/3'UTR haplotypes followed the patterns already described in worldwide populations. HLA-E was the most conserved, exhibiting mainly two full-length encoded-molecules (E*01:01 and E*01:03), followed by HLA-F, three full-length proteins (F*01:01, F*01:02 and F*01:03) and HLA-G, four proteins: three full-length (G*01:01, G*01:03 and G*01:04) and one truncated (G*01:05N). Although HLA-G/E/F alleles in the Toffin population were the most frequently observed worldwide, the frequencies of the coding haplotypes were closely similar to those described for other African populations (Guinea-Conakry and Burkina-Faso), when compared to non-African ones (Brazilian), indicating that variable sites along these genes were present in Africa before human dispersion.

Extended HLA-G genetic diversity and ancestry composition in a Brazilian admixed population sample: Implications for HLA-G transcriptional control and for case-control association studies.

Human leukocyte antigen-G (HLA-G) is a nonclassical Major Histocompatibility Complex (MHC) molecule with immunomodulatory function and restricted tissue expression. The genetic diversity of HLA-G has been extensively studied in several populations, however, the segment located upstream -1406 has not yet been evaluated. We characterized the nucleotide variation and haplotype structure of an extended distal region (-2635), all exons and the 3'UTR segment of HLA-G by next-generation sequencing (NGS) in a sample of 335 Brazilian individuals. We detected 29 variants at the HLA-G distal promoter region, arranged into 19 haplotypes, among which we identified sites that may influence transcription factor targeting. Although the variation pattern in the distal region resembled the one observed in the conventional promoter segment, molecular signature for balancing selection was observed in the promoter segment from -1406 to -1 (Tajima's D = 2.315, P = 0.017), but not in this distal segment (D = 1.049, P = 0.118). Furthermore, the ancestry composition of this Brazilian population sample was determined by the analysis of SNPforID 34-plex ancestry informative marker (AIM) SNP panel. The distribution of HLA-G haplotypes was ancestry-dependent, corroborating previous findings and emphasizing the importance of considering the ancestry information in association studies.