Macrophages: key players in erythrocyte turnover

ABSTRACT The development of red blood cells (RBCs), or erythropoiesis, occurs in specialized niches in the bone marrow, called erythroblastic islands, composed of a central macrophage surrounded by erythroblasts at different stages of differentiation. Upon anemia or hypoxemia, erythropoiesis extends to extramedullary sites, mainly spleen and liver, a process known as stress erythropoiesis, leading to the expansion of erythroid progenitors, iron recruitment and increased production of reticulocytes and mature RBCs. Macrophages are key cells in both homeostatic and stress erythropoiesis, providing conditions for erythroid cells to survive, proliferate and differentiate. During RBCs aging and injury, macrophages play a fundamental role again, performing the clearance of these cells and recycling iron for new erythroblasts in development. Thus, macrophages are crucial components of the RBCs turnover and in this review, we aimed to cover the main known mechanisms involved in the process of birth and death of RBCs, highlighting the importance of macrophage functions in the whole RBC lifecycle.

Characteristics of bone marrow compensatory erythropoiesis in hereditary spherocytosis / 中华血液学杂志

Objective: To reveal the compensatory features of bone marrow (BM) erythropoiesis in hereditary spherocytosis (HS) and to explore the effect of diferent hemoglobin levels on this compensation. Methods: Clinical and laboratory data of patients with HS were collected, and the peripheral blood absolute reticulocytes counts value was taken as the surrogate parameter to evaluate the ability of erythropoiesis compensation. BM erythropoiesis compensation in HS with diferent degrees of anemia were evaluated. Results: ①Three hundred and two patients were enrolled, including 115 with compensated hemolytic disease, 74 with mild anemia, 90 with moderate anemia, and 23 with severe anemia. ②Hemoglobin (HGB) was negatively correlated with serum erythropoietin in the decompensated hemolytic anemia group (EPO; rs=-0.585, P<0.001) . ③The median absolute reticulocyte count (ARC) of HS patients was 0.34 (0.27, 0.44) ×10(12)/L, up to 4.25 times that of normal people. The maximum ARC was 0.81×10(12)/L, about 10 times that of normal people. The median ARC of patients with compensated hemolytic disease was 0.29 (0.22, 0.38) ×10(12)/L, up to 3.63 times that of normal people. The median ARC of patients with hemolytic anemia was 0.38 (0.30, 0.46) ×10(12)/L, which was significantly higher than the patients with compensated hemolytic disease, up to 4.75 times that of normal people (z=4.999, P=0.003) . ④ ARC was negatively correlated with HGB in the compensated hemolytic disease group (rs=-0.177, P=0.002) and positively correlated with HGB in the decompensated hemolytic anemia group (rs=0.191, P=0.009) . There was no significant difference in the ARC among patients with mild, moderate, and severe anemia (χ(2)=4.588, P=0.101) . ⑤The median immature reticulocyte production index of the mild, moderate, and severe anemia groups was 13.1% (9.1%, 18.4%) , 17.0% (13.4%, 20.8%) , and 17.8% (14.6%, 21.8%) , respectively; the mild anemia group had lower index values than the moderate and severe anemia groups (P(adj) values were both<0.05) , but there was no significant difference between the latter groups (P(adj)=1.000) . The median immature reticulocyte count of patients in the mild, moderate, and severe groups was 5.09 (2.60, 7.74) ×10(10)/L, 6.24 (4.34, 8.83) ×10(10)/L, and 7.00 (3.07, 8.22) ×10(10)/L, respectively; there was no significant difference among the groups (χ(2)=3.081, P=0.214) . Conclusion: HGB can be maintained at a normal level through bone marrow erythropoiesis, while red blood cells are reduced in HS. However, once anemia develops, the bone marrow exerts its maximum erythropoiesis capacity and does not increase, regardless of anemia aggravation or serum EPO increase.

Do blood parasites increase immature erythrocytes and mitosis in amphibians? / ¿Causan los parásitos sanguíneos un aumento en los eritrocitos inmaduros y la mitosis en los anfibios?

Abstract Introduction: In amphibians, blood may act as a hematopoietic tissue. However, the knowledge concerning hematological features is scarce, there is not much information that allows an analysis about the possible explanations of this physiological feature. Objective: This study aimed to evaluate the relationship between immature red blood cells (RBCs) mitosis and the presence of blood parasites in amphibians. Methods: We sampled 116 amphibians (31 species) in six Colombian localities. Blood was taken by cardiac puncture or maxillary vein puncture. Smears were prepared, fixed, and Giemsa stained for microscopical analysis. The variables analyzed were the percentage of immature RBCs, mitotic cells in peripheral blood, and blood parasite infection. Data were analyzed using Wilcoxon's rank test and exact Fisher statistical tests. Results: Sixty-two individuals showed mitosis in peripheral blood, and these mitotic RBCs shared morphological features with immature RBCs. Overall, parasite prevalence was 30.1 %, distributed as follows: Trypanosoma (24.1 %), Hepatozoon-like (6 %), Dactylosoma (4.3 %), Karyolysus-like (0.9 %), and Filarioidea (2.6 %). A positive association between the percentage of immature RBCs and the presence of mitotic RBCs was found, and also between the blood parasite infection and the percentage of immature RBCs. Conclusions: In this study, we found that the presence of blood parasites, immature RBCs, and RBCs mitosis are frequent events in amphibians' peripheral blood, and our analysis suggests an association between those features. Thus, the release of immature RBCs and the mitosis of those cells in peripheral blood may be a physiological response to blood parasite infection. Further studies characterizing hematology in amphibians and wildlife, in general, are desirable.

Resumen Introducción: En anfibios, la sangre puede actuar como un tejido hematopoyético. Sin embargo, el conocimiento acerca de las características hematológicas es escaso y no hay información que permita un análisis acerca de las posibles explicaciones a este rasgo fisiológico. Objetivo: La intención de este estudio fue evaluar la relación entre la presencia de eritroblastos, mitosis de glóbulos rojos (GRs) y la infección por hemoparásito en sangre periférica de anfibios. Métodos: Se muestrearon 116 anfibios (31 especies) en seis localidades de Colombia. Se tomaron muestras de sangre mediante punción cardiaca o punción a la vena maxilar. Se prepararon extendidos sanguíneos, se fijaron y tiñeron con Giemsa para su posterior análisis por microscopía. Se analizaron variables como porcentaje de GRs inmaduros, células mitóticas en sangre periférica e infección por hemoparásitos. Los datos fueron analizados mediante el test de rango de Willcoxon y el test exacto de Fisher. Resultados: sesenta y dos individuos evidenciaron mitosis en sangre periférica y dichas mitosis compartían características morfológicas con GRs inmaduros. La prevalencia general de parásitos fue del 30.1 %, distribuido de la siguiente forma: Trypanosoma (24.1 %), Hepatozoon-like (6 %), Dactylosoma (4.3 %), Karyolysus-like (0.9 %), y Filarioidea (2. 6 %). Hay una asociación positiva entre el porcentaje de GRs inmaduros y la presencia de células mitóticas, también se encontró una relación entre la infección por hemoparásitos y el porcentaje de GRs inmaduros. Conclusiones: En este estudio encontramos que la presencia de parásitos sanguíneos, GRs inmaduros y mitosis de GRs son eventos frecuentes en sangre periférica de anfibios, y nuestros resultados sugieren una asociación entre dichas características. Por tanto, la liberación de GRs inmaduros y la mitosis de estas células en sangre periférica podría ser una respuesta fisiológica a infecciones parasitarias. Posteriores estudios que caractericen la hematología en anfibios y en vida silvestre en general, son deseables.

Aportes de la biotecnología al diagnóstico y tratamiento de la enfermedad renal crónica y comorbilidades frecuentes / Biotechnology contribution to the diagnosis and treatment of chronic kidney disease and frequent comorbidities

Este trabajo tiene como objetivo revisar las contribuciones de la biotecnología, en relación con el tratamiento, diagnóstico y la monitorización de la enfermedad renal crónica (ERC) y sus comorbilidades más frecuentes, especialmente la anemia. En relación con los tratamientos, enfocamos el desarrollo de productos biofarmacéuticos como los agentes estimulantes de la eritropoyesis (ESA), que fueron los primeros biofármacos utilizados para el tratamiento de la anemia asociada a la ERC; analizamos sus características y utilización actual después de varios años de experiencia clínica, así como también otras alternativas en desarrollo. Revisamos distintos tipos de bioterapias, la utilización de las células estromales mesenquimales de médula ósea (MSC) y tratamientos alternativos con modificaciones dietarias, que se basan en la asociación entre la microbiota intestinal de los pacientes renales crónicos y sus condiciones fisiopatológicas. Finalmente, en relación con el diagnóstico y monitorización, nos referimos al estudio y validación de biomarcadores diagnósticos, predictivos y terapéuticos que han permitido optimizar los resultados clínicos en este tipo de pacientes. (AU)

The aim of this work is to review the contributions of biotechnology, in relation to the treatment, diagnosis and monitoring of chronic kidney disease (CKD) and its most frequent comorbidities, especially anemia. Regarding the treatment, we focus on the development of biopharmaceutical products such as erythropoiesis stimulating agents (ESA), which were the first biopharmaceuticals used to treat anemia associated with chronic kidney disease (CKD). We analyzed their characteristics and their current use after several years of clinical experience, as well as other alternatives in development. We also review different types of biotherapies, the use of bone marrow mesenchymal stromal cells (MSC) and alternative treatments with dietary modifications, which are based on the association between the intestinal microbiota of chronic kidney patients and their pathophysiological conditions. Finally, in relation to diagnosis and monitoring, we refer to the study and validation of diagnostic, predictive and therapeutic biomarkers that have made clinical results possible to be optimized in this type of patient. (AU)

Anemias hemolíticas hereditarias por defectos en la síntesis de globina / Hereditary hemolytic anemias due to defective globin synthesis

Introducción: Los defectos genéticos en la molécula de hemoglobina se dividen en aquellos que tienen una tasa reducida de producción de una o más cadenas de globina, las talasemias; y en los que se producen cambios estructurales que conducen a inestabilidad o transporte anormal de oxígeno. Objetivo: Explicar los diferentes mecanismos por los cuales ocurren las talasemias y otras alteraciones en la síntesis de las cadenas de globina, así como las características moleculares, fisiopatogénicas y los cambios hematológicos. Métodos: Se realizó una revisión de la literatura, en inglés y español, a través del sitio web PubMed y el motor de búsqueda Google académico de artículos publicados en los últimos 10 años. Se hizo un análisis y resumen de la bibliografía revisada. Análisis y síntesis de la información: Las talasemias son un grupo heterogéneo de defectos genéticos en la síntesis de hemoglobina, que causa una disminución en la tasa de producción de una o más cadenas de la molécula. De acuerdo a la cadena de globina que presenta el defecto se dividen en α-β-, δβ- o γδβ-talasemias. Conclusiones: Las talasemias y las hemoglobinopatías son las enfermedades hemolíticas hereditarias más comunes en muchas partes del mundo, caracterizadas por complejas interacciones entre anemia, eritropoyesis ineficaz y alteraciones del metabolismo del hierro(AU)

Introduction: Genetic disorders in the hemoglobin molecule are divided into those that have a reduced rate of production of one or more globin chains, thalassemias; and those in which structural changes occur that lead to instability or abnormal oxygen transport. Objective: To explain the different mechanisms by which thalassemias and other alterations in the synthesis of globin chains occur, as well as molecular, physiopathogenic and hematological changes. Methods: A review of the literature in English and Spanish was carried out through the PubMed website and the Google Scholar search engine, searching for articles published in the last ten years. The revised bibliography was analyzed and summarized. Information analysis and synthesis: Thalassemias make up a heterogeneous group of genetic defects in the synthesis of hemoglobin, which causes a decrease in the rate of production of one or more chains of the molecule. According to the globin chain that presents the defect, they are divided into α-β-, δβ- or γδβ-thalassemias. Conclusions: Thalassemias and hemoglobinopathies are the most common hereditary hemolytic diseases in many parts of the world. They are characterized by complex interactions between anemia, ineffective erythropoiesis, and alterations in iron metabolism(AU)

New Oral Agent for Treatment of Anemia in Patient with Chronic Kidney Disease: Prolyl Hydroxylase Inhibitor / 대한내과학회지

Hypoxia inducible factor (HIF)-stabilizers are being developed for the renal anemia treatment. This small molecules inhibit prolyl hydroxylase domain (PHD)-containing enzymes, causing HIF activation instead of degradation under the state of normoxia, finally increase production of intrinsic erythropoiesis. Current treatment guidelines suggest that renal anemia should be treated mainly with iron and erythropoiesis stimulating agents (ESAs). But there are several complications and concerns such as hypertension, ESA refractory anemia and increased cardiovascular mortality in using ESAs. Advantages of HIF stabilizers over ESAs are orally available, no dose-up requirement for inflammation. So far new HIF stabilizers showed efficacy and safety in renal anemia treatment. This new therapeutic agent may emerge as a standard treatment option for renal anmia treatment.

Epoetin alfa resistance in hemodialysis patients with chronic kidney disease: a longitudinal study

Anemia is an inevitable complication of hemodialysis, and the primary cause is erythropoietin deficiency. After diagnosis, treatment begins with an erythropoiesis-stimulating agent (ESA). However, some patients remain anemic even after receiving this medication. This study aimed to investigate the factors associated with resistance to recombinant human erythropoietin therapy with epoetin alfa (αEPO). We performed a prospective, longitudinal study of hemodialysis patients receiving treatment with αEPO at our reference hospital from July 2015 to June 2016. Clinical data was collected, and the response to αEPO treatment was evaluated using the erythropoietin resistance index (ERI). The ERI was defined as the weekly weight-adjusted αEPO dose (U/kg per week)/hemoglobin level (g/dL). A longitudinal linear regression model was fitted with random effects to verify the relationships between clinical and laboratory data and ERI. We enrolled 99 patients (average age, 45.7 (±17.6) years; male, 51.5%; 86.8% with hypertension). The ERI showed a significant positive association with serum ferritin and C-reactive protein, percentage interdialytic weight gain, and continuous usage of angiotensin receptor blocker (ARB) hypertension medication. The ERI was negatively associated with serum iron and albumin, age, urea reduction ratio, and body mass index. Our findings indicate that resistance to αEPO was related to a low serum iron reserve, an inflammatory state, poor nutritional status, and continuous usage of ARBs.

Niveles de eritropoyetina y reticulocitos en residentes de bajas alturas migrantes a medianas alturas / Erythropoietin and reticulocyte levels in lowland residents migrating to middle altitudes

RESUMEN Introducción: La eritropoyetina es una alfa globulina glicosilada con producción renal en más del 90% en la vida adulta. Es la principal hormona en el mantenimiento constante de la masa eritrocitaria, aunque existen modificaciones en sus niveles asociados con el tabaquismo, anemias, EPOC y la migración de bajas a medianas o altas alturas. Esto último desencadena un proceso hipóxico que puede llegar a producir mal agudo de montaña. Objetivo: Describir el comportamiento de la eritropoyetina, el recuento de reticulocitos y su influencia, en procesos de adaptación a la altura. Metodología: Estudio descriptivo de corte transversal que incluyó 11 participantes provenientes de bajas alturas a quienes se les determinó la concentración sérica de eritropoyetina y el recuento de reticulocitos en un periodo de 28 días. Resultados: Ocho de los participantes presentaron un ascenso progresivo en los niveles séricos de eritropoyetina, uno mantuvo una curva plana y dos presentaron comportamiento atípico respecto a lo reportado en la literatura. Conclusión: La eritropoyetina es un factor fundamental que marca el comienzo de la eritropoyesis, cuya finalidad es mejorar el aporte de oxígeno en procesos de adaptación a la altura. Además, la hipoxia es un factor determinante en el inicio y desarrollo del mal agudo de montaña. El recuento de reticulocitos depende del estímulo proliferativo y anti-apoptótico de la eritropoyetina, así como de las concentraciones séricas de vitamina B12, hierro y ácido fólico.

ABSTRACT Introduction: EPO is a glycosylated alpha globulin produced in more than 90% by kidneys through adult life, being a key hormone that regulated the erythrocytic mass. However, there are some modifications in the levels of this hormone that may be related to smoking, anemia, EPOC and migration to from low to higher altitudes, inducing hypoxic processes. Depending on the individual, it may produced the disease named as acute mountain sickness. Objective: To describe erythropoietin level modifications, reticulocytes count and its influence, on the adaptive process to altitude. Methodology: This is a transversal descriptive study including 11 participants from low altitudes places, whose EPO serum concentration and reticulocytes count was determined during 28 days. Results: Eight participants presented a progressive increase in EPO serum levels, one participant exhibited a constant level and two more showed atypical results according to previous literature. Conclusion: EPO is a key factor for determining the erythropoiesis beginning, as its objective is to improve the oxygen provision during altitude adaptation processes by increasing its concentration in blood due to hypoxic stimulus. Besides, hypoxia is a determinant factor in the beginning and development of acute mountain sickness. The reticulocytes count depends also on the EPO proliferative and anti-apoptotic stimulus, and on the serum concentrations of B12 vitamin, iron and folic acid.

TET2 is upregulated during erythroid differentiation of CD34+ cells from healthy donors and myelodysplastic syndrome patients

Background: Tet methylcytosine dioxygenase 2 (TET2) is frequently mutated and/or downregulated in myeloid neoplasm, including myelodysplastic syndromes. Despite the extensive studies, the specific contribution of TET2 in disease phenotype of myeloid neoplasms is not fully elucidated. Recent findings have grown attention on the role of TET2 in normal and malignant erythropoiesis. Methods: In the present study, we investigated TET2 mRNA levels by quantitative PCR during erythropoietin-induced erythroid differentiation CD34+ cells from healthy donor and myelodysplastic syndrome patients. Statistical analyses were performed using the ANOVA and Bonferroni post hoc test and a p-value <0.05 was considered statically significant. Results: TET2 expression is upregulated during erythroid differentiation of CD34+ cells from healthy donor and myelodysplastic syndrome patients. Conclusions: Our findings corroborate that TET2 is involved in the erythrocyte differentiation (AU)

Treatment of renal anemia: Erythropoiesis stimulating agents and beyond

Anemia, complicating the course of chronic kidney disease, is a significant parameter, whether interpreted as subjective impairment or an objective prognostic marker. Renal anemia is predominantly due to relative erythropoietin (EPO) deficiency. EPO inhibits apoptosis of erythrocyte precursors. Studies using EPO substitution have shown that increasing hemoglobin (Hb) levels up to 10–11 g/dL is associated with clinical improvement. However, it has not been unequivocally proven that further intensification of erythropoiesis stimulating agent (ESA) therapy actually leads to a comprehensive benefit for the patient, especially as ESAs are potentially associated with increased cerebro-cardiovascular events. Recently, new developments offer interesting options not only via stimulating erythropoeisis but also by employing additional mechanisms. The inhibition of activin, a member of the transforming growth factor superfamily, has the potential to correct anemia by stimulating liberation of mature erythrocyte forms and also to mitigate disturbed mineral and bone metabolism as well. Hypoxia-inducible factor prolyl hydroxylase inhibitors also show pleiotropic effects, which are at the focus of present research and have the potential of reducing mortality. However, conventional ESAs offer an extensive body of safety evidence, against which the newer substances should be measured. Carbamylated EPO is devoid of Hb augmenting effects whilst exerting promising tissue protective properties. Additionally, the role of hepcidin antagonists is discussed. An innovative new hemodialysis blood tube system, reducing blood contact with air, conveys a totally different and innocuous option to improve renal anemia by reducing mechanical hemolysis.

Regulation of the embryonic erythropoietic niche: a future perspective

The production of red blood cells, termed erythropoiesis, occurs in two waves in the developing mouse embryo: first primitive erythropoiesis followed by definitive erythropoiesis. In the mouse embryo, both primitive and definitive erythropoiesis originates in the extra-embryonic yolk sac. The definitive wave then migrates to the fetal liver, fetal spleen and fetal bone marrow as these organs form. The fetal liver serves as the major organ for hematopoietic cell expansion and erythroid maturation after mid-gestation. The erythropoietic niche, which expresses critical cytokines such as stem cell factor (SCF), thrombopoietin (TPO) and the insulin-like growth factors IGF1 and IGF2, supports hematopoietic expansion in the fetal liver. Previously, our group demonstrated that DLK1⁺ hepatoblasts support fetal liver hematopoiesis through erythropoietin and SCF release as well as extracellular matrix deposition. Loss of DLK1⁺ hepatoblasts in Map2k4(−/−) mouse embryos resulted in decreased numbers of hematopoietic cells in fetal liver. Genes encoding proteinases and peptidases were found to be highly expressed in DLK1⁺ hepatoblasts. Capitalizing on this knowledge, and working on the assumption that these proteinases and peptidases are generating small, potentially biologically active peptides, we assessed a range of peptides for their ability to support erythropoiesis in vitro. We identified KS-13 (PCT/JP2010/067011) as an erythropoietic peptide-a peptide which enhances the production of red blood cells from progenitor cells. Here, we discuss the elements regulating embryonic erythropoiesis with special attention to niche cells, and demonstrate how this knowledge can be applied in the identification of niche-derived peptides with potential therapeutic capability.

Management of Blood Loss in Hip Arthroplasty: Korean Hip Society Current Consensus / 대한고관절학회지

The volume of hip arthroplasty is stiffly increasing because of excellent clinical outcomes, however it has not been shown to decrease the incidence of transfusions due to bleeding related to this surgery. This is an important consideration since there are concerns about the side effects and social costs of transfusions. First, anemia should be assessed at least 30 days before elective hip arthroplasty, and if the subject is diagnosed as having anemia, an additional examination of the cause of the anemia should be carried and steps taken to address the anemia. Available iron treatments for anemia take 7 to 10 days to facilitate erythropoiesis, and preoperative iron supplementation, either oral or intravenous, is recommended. When using oral supplements for iron storage, administer elemental iron 100 mg daily for 2 to 6 weeks before surgery, and calculate the dose using intravenous supplement. Tranexamic acid (TXA) is a synthetic derivative of the lysine component, which reduces blood loss by inhibiting fibrinolysis and clot degradation. TXA is known to be an effective agent for reducing postoperative bleeding and reducing the need for transfusions in primary and revision total hip arthroplasties. Patient blood management has improved the clinical outcome after hip arthroplasty through the introduction and research of various agents, thereby reducing the need for allogeneic blood transfusions and reducing the risk of transfusion-related infections and the duration of hospitalizations.

Eritropoyetina como agente eritropoyético y no eritropoyético: consideraciones terapéuticas / Erythropoietic and non-erythropoietic functions of erythropoietin: therapeutic considerations / Eritropoietina como agente eritropoiético e não eritropoiético: considerações terapêuticas / Erythropoietic and non-erythropoietic functions of erythropoietin: therapeutic considerations

La producción de glóbulos rojos es controlada continuamente para suplir la desaparición de las células envejecidas y garantizar un aporte de oxígeno adecuado a todo el organismo. La citoquina pleitrópica eritropoyetina (Epo), originalmente definida por su rol en la eritropoyesis para prevenir la muerte programada de progenitores eritroides en la médula ósea, ha demostrado un rol antiapoptótico protector sobre diversos tejidos no hematopoyéticos. A la reconocida eficacia del tratamiento con eritropoyetina recombinante humana (rhuEpo) para contrarrestar la anemia que acompaña a patologías muy diversas, se agregan algunos aspectos que impiden lograr los resultados terapéuticos esperados, ya sea por resistencia al tratamiento o por el desarrollo de efectos adversos. Con el fin de prevenir estos efectos, así como reducir las dosis de rhuEpo en tratamientos crónicos se han desarrollado nuevos agentes que presentan modificaciones estructurales de la Epo, o bien alteraciones en las propiedades/actividad de la Epo nativa. Dado que, actualmente, los resultados sobre los efectos de la Epo sobre morbilidad/ mortalidad en diversas patologías no están suficientemente claros, nuevas investigaciones serán útiles para resolver dudas sobre la efectividad de la eritropoyetina y sus derivados o agentes alternativos con el fin de proveer bases sólidas para el desarrollo de ensayos clínicos concluyentes.

Erythropoietin (Epo), the cytokine required for promoting erythropoiesis through the proliferation and differentiation of erythroid cells, has been reported to act as a pleiotropic cytokine beyond the hematopoietic system. In contrast with the potentially beneficial effects attributed to recombinant human erythropoietin (rhuEpo), research has advanced to indicate that mortality and morbidity rates are increased in some patient groups when treated with rhuEpo. Some cardiac and systemic conditions may predispose to adverse events, and other factors, such as proinflammatory agents, may lead to resistance to erythropoietin treatment. Many compounds are currently under investigation in order to avoid these unwanted effects and to reduce the rhuEpo dose during chronic therapies. They are either erythropoiesis-stimulating agents different from erythropoietin or structurally modified erythropoietins with altered properties and activities. In recent reports, contrasting data have raised several concerns regarding the effectiveness of erythropoietin treatment to prevent adverse events. Therefore, much investigation is needed to provide a solid basis for the development of conclusive clinical trials.

A produção de glóbulos vermelhos é controlada continuamente para suprir o desaparecimento das células envelhecidas e garantir uma contribuição de oxigênio adequado a todo o organismo. A citocina pleiotrópica eritropoietina (Epo), originalmente definida por seu papel na eritropoiese para prevenir a morte programada de progenitores eritroides na medula óssea, tem demonstrado um papel anti-apoptótico protetor sobre diversos tecidos não hematopoiéticos. Adicionam-se à reconhecida eficácia do tratamento com eritropoietina recombinante humana (rhuEpo), para contra-arrestar a anemia que acompanha patologias muito diversas, alguns aspectos que impedem alcançar os resultados terapêuticos esperados, quer seja por resistência ao tratamento ou pelo desenvolvimento de efeitos adversos. Com o fim de prevenir estes efeitos, bem como reduzir as doses de rhuEpo em tratamentos crônicos foram desenvolvidos novos agentes que apresentam modificações estruturais da Epo, ou então alterações nas propriedades/atividade da Epo nativa. Devido a que, atualmente, os resultados sobre os efeitos da Epo sobre morbidade/mortalidade em diversas patologias não estão suficientemente claros, novas pesquisas serão úteis para resolver dúvidas sobre a efetividade da eritropoietina e seus derivados ou agentes alternativos visando a fornecer bases sólidas para o desenvolvimento de ensaios clínicos concludentes.

Expresión diferencial de isoformas del factor inducible por hipoxia (hif-1αy hif-2 α) en bazo y médula ósea en un modelo murino / Differential expression of hypoxia inducible factor isoforms (hif-1α and hif-2 α) in spleen and bone marrow in a murine model

Los factores inducibles por hipoxia (HIFs) regulan la adaptación a la hipoxia (H) y protegen a las células induciendo la transcripción de múltiples genes. Se propone estudiar la expresión de las isoformas HIF1αy HIF2αen tejidos hematopoyéticos MO y Bz durante 15 días de Hipoxia Hipobárica (HH) relacionarlos con la cinética de expresión de EPO-R y factores determinantes de la eritropoyesis GATA-1 y NFE2. Se utilizaron ratones CF1, sometidos a HH 0,4 atm de 0 a 15 días. A cada tiempo, se extrajeron fémures y Bz para la obtención de extractos, fraccionamiento proteico e inmunoblotting. Se determinaron parámetros hematológicos standard. La apoptosis fue cuantificada por TUNEL. HIFαfue evaluado en sus dos isoformas. En MO y Bz el factor de transcripción aumenta y es mayor desde el día 1 de HH en Bz. Niveles máximos de HIF1αse verifican al día 3 en MO y a partir del día 5 en Bz. HIF2αen MO presenta expresión máxima al día 2 con posterior descenso, donde la MO retoma el control de la eritropoyesis. En Bz, HIF2 exhibe patrón irregular, conservando aumentos de su inmunodetección en función de la H. Epo-R se expresa desde día 1 en MO y Bz en un patrón similar de comportamiento a GATA-1. NFE2 tiene una cinética diferencial con progresión de ascenso en MO conforme aumentan los días de H observándose un máximo al día 15 mientras que el Bz muestra paulatino descenso en función de la adaptación medular a la H. Esto sugiere que en el Bz y MO se verifican procesos adaptativos coexistentes de expansión/sobrevivencia y apoptosis de progenitores eritroides. En Bz predomina una eritropoyesis compensatoria. En MO por el contrario predomina apoptosis temprana con posterior recomposición y control de la expansión del compartimiento eritroideo. HIF1 y HIF2 se sobreexpresan en ambos tejidos, sin embargo la eritropoyesis esplénica está ligada al control de HIF1αya que aparentemente HIF2αestaría asociada a la supervivencia de otros tipos celulares esplénicos durante el estrés hipóxicoPalabras clave: Hipoxia hipobárica; Factores Inducibles por Hipoxia; Eritropoyesis

Avaliação de variantes do gene TMPRSS6 e sua relação com o status de ferro em condições de eritropoese normal e aumentada / Evaluation of TMPRSS6 genetic variants and its relationship with iron status of women with normal and increased erythropoiesis

Introdução: A deficiência de ferro é frequente entre mulheres na idade reprodutiva e é considerada problema de saúde pública mundial. Por outro lado, patologias associadas com aumentada atividade eritropoética e sobrecarga de ferro, como as talassemias e a anemia falciforme, estão entre as doenças monogênicas mais frequentes em muitas populações. Os genes HFE e TMPRSS6 codificam as proteínas hemocromatose hereditária (HFE) e matriptase-2 (MT2) que, no fígado, modulam a produção de hepcidina, o hormônio regulador central do metabolismo de ferro. O objetivo deste estudo foi avaliar a relação entre o consumo alimentar de ferro, as variantes rs855791 (MT2736V), rs4820268 (MT2521V), rs1799945 (HFE63D) e rs1800562 (HFE282Y) e o status corporal do mineral entre mulheres com atividade eritropoética normal (aparentemente saudáveis) ou levemente aumentada (com ß-talassemia menor). Casuística e métodos: Inicialmente, foram incluídas 127 estudantes universitárias na idade reprodutiva (18 a 42 anos) e em aparente balanço estacionário do ferro corporal (necessidades fisiológicas e consumo alimentar de ferro pouco variáveis há pelo menos 12 meses). Em um segundo estudo, foram incluídos 33 casos de ß-talassemia menor (18 na pós-menopausa), registrados em serviços de hematologia de dois hospitais de São Paulo-SP e um de Sorocaba-SP. Essas foram pareadas com 66 controles, segundo idade, índice de massa corporal, status reprodutivo e uso de anticoncepcionais hormonais. A partir de inquéritos feitos com registros alimentares ou recordatórios de 24 horas foi estimado o consumo de ferro total e biodisponível. Amostras de sangue foram utilizadas para a extração de DNA e determinações de ferritina sérica, saturação da transferrina e hemoglobina. As genotipagens do TMPRSS6 e do HFE foram realizadas por PCR em tempo real. Resultados: Considerando as 226 mulheres avaliadas, as frequências dos alelos MT2736V, MT2521D, HFE63D e HFE282Y foram estimadas em 40,3%, 44,0%, 16,3% e 1,5% respectivamente. No primeiro estudo, foi estimada média de consumo de ferro 10 de 10,9 mg/dia e prevalência de deficiência do mineral de 12,6%. Estimativas de consumo de ferro biodisponível, mas não de ferro total, foram correlacionadas com os valores de ferritina e saturação da transferrina. As associações entre a biodisponibilidade dietética de ferro e seus biomarcadores foram especialmente evidentes entre as carreadoras da variante HFE63D, indicando uma significante interação gene-nutriente. Por outro lado, valores relativamente menores de saturação da transferrina foram associados à presença do alelo MT2736V, independentemente do consumo de ferro biodisponível. Mulheres com ß-talassemia menor e suas controles não diferiram quanto à frequência das variantes TMPRSS6 e HFE ou à biodisponibilidade dietética de ferro. Na pós-menopausa, a ß-talassemia menor foi associada com valores duas vezes maiores de ferritina, 20% maiores de saturação da transferrina e com probabilidade 3,5 vezes maior de hiperferritinemia. Entretanto, para casos e controles na idade reprodutiva, foi estimada probabilidade de inadequação do consumo de 20,7% e prevalências de deficiência do mineral de 13,3% e 10,0%, respectivamente. Entre essas mulheres, o genótipo positivo ou negativo para a variante MT2736V foi também associado com diferenças nas médias de saturação da transferrina. No entanto, o contraste nesses valores foi relativamente maior entre as mulheres com ß-talassemia menor. Além disso, mais acentuada hipocromia acompanhou a presença da variante MT2736V nessa condição. Conclusão: Os achados sugerem que, entre mulheres com eritropoese normal ou com ß-talassemia menor, a variante MT2736V não afeta tão fortemente as reservas de ferro corporal como faz a adequação do consumo desse mineral. Ainda assim, a variante HFE63D pode modificar a relação da biodisponibilidade de ferro com seu status corporal e, portanto, ser um importante marcador preditivo de resposta diferencial à dieta. A variante MT2736V foi associada com menor disponibilidade de ferro na circulação de mulheres na idade reprodutiva e, entre aquelas com ß.;-talassemia menor, com indício de acentuada alteração morfológica dos eritrócitos

Introduction: Iron deficiency is common among women at childbearing age and it is regarded as a worldwide public health issue. On the other hand, disorders associated with increased erythropoietic activity and iron overload, such as thalassemias and sickle cell disease, are among the most frequent Mendelian diseases in many populations. HFE and TMPRSS6 genes encode the hereditary hemochromatosis protein (HFE) and the matriptase-2 (MT2) both of which modulate the hepatic production of hepcidin, the main hormone regulator of iron metabolism. The aim of this study was to evaluate the relationship among dietary iron intake, rs855791 (MT2736V), rs4820268 (MT2521D), rs1799945 (HFE63D) and rs1800562 (HFE282Y) genetic variants and body iron status of women with normal (apparently healthy) or slightly increased (ß-thalassemia minor) erythropoietic activity. Casuistic and methods: Initially, 127 university students at childbearing age (18 to 42 years old) and with steady state body iron (few variations on physiological requirements and dietary iron intake at least for the last 12 months) were included. In a second study, it was included 33 cases of ß-thalassemia minor (18 of them post-menopaused) registered in Hematology Services from two hospitals from São Paulo-SP and one from Sorocaba-SP. They were paired with 66 controls by age, body mass index, reproductive status and hormonal contraceptive use. Using food diaries or 24 hours food recalls, total and bioavailable dietary iron intakes were estimated. Blood samples were used for DNA extraction and for determinations of ferritin, transferrin saturation with iron and hemoglobin. TMPRSS6 and HFE genotyping were performed by real time PCR. Results: Considering all the 226 women studied, the allelic frequencies of MT2736V, MT2521D, HFE63D e HFE282Y genetic variants were em 40.3%, 44.0%, 16.3% e 1.5%, respectively. In the first study, a total dietary iron intake of 10.9 mg/day and an iron deficiency prevalence of 12.6% were estimated. There were correlations among ferritin and transferrin saturation values with estimates of bioavailable, but not of total dietary iron intake s. Associations between dietary iron bioavailability and iron biomarkers were especially evident among carriers of HFE63D variant, indicating a significant gene-diet interaction. On the other hand, lower levels of transferrin saturation were associated with the presence of MT2736V variant allele, irrespective of bioavailable iron intake. TMPRSS6 and HFE variants frequencies and dietary iron bioavailability estimates did not differ between women with ß-thalassemia minor and their controls. Among postmenopausal women, ß-thalassemia minor was associated with two times higher ferritin values, 20% higher transferrin saturation values and 3.5 times higher chance for hiperferritinemia. However, among cases and controls at childbearing age, it was estimated a probability of inadequacy in the dietary iron intake of 20.7% and iron deficiency prevalences of 13.3% and 10.0%, respectively. Among these women, a positive or negative genotype for MT2736V was also associated with differences in transferrin saturation. Nevertheless, the contrast between these values was relatively higher among women with ß-thalassemia minor. Moreover, a more accentuated hypochromia accompanied the presence of the MT2736V variant in this condition. Conclusions: Our findings suggest that, among women with normal erythropoiesis or ß-thalassemia minor, the presence of the MT2736V variant does not strongly impact the body iron stores as does dietary iron adequacy. Nevertheless, the HFE63D variant may modify the relationship between the dietary iron bioavailability and the body iron status. Therefore, it might be an important predictive marker of women's differential response to diet. The MT2736V variant was associated with lower availability of circulating iron in women at childbearing age and, among those with ß-thalassemia minor, with suggestive accentuated morphological alteration of erythrocytes

Effect of AB serum on human terminal erythroid differentiation ex vivo / 中南大学学报(医学版)

To demonstrate the effect of AB serum on terminal erythroid differentiation ex vivo.
 Methods: After separation of CD34+ cells from cord blood, the cells were cultured and divided into a control group and an experimental group. The effects of AB serum were examined by the expressions of different markers (GPA, Band3 and α4-integrin) for erythroblast differentiation and enucleation by flow cytometry. 
 Results: The CD34+ cells were successfully differentiated to enucleated red blood cells. There were evident differences among the expressions of GPA, Band3 and α4-integrin between the 2 groups. The percentage of GPA positive cells in the experimental group was bigger than that in the control group in every time point. The expression of Band3 in the experimental group was higher than that in the control group. The expression of α4-integrin in the experimental group was lower than that in the control group. In addition, the enucleation rate in the experimental group was higher than that in the control group.
 Conclusion: AB serum can promote the cell differentiation and enucleation during terminal erythroid differentiation in vitro.

Relationship between macrophages and erythropoiesis / 中国当代儿科杂志

Macrophages have two major roles in regulating the dynamic equilibrium in erythropoiesis, promoting the differentiation and maturation of nucleated red blood cells into reticulocytes and removing old red blood cells. A recent mouse study has demonstrated that the phenotype of macrophages in erythroblastic islands is CD169+ VCAM-1+ ER-HR3+ CD11b+ F4/80+ Ly-6G+. Molecular connections between erythroid progenitor cells and central macrophages help to maintain the function and integrity of erythroblastic islands. New research advances in Kruppel-like factor 1 (KLF1) provide new evidence for the important role of macrophages in erythroblastic islands. Macrophages play an important role in erythropoiesis both in sickness and in health, and provide a potential targeted therapy for diseases such as polycythemia vera and beta-thalassemia in the future.

Recombinant Human Erythropoietin Therapy for a Jehovah's Witness Child With Severe Anemia due to Hemolytic-Uremic Syndrome / 소아과

Patients with hemolytic-uremic syndrome (HUS) can rapidly develop profound anemia as the disease progresses, as a consequence of red blood cell (RBC) hemolysis and inadequate erythropoietin synthesis. Therefore, RBC transfusion should be considered in HUS patients with severe anemia to avoid cardiac or pulmonary complications. Most patients who are Jehovah's Witnesses refuse blood transfusion, even in the face of life-threatening medical conditions due to their religious convictions. These patients require management alternatives to blood transfusions. Erythropoietin is a glycopeptide that enhances endogenous erythropoiesis in the bone marrow. With the availability of recombinant human erythropoietin (rHuEPO), several authors have reported its successful use in patients refusing blood transfusion. However, the optimal dose and duration of treatment with rHuEPO are not established. We report a case of a 2-year-old boy with diarrhea-associated HUS whose family members are Jehovah's Witnesses. He had severe anemia with acute kidney injury. His lowest hemoglobin level was 3.6 g/dL, but his parents refused treatment with packed RBC transfusion due to their religious beliefs. Therefore, we treated him with high-dose rHuEPO (300 IU/kg/day) as well as folic acid, vitamin B12, and intravenous iron. The hemoglobin level increased steadily to 7.4 g/dL after 10 days of treatment and his renal function improved without any complications. To our knowledge, this is the first case of successful rHuEPO treatment in a Jehovah's Witness child with severe anemia due to HUS.

Genetically Engineered In Vitro Erythropoiesis

BACKGROUND: Engineered blood has the greatest potential to combat a predicted future shortfall in the US blood supply for transfusion treatments. Engineered blood produced from hematopoietic stem cell (HSC) derived red blood cells in a laboratory is possible, but critical barriers exist to the production of clinically relevant quantities of red blood cells required to create a unit of blood. Erythroblasts have a finite expansion capacity and there are many negative regulatory mechanisms that inhibit in vitro erythropoiesis. In order to overcome these barriers and enable mass production, the expansion capacity of erythroblasts in culture will need to be exponentially improved over the current state of art. This work focused on the hypothesis that genetic engineering of HSC derived erythroblasts can overcome these obstacles. OBJECTIVES: The objective of this research effort was to improve in vitro erythropoiesis efficiency from human adult stem cell derived erythroblasts utilizing genetic engineering. The ultimate goal is to enable the mass production of engineered blood. METHODS: HSCs were isolated from blood samples and cultured in a liquid media containing growth factors. Cells were transfected using a Piggybac plasmid transposon. RESULTS: Cells transfected with SPI-1 continued to proliferate in a liquid culture media. Fluorescence-activated cell sorting (FACS) analysis on culture day 45 revealed a single population of CD71+CD117+ proerythroblast cells. The results of this study suggest that genetically modified erythroblasts could be immortalized in vitro by way of a system modeling murine erythroleukemia. CONCLUSION: Genetic modification can increase erythroblast expansion capacity and potentially enable mass production of red blood cells.

¿Búsqueda de biomarcadores o construcción de un panel diagnósticopara la eritropoyesis deficiente de hierro?[Editorial] / Searching for biomarkers or better designing a diagnostic panel f: or iron-deficient erythropoiesis?[Editorial]

En la práctica clínica los exámenes de laboratorio son por lo general la piedra angular para el diagnóstico de los pacientes y la toma de decisiones médicas que se derivan de ello, por lo que la selección de los exámenes adecuados de acuerdo con las manifestaciones clínicas y la impresión diagnóstica resulta clave en la medida que permitan un diagnóstico preciso y oportuno del paciente, favorezcan el uso racional de ayudas diagnósticas y no se conviertan en un factor de confusión para el personal médico.Clásicamente, las investigaciones relacionadas con el desarrollo tecnológico del laboratorio clínico se han enfocado en desarrollar pruebas que se comporten como el estándar de oro de una enfermedadespecífica, que cuenten con alta sensibilidad, especificidad, valor pronóstico positivo y valor pronóstico negativo, en otras palabras, que sean ®invencibles¼ y permitan un diagnóstico irrefutable de la enfermedad para la cual se pretende su uso. No obstante, todas las pruebas, independiente de su complejidad, tienen limitantes preanalíticas, analíticas y posanalíticas, por lo que su aplicación clínica se debería apoyar con pruebas complementarias apropiadas para cada enfermedad y en la interpretaciónconjunta de los resultados. Por lo tanto, más que la identificación de pruebas que puedancomportarse como estándares de oro, las investigaciones y la práctica clínica se debería enfocar en el diseño de paneles diagnósticos, específicos y estandarizados para cada sospecha diagnóstica.