RESUMEN
Variations at the KLF1 gene have been associated with a series of human erythroid phenotypes including the In-(Lu) phenotype, hereditary persistence of fetal hemoglobin, congenital dyserythropoietic anemia, borderline HbA(2) and increased red blood cell protoporphyrin. Natural mutations have shown that KLF1 regulates gamma globin gene expression and its role in the switching from fetal to adult globin expression has been suggested by experimental studies. In this paper we report that subjects with S270X KLF1 mutations show a decrease of HbF levels with increasing age, supporting in vivo the role of KLF1 in hemoglobin switching in humans.
Asunto(s)
Envejecimiento/genética , Hemoglobina Fetal/genética , Regulación del Desarrollo de la Expresión Génica/fisiología , Hemoglobina A/genética , Factores de Transcripción de Tipo Kruppel/genética , Polimorfismo de Nucleótido Simple , Adolescente , Niño , Preescolar , Hemoglobina Fetal/metabolismo , Técnicas de Genotipaje , Hemoglobina A/metabolismo , Humanos , Lactante , Italia , Estudios Longitudinales , Mutación , Adulto Joven , gamma-Globinas/genéticaRESUMEN
The persistence of high fetal hemoglobin level in adults may ameliorate the clinical phenotype of beta-thalassemia and sickle cell anemia. Several genetic variants responsible for hereditary persistence of fetal hemoglobin, linked and not linked to the beta globin gene cluster, have been identified in patients and in normal individuals. Monoallelic loss of KLF1, a gene with a key role in erythropoiesis, has been recently reported to be responsible for persistence of high levels of fetal hemoglobin. In a Sardinian family, high levels of HbF (22.1-30.9%) were present only in compound heterozygotes for the S270X nonsense and K332Q missense mutations, while the isolated S270X nonsense (haploinsufficiency) or K332Q missense mutation were associated with normal HbF levels (<1.5%). Functionally, the K332Q Klf1 mutation impairs binding to the BCl11A gene and activation of the γ- and ß-globin promoters. Moreover, we report for the first time the association of KLF1 mutations with very high levels of zinc protoporphyrin.
Asunto(s)
Eritrocitos/metabolismo , Hemoglobina Fetal/metabolismo , Factores de Transcripción de Tipo Kruppel/genética , Mutación , Protoporfirinas/metabolismo , Adulto , Secuencia de Bases , Análisis Mutacional de ADN , Salud de la Familia , Femenino , Fluorometría , Células HEK293 , Heterocigoto , Humanos , Italia , Masculino , Persona de Mediana Edad , Linaje , Reacción en Cadena de la PolimerasaRESUMEN
BACKGROUND AND OBJECTIVES: Patients with beta-thalassemia, like those with genetic hemochromatosis, develop iron overload due to increased iron absorption, and their iron burden is further exacerbated by transfusion therapy. Hepcidin, a hepatic hormone, regulates systemic iron homeostasis by inhibiting the absorption of iron from the diet and the recycling of iron by macrophages. In turn, hepcidin release is increased by iron loading and inhibited by erythropoietic activity. Hepcidin deficiency is the cause of iron overload in most forms of hereditary hemochromatosis. We sought to determine hepcidin's role in the pathogenesis of iron overload in b-thalassemia. DESIGN AND METHODS: We assessed the degree of iron overload in thalassemia intermedia and major patients by measuring hepatic iron concentration in liver biopsy samples and serum ferritin, estimated erythropoietic drive by assaying soluble transferrin receptor and serum erythropoietin levels and correlated these with urinary hepcidin measurements. RESULTS: Urinary hepcidin levels in beta-thalassemia demonstrate severe hepcidin deficiency in thalassemia intermedia. There was a strong inverse relationship between urinary hepcidin levels and both erythropoietin and soluble transferrin receptor, markers of erythropoietic activity. In contrast, hepcidin levels were elevated in thalassemia major, presumably due to transfusions that reduce erythropoietic drive and deliver a large iron load. Despite similar liver iron concentrations in the two conditions, serum ferritin was much lower in thalassemia intermedia. INTERPRETATION AND CONCLUSIONS: In thalassemia intermedia, high erythropoietic drive causes severe hepcidin deficiency. The lack of hepcidin results in hyperabsorption of dietary iron, but also in iron depletion of macrophages, lowering their secretion of ferritin and, consequently, serum ferritin levels. In contrast, in thalassemia major, transfusions decrease erythropoietic drive and increase the iron load, resulting in relatively higher hepcidin levels. In the presence of higher hepcidin levels, dietary iron absorption is moderated and macrophages retain iron, contributing to higher serum ferritin. In the future, hepcidin measurements may allow a more accurate assessment of the degree of iron overload and the maldistribution of iron in thalassemia.
Asunto(s)
Péptidos Catiónicos Antimicrobianos/fisiología , Sobrecarga de Hierro/etiología , Hierro/metabolismo , Hígado/metabolismo , Talasemia beta/metabolismo , Adolescente , Adulto , Péptidos Catiónicos Antimicrobianos/deficiencia , Péptidos Catiónicos Antimicrobianos/genética , Péptidos Catiónicos Antimicrobianos/orina , Transfusión Sanguínea , Terapia por Quelación , Terapia Combinada , Deferoxamina/uso terapéutico , Eritropoyesis , Femenino , Ferritinas/sangre , Ferritinas/metabolismo , Regulación de la Expresión Génica , Hepcidinas , Humanos , Absorción Intestinal , Hierro/análisis , Quelantes del Hierro/uso terapéutico , Sobrecarga de Hierro/metabolismo , Hierro de la Dieta/farmacocinética , Hígado/química , Macrófagos/metabolismo , Masculino , Índice de Severidad de la Enfermedad , Transferrina/análisis , Talasemia beta/tratamiento farmacológico , Talasemia beta/genética , Talasemia beta/terapiaRESUMEN
Two beta-thalassaemia patients, whose constitutive genotype was beta(39C)/beta(39C-->T), had the clinical phenotype beta-thalassaemia intermedia. Analysis of leucocyte DNA showed the presence of the mutated beta(39C-->T)-gene exclusively, while the normal beta(39C)-gene was also present in reticulocyte RNA. Deletional analysis of chromosome 11p15.5 on leucocyte DNA showed large deletions including the beta-globin gene. Two populations of erythroid progenitors, one heterozygous and the other hemizygous for the beta(39C-->T) mutation, were demonstrated in one case. This confirms that, in heterozygous individuals, beta-thalassaemia intermedia may be caused by inactivation of the beta-locus in trans as a result of chromosome 11p15.5 deletions in a subpopulation of haematopoietic cells.
Asunto(s)
Cromosomas Humanos Par 11 , Eliminación de Gen , Globinas/genética , Talasemia/genética , Adulto , Cromatografía Líquida de Alta Presión , Femenino , Hemoglobina Fetal/análisis , Hemoglobina A/análisis , Hemoglobina A2/análisis , Heterocigoto , Humanos , Masculino , ARN Mensajero/análisis , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Talasemia/sangre , Talasemia beta/sangre , Talasemia beta/genéticaRESUMEN
The phylogeography of Y-chromosome haplogroups E (Hg E) and J (Hg J) was investigated in >2400 subjects from 29 populations, mainly from Europe and the Mediterranean area but also from Africa and Asia. The observed 501 Hg E and 445 Hg J samples were subtyped using 36 binary markers and eight microsatellite loci. Spatial patterns reveal that (1). the two sister clades, J-M267 and J-M172, are distributed differentially within the Near East, North Africa, and Europe; (2). J-M267 was spread by two temporally distinct migratory episodes, the most recent one probably associated with the diffusion of Arab people; (3). E-M81 is typical of Berbers, and its presence in Iberia and Sicily is due to recent gene flow from North Africa; (4). J-M172(xM12) distribution is consistent with a Levantine/Anatolian dispersal route to southeastern Europe and may reflect the spread of Anatolian farmers; and (5). E-M78 (for which microsatellite data suggest an eastern African origin) and, to a lesser extent, J-M12(M102) lineages would trace the subsequent diffusion of people from the southern Balkans to the west. A 7%-22% contribution of Y chromosomes from Greece to southern Italy was estimated by admixture analysis.
Asunto(s)
Cromosomas Humanos Y/genética , Variación Genética , Haplotipos/genética , Polimorfismo Genético , África del Norte , Alelos , Europa (Continente) , Frecuencia de los Genes , Genética de Población , Geografía , Humanos , Masculino , Región Mediterránea , Repeticiones de Microsatélite , Medio Oriente , Filogenia , Recombinación GenéticaRESUMEN
An immunologically mediated pathway has been largely accepted to be one of the mechanisms involved in the clearance of senescent or prematurely damaged RBC. According to this pathway, RBC removal is mediated by binding of naturally occurring IgG to clustered integral membrane proteins, followed by complement deposition. The validation of an immunoenzymatic method for the detection of RBC-bound autologous IgG is presented. The use of RBC-bound IgG as an index related to red cell age was evaluated by measuring IgG binding in RBC treated with the clustering agent ZnCl2, in density fractionated RBC and in a selected group of patients expected to have an altered RBC life span. The immunoenzymatic method for IgG detection resulted to be reproducible (CV = 3.4%). IgG binding to in vitro clustered RBC was found to be enhanced to a very great extent, about 20 times higher with respect to untreated RBC. A slight but significant increase (about 1.8-fold) in membrane-bound IgG was observed in the highest density fraction of normal RBC, which constituted 1% of the total cells. A significantly greater number of RBC-bound IgG was measured in splenectomized beta-thalassemia intermedia patients and in subjects with secondary decreases in the C3 complement fraction concentration.
Asunto(s)
Envejecimiento Eritrocítico , Eritrocitos/inmunología , Inmunoglobulina G/análisis , Biomarcadores/análisis , Estudios de Casos y Controles , Membrana Eritrocítica/inmunología , Eritrocitos/fisiología , Humanos , Métodos , Reproducibilidad de los Resultados , Talasemia beta/sangreRESUMEN
The clinical phenotype of homozygous beta thalassemia varies in severity from the mild thalassemia intermedia to the severe thalassemia major. This variability depends largely on the molecular heterogeneity of beta thalassemia defects. We report the first case of a homozygous state for nondeletion Sardinian delta-beta(0) thalassemia, which resulted in a symptomless clinical phenotype with a peculiar hemoglobin (Hb) pattern (99.8% Hb F and 0.2% Hb A(2)). The molecular defect was characterized by the presence of 2 nucleotide substitutions: -196C>T in the promoter of the Agamma-globin gene and beta 39C>T nonsense mutation. The absence of typical beta thalassemia clinical findings was due to the high Hb F output, which compensated for the absence of beta chains. The near absence of Hb A(2) may have resulted from either alterations in the globin gene transcriptional complex with preferential activation of gamma-globin genes and suppression of delta-globin genes or preferential survival of red blood cells with the highest Hb F content and low Hb A(2) level.
Asunto(s)
Globinas/genética , Talasemia beta/genética , Femenino , Homocigoto , Humanos , Lactante , Familia de Multigenes , Mutación , Eliminación de Secuencia , Talasemia beta/fisiopatologíaRESUMEN
We describe a new two-step culture method for mass production in vitro of erythroid cells from either CD34+ (10(5) cells/mL) or light-density (10(6) cells/mL) cells purified from the blood of normal donors and thalassemic patients. The method includes (i) culture of the cells in the presence of dexamethasone and estradiol (10(-6) M each) and (ii) the growth factors SCF (50 ng/mL), IL-3 (1 ng/mL), and EPO (1 U/mL). In their proliferative phase, these cultures generated approximately 1.2 x 10(7) erythroblasts for each milliliter of blood collected from normal donors or thalassemic patients. They were composed mostly (90%) of CD45(low)/glycophorin (GPA)(neg)/CD71(1ow) cells at day 7, 50-60\% of which became CD45(neg)/GPA+/CD71high by days 15-20. However, when cells from days 7 to 12 of the proliferative phase were transferred in differentiation medium containing EPO and insulin, they progressed to mature erythroblasts (g90% benzidine(pos) and CD45(neg)/GPA+/CD71medium) in 4 days. Because of the high number of erythroid cells that are generated from modest volumes of blood, this method will prove useful in donor-specific studies of erythroid differentiation.