RESUMO
Sickle cell disease (SCD) is a hereditary hemoglobinopathy, caused by a mutation at position 6 of the ß-globin chain and patients are frequently exposed to several blood transfusions in order to maintain physiological function. Transfusion blood bags are composed of PVC and phthalates (as DEHP) are often introduced to the material in order to confer malleability. In this sense, DEHP can easily elute to the blood and cause harmful effects. This study aimed to unravel DEHP effect on SCD patient's hemoglobin function. We found that HbS polymerization using whole erythrocytes is decreased by DEHP in ex vivo experiments and this effect might be mediated by the DEHP-VAL6 interaction, evaluated in silico. Isolated HbS exhibited less polymerization at low DEHP concentrations and increased polymerization rate at higher concentration. When analyzing the propensity to aggregate, HbS is more inclined to aggregate when compared to HbA due to the residue 6 mutation. Circular dichroism showed characteristic hemoglobin peaks for oxygenated HbS that are lost when oxygen is sequestered, and DEHP at higher concentration mildly recovers a peak close to the second hemoglobin one. Finally, by transmission electron microscopy we demonstrated that high DEHP concentration increased polymer formation with a more organized structure. These findings show for the first-time the beneficial effect of low-dose DEHP on HbS polymerization.
Assuntos
Anemia Falciforme , Dietilexilftalato , Eritrócitos , Hemoglobina Falciforme , Polimerização , Humanos , Anemia Falciforme/genética , Anemia Falciforme/metabolismo , Hemoglobina Falciforme/genética , Hemoglobina Falciforme/metabolismo , Eritrócitos/efeitos dos fármacos , Eritrócitos/metabolismo , Dietilexilftalato/toxicidade , Simulação por ComputadorRESUMO
OBJECTIVES: Malaria is an important selective force for human genetic adaptation due to the sustained, lethal impact it has had on populations worldwide. High frequencies of both hemoglobin S and the null allele FYBES of the Duffy blood group have been found in areas where this disease is endemic, attributed to the protective action of the carriers of these variants against malaria infection. The objective of this work was to perform ancestral reconstruction and analyze the correlation of the frequencies of these alleles throughout the phylogeny of 24 human populations. METHODS: A tree topology and the allelic frequencies reported in the literature for the 24 populations were used. The ancestral frequencies for the two alleles were reconstructed using the maximum likelihood method and the Brownian model of evolution (CI = 95%), and the correlation analysis was performed using phylogenetically independent contrasts (PICs). Statistical analyses were performed with the statistical software R version 3.4.1. RESULTS: For both alleles, a correspondence was found in the reconstruction of the ancestral frequencies, and a significant statistical correlation (p = .001) was observed between the S and FYBES alleles. CONCLUSIONS: These results provide evidence of an epistatic relationship between the two alleles, which may influence the fitness of the individuals who present with them when they are subjected to a selective force such as malaria.
Assuntos
Hemoglobina Falciforme , Malária , Humanos , Hemoglobina Falciforme/genética , Genótipo , Alelos , Sistema do Grupo Sanguíneo Duffy/genética , Frequência do Gene , Malária/genéticaRESUMO
Pregnancy in Sickle Cell Disease (SCD) women is associated to increased risk of clinical and obstetrical complications. Placentas from SCD pregnancies can present increased abnormal findings, which may lead to placental insufficiency, favoring adverse perinatal outcome. These placental abnormalities are well known and reported, however little is known about the molecular mechanisms, such as epigenetics. Thus, our aim was to evaluate the DNA methylation profile in placentas from women with SCD (HbSS and HbSC genotypes), compared to uncomplicated controls (HbAA). We included in this study 11 pregnant women with HbSS, 11 with HbSC and 21 with HbAA genotypes. Illumina Methylation EPIC BeadChip was used to assess the whole placental DNA methylation. Pyrosequencing was used for array data validation and qRT-PCR was applied for gene expression analysis. Our results showed high frequency of hypermethylated CpGs sites in HbSS and HbSC groups with 73.5% and 76.2% respectively, when compared with the control group. Differentially methylated regions (DMRs) also showed an increased hypermethylation status for the HbSS (89%) and HbSC (86%) groups, when compared with the control group methylation data. DMRs were selected for methylation validation (4 DMRs-HbSS and 3 DMRs the HbSC groups) and after analyses three were validated in the HbSS group, and none in the HbSC group. The gene expression analysis showed differential expression for the PTGFR (-2.97-fold) and GPR56 (3.0-fold) genes in the HbSS group, and for the SPOCK1 (-2.40-fold) and ADCY4 (1.80-fold) genes in the HbSC group. Taken together, these data strongly suggest that SCD (HbSS and HbSC genotypes) can alter placental DNA methylation and lead to gene expression changes. These changes possibly contribute to abnormal placental development and could impact in the clinical course, especially for the fetus, possibly leading to increased risk of abortion, fetal growth restriction (FGR), stillbirth, small for gestational age newborns and prematurity.
Assuntos
Anemia Falciforme , Doença da Hemoglobina SC , Anemia Falciforme/complicações , Anemia Falciforme/genética , Epigênese Genética , Feminino , Doença da Hemoglobina SC/genética , Hemoglobina Falciforme/genética , Humanos , Recém-Nascido , Placenta/metabolismo , Gravidez , Proteoglicanas/metabolismoRESUMO
Hemoglobin S is caused by a nucleotide change in HBB gene (HBB:c.20A>T, p.Glu6Val), is presented in diverse forms: simple carriers (HbSA), homozygotes (HbSS) also known as sickle cell anemia, and compound heterozygotes with other ß-hemoglobinopathies. It is worldwide distributed, in Mexico, is frequently observed in the southern states Guerrero, Oaxaca and Chiapas. Elevated fetal hemoglobin (HbF) is associated with mild phenotype; single-nucleotide variants (SNVs) in modifier genes, such as BCL11A, HBG2, HBBP1 pseudogene and HBS1L-MYB intergenic region, upregulate HbF synthesis. The aim of this study was to identify HbF regulating genetic variants in HbSS and HbSA Mexican subjects. We studied 39 individuals (HbSS = 24, 61%, HbSA = 15, 39%) from Chiapas (67%) and Guerrero (33%), peripheral blood was collected in ethylenediamine tetraacetic acid (EDTA) for molecular and hematological studies, DNA was isolated by salting-out technic and genotyping was performed through allelic discrimination by real time polymerase chain reaction (RT-PCR) using Taqman® probes for 15 SNV (in BCL11A: rs6706648, rs7557939, rs4671393, rs11886868, rs766432, rs7599488, rs1427407; HBS1L-MYB: rs28384513, rs7776054, rs9399137, rs4895441, rs9402686, rs1320963; HBG2: rs7482144; and HBBP1: rs10128556). The obtained data were analyzed using IMB SPSS v.22.0 software. All minor alleles were observed in frequencies over 0.05, the most frequent was rs9402686 (0.82), while the less frequent was rs101028556 (0.08). In HbSS group, the mean fetal hemoglobin was 11.9 ± 5.9% and was significantly elevated in BCL11A rs11886868 wildtype homozygotes and in carriers of HBS1L-MYB intergenic region rs7776054 (p = 0.04 and p = 0.03, respectively). In conclusion, in HbSS Mexican patients, two SNVs were observed related to increased HbF; BCL11A rs11886868 and HBS1L-MYB rs7776054.
Sickle cell anemia (SCA) is one of the most common types of hemoglobinopathies in people of African ancestry, it is caused by homozygosity of HbS mutation (HBB:c.20A>T). It is known that fetal hemoglobin plays a key role in decreasing HbS polymerization which damages the erythrocyte structure and is responsible for the characteristic hemolytic crises endured by these patients. Single-nucleotide variant (SNV) in genes that regulate fetal hemoglobin (HbF) after birth have been associated with its increment, thus ameliorating the hematologic phenotype of this pathology and other ß-hemoglobinopathies. Therefore, in this study, we identified, for the first time in Mexican patients with SCA (HbSS) and HbS carriers (HbSA), the presence of 15 SNVs on BCL11A, HBS1L-MYB and HBG2; all HbSS patients had anemia and elevated HbF; 2 variants were related to increased HbF rs11688888C of BCL11A and rs7776054G of HBSIL-MYB; and finally, all minor alleles were found at a frequency higher than 0.05.
Assuntos
Anemia Falciforme , Hemoglobina Fetal , DNA Intergênico , Ácido Edético , Hemoglobina Fetal/genética , Hemoglobina Falciforme/genética , Heterozigoto , Homozigoto , Humanos , México , Nucleotídeos , Polimorfismo de Nucleotídeo Único , Proteínas Repressoras/genéticaRESUMO
OBJECTIVE: To report the diagnostic challenges of newborn screening for abnormal haemoglobins. SETTING: Cord blood samples from 13 hospitals in southwest Jamaica taken in 2008-2019. METHODS: Blood spots, collected from the umbilical cord, were analysed by high pressure liquid chromatography (HPLC) to reveal phenotypes for HbSS and HbCC, but genotype confirmation may require parental studies or gene sequencing. Such cases that were successfully traced were analysed in this follow-up study. RESULTS: HPLC screening of 121,306 samples detected HbAS in 11,846 (9.8%), HbAC in 4508 (3.7%) and other electrophoretic abnormalities in 1090 babies. Among 101 previously unconfirmed cases, 34/90 (38%) with HPLC evidence of a HbSS phenotype had other genotypes, and 7/11 (64%) with a HbCC phenotype had other genotypes. Syndromes from the interaction of ß thalassaemia occurred in 112 babies (85 with HbS, 27 with HbC) and of genes for hereditary persistence of fetal haemoglobin (HPFH) in 18 (12 with HbS, 6 with HbC). Variants other than HbS and HbC occurred in 270 babies, 16 in combination with either HbS or HbC, and 254 as traits. Most variants are benign even when inherited with HbS, although HbO Arab, HbD Punjab, or Hb Lepore Washington, which occurred in 6 cases, may cause sickle cell disease. CONCLUSIONS: Genes for ß thalassaemia and HPFH are common in western Jamaica and when associated with HbS may present diagnostic challenges in newborns, as HbF and HbA2 have not reached diagnostic levels. Family and DNA studies may be necessary for genotype confirmation.
Assuntos
Anemia Falciforme , Hemoglobinas Anormais , Talassemia beta , Anemia Falciforme/diagnóstico , DNA , Seguimentos , Hemoglobina Falciforme/genética , Hemoglobinas Anormais/genética , Humanos , Recém-Nascido , Jamaica , Triagem Neonatal/métodosRESUMO
INTRODUCTION: The hematological and clinical features vary markedly between the different genotypes of sickle cell disease. Even within the single genotype of homozygous sickle cell disease (HbSS), there is marked variability that is presumed to result from interacting genetic and environmental factors. AREAS COVERED: The classification of the different genotypes of sickle cell disease with approximate prevalence at birth in different communities and some of the major clinical and hematological differences. This assessment includes three potential genetic factors influencing hematology and clinical outcome in HbSS, the beta globin haplotype, alpha thalassemia, and persistence of fetal hemoglobin (HbF). EXPERT OPINION: The author is a clinician with experience of sickle cell disease primarily in Jamaica but also in Greece, Uganda, Saudi Arabia, and India. It is therefore necessarily an account of clinical data and does not address current debates on molecular mechanisms. Most data derive from Jamaica where efforts have been made to reduce any symptomatic bias by long-term follow-up of patients all over the Island and further reduced by a cohort study based on newborn screening, which has been in operation for over 48 years.
Assuntos
Anemia Falciforme , Talassemia alfa , Anemia Falciforme/diagnóstico , Anemia Falciforme/epidemiologia , Anemia Falciforme/genética , Variação Biológica da População , Estudos de Coortes , Hemoglobina Fetal/genética , Haplótipos , Hemoglobina Falciforme/genética , Humanos , Recém-Nascido , Talassemia alfa/epidemiologia , Talassemia alfa/genética , Globinas beta/genéticaRESUMO
BACKGROUND: Sickle cell disease (SCD) is an autosomal recessive genetic disease in which a mutation occurs in the ß-globin chain gene, resulting in abnormal hemoglobin levels. In an environment with reduced oxygen concentration, red blood cells change their conformation, resulting in chronic hemolysis and consequent anemia and vaso-occlusive crises with injuries to several organs, with a significant impairment of the osteoarticular system. This study aimed to verify the chronic osteoarticular alterations and their association with clinical and laboratory characteristics of patients with SCD with a more severe phenotype (SS and Sß0), on a steady-state fasis. METHODS: Fifty-five patients were referred to a medical consultation with a specialized assessment of the locomotor system, followed by laboratory tests and radiographic examinations. RESULTS: In total, 74.5% patients had hemoglobinopathy SS; 67.3% were female; and 78.2% were non-whites. The mean patient age was 30.5 years. Most patients (61.8%) reported up to three crises per year, with a predominance of high-intensity pain (65.5%). Radiographic alterations were present in 80% patients. A total of 140 lesions were identified, most which were located in the spine, femur, and shoulders. Most lesions were osteonecrosis and osteoarthritis and were statistically associated with the non-use of hydroxyurea. CONCLUSIONS: There was a high prevalence of chronic osteoarticular alterations, which was statistically associated only with the non-regular use of hydroxyurea.
Assuntos
Anemia Falciforme/complicações , Osteoartrite/diagnóstico por imagem , Osteonecrose/diagnóstico por imagem , Adolescente , Adulto , Anemia Falciforme/sangue , Anemia Falciforme/tratamento farmacológico , Antidrepanocíticos/uso terapêutico , Estudos Transversais , Feminino , Hemoglobina Falciforme/genética , Humanos , Hidroxiureia/uso terapêutico , Masculino , Osteoartrite/epidemiologia , Osteonecrose/epidemiologia , Pigmentação da Pele , Adulto JovemRESUMO
BACKGROUND AND AIMS: Hepatic crisis is an emergent complication affecting patients with sickle cell disease (SCD); however, the molecular mechanism of sickle cell hepatobiliary injury remains poorly understood. Using the knock-in humanized mouse model of SCD and SCD patient blood, we sought to mechanistically characterize SCD-associated hepato-pathophysiology applying our recently developed quantitative liver intravital imaging, RNA sequence analysis, and biochemical approaches. APPROACH AND RESULTS: SCD mice manifested sinusoidal ischemia, progressive hepatomegaly, liver injury, hyperbilirubinemia, and increased ductular reaction under basal conditions. Nuclear factor kappa B (NF-κB) activation in the liver of SCD mice inhibited farnesoid X receptor (FXR) signaling and its downstream targets, leading to loss of canalicular bile transport and altered bile acid pool. Intravital imaging revealed impaired bile secretion into the bile canaliculi, which was secondary to loss of canalicular bile transport and bile acid metabolism, leading to intrahepatic bile accumulation in SCD mouse liver. Blocking NF-κB activation rescued FXR signaling and partially ameliorated liver injury and sinusoidal ischemia in SCD mice. CONCLUSIONS: These findings identify that NF-κB/FXR-dependent impaired bile secretion promotes intrahepatic bile accumulation, which contributes to hepatobiliary injury of SCD. Improved understanding of these processes could potentially benefit the development of therapies to treat sickle cell hepatic crisis.
Assuntos
Anemia Falciforme/complicações , Bile/metabolismo , Colestase/etiologia , Insuficiência Hepática/etiologia , Fígado/patologia , Adolescente , Adulto , Anemia Falciforme/sangue , Anemia Falciforme/tratamento farmacológico , Anemia Falciforme/genética , Animais , Ductos Biliares Intra-Hepáticos/diagnóstico por imagem , Ductos Biliares Intra-Hepáticos/patologia , Colestase/patologia , Colestase/prevenção & controle , Modelos Animais de Doenças , Feminino , Técnicas de Introdução de Genes , Hemoglobina Falciforme/genética , Insuficiência Hepática/patologia , Insuficiência Hepática/prevenção & controle , Humanos , Microscopia Intravital , Fígado/diagnóstico por imagem , Masculino , Camundongos , Pessoa de Meia-Idade , NF-kappa B/antagonistas & inibidores , NF-kappa B/efeitos dos fármacos , NF-kappa B/metabolismo , Receptores Citoplasmáticos e Nucleares/metabolismo , Transdução de Sinais/efeitos dos fármacos , Adulto JovemRESUMO
We described the clinical, laboratory and molecular characteristics of individuals with Hb S (HBB: c.20A>T)/ß-thalassemia (Hb S/ß-thal) participating in the Recipient Epidemiology and Donor Evaluation Study (REDS-III) Brazil Sickle Cell Disease cohort. HBB gene sequencing was performed to genotype each ß-thal mutation. Patients were classified as Hb S/ß0-thal, Hb S/ß+-thal-severe or Hb S/ß+-thal based on prior literature and databases of hemoglobin (Hb) variants. Characteristics of patients with each ß-thal mutation were described and the clinical profile of patients grouped into Hb S/ß0-thal, Hb S/ß+-thal and Hb S/ß+-thal-severe were compared. Of the 2793 patients enrolled, 84 (3.0%) had Hb S/ß0-thal and 83 (3.0%) had Hb S/ß+-thal; 40/83 (48.2%) patients with Hb S/ß+-thal had mutations defined as severe. We identified 19 different ß-thal mutations, eight Hb S/ß0-thal, three Hb S/ß+-thal-severe and eight Hb S/ß+-thal. The most frequent ß0 and ß+ mutations were codon 39 (HBB: c.118C>T) and IVS-I-6 (T>C) (HBB: c.92+6T>C), respectively. Individuals with Hb S/ß0-thal had a similar clinical and laboratory phenotype when compared to those with Hb S/ß+-thal-severe. Individuals with Hb S/ß+-thal-severe had significantly lower total Hb and Hb A levels and higher Hb S, white blood cell (WBC) count, platelets and hemolysis markers when compared to those with Hb S/ß+-thal. Likewise, individuals with Hb S/ß+-thal-severe showed a significantly higher occurrence of hospitalizations, vaso-occlusive events (VOE), acute chest syndrome (ACS), splenic sequestration, blood utilization, and hydroxyurea (HU) therapy.
Assuntos
Anemia Falciforme/epidemiologia , Anemia Falciforme/genética , Hemoglobina Falciforme/genética , Mutação , Globinas beta/genética , Talassemia beta/epidemiologia , Talassemia beta/genética , Adolescente , Adulto , Alelos , Anemia Falciforme/diagnóstico , Anemia Falciforme/patologia , Brasil/epidemiologia , Criança , Códon , Estudos de Coortes , Análise Mutacional de DNA , Feminino , Expressão Gênica , Frequência do Gene , Genótipo , Humanos , Incidência , Masculino , Fenótipo , Índice de Gravidade de Doença , Talassemia beta/diagnóstico , Talassemia beta/patologiaRESUMO
In regions with an Afro-descendant population and where malaria is endemic, high frequencies of polymorphisms have been found that confer resistance to this disease, such as the haemoglobin S (HbS) and Duffy genes, which provide resistance to P. falciparum and P. vivax infection, respectively. The objective of this study was to evaluate the individual and joint selection actions of these two genes in an Afro-descendant Colombian population. A total of 819 individuals were analysed using stratified random sampling. PCR-RFLP and Hardy-Weinberg equilibrium deviation analysis (H-W eq.), linkage disequilibrium (LD), D'IS2 and D'ST2 indexes, neutrality tests, correlations and fitness were performed using Arlequin 3.5.2.2 and R 3.4.1 software. In general, the population showed neutrality and H-W eq. for the HbS gene but not for the Duffy gene (FYA/FYB, FYA/FYBES and FYB/FYBES genotypes were responsible for this deviation). LD between the HbS locus and the promoter region of the Duffy gene, a value D'IS2 = 0.001 and D'ST2 = 0.020 was found, an increase in fitness of the AS*FYBES/FYBES genotype combination (marked in adolescents and adults), and a strong correlation between these genotypes (Rho = 90%, p = .001) were found, evidencing a possible joint selection action for these two alleles. This work presents evidence of the action of natural selection, both individually and jointly, on malaria resistance genes, HbS and Duffy, in the Buenaventura population.