Resistencia osmótica eritrocitaria en el diagnóstico de anemias hereditarias en Tucumán, Argentina / Red cell osmotic resistance in the hereditary anemias diagnosis in Tucumán, Argentina / Resistência osmótica eritrocitária no diagnóstico de anemias hereditárias em Tucumán, Argentina

Las anemias hereditarias más frecuentes en Tucumán (Argentina) son el rasgo beta talasémico (RBT), las hemoglobinopatías estructurales (HBP) y la esferocitosis hereditaria (EH). La resistencia osmótica eritrocitaria inmediata y 24 horas post-incubación constituye el método diagnóstico de la EH, y como tubo único (ROETU) es usada para cribado de RBT. El propósito del trabajo fue determinar el comportamiento de ROETU (4,0 y 5,5 g/L de NaCl) en el diagnóstico de anemias hereditarias. Se estudiaron 125 pacientes: 34 normales (GN), 59 con RBT (GRBT), 21 con HBP (GHBP) y 11 con EH (GEH), que fueron agrupados en niños (≤12 años), mujeres y hombres (>12 años). Se realizaron hemograma (Coulter AcT10 y Sysmex KX-21N), índices de Mentzer y de Shine&Lal, ROETU, hierro, transferrina y saturación de transferrina (Wiener Lab), reticulocitos (azul brillante de cresilo), prueba de falciformación y electroforesis de hemoglobina a pH alcalino y ácido. GRBT presentó anemia microcítica hipocrómica, y GEH y GHBP, anemia normocítica normocrómica. El hierro fue normal. GRBT y GHBP fueron resistentes en ROETU 4,0 g/L, aunque GRBT mostró mayor resistencia (p<0,05). GEH fue menos resistente que GN en ROETU 5,5 g/L (p<0,05). ROETU 4,0 y 5,5 g/L serían recomendables en el diagnóstico presuntivo de RBT y EH, respectivamente.

Beta thalassaemia trait (BTT), structural hemoglobinopathies (SHB) and hereditary spherocytosis (HS) are the most frequent hereditary anaemias in Tucumán (Argentina). Immediately and 24 hours post-incubation red cell osmotic resistance is the diagnosis method of HS, and as a single tube (RORST), it is used for clínicamenBTT screening. The purpose of this study was to determine the RORST (NaCl 4.0 and 5.5 g/L) behaviour in the diagnosis of hereditary anemia. The study encompassed 125 patients : 34 normal patients (NG), 59 with BTT (BTTG), 21 with SHB (SHBG) and 11 with HS (HSG), who were divided into children (≤12 years), women and men (> 12 years). Blood count (Coulter AcT10 and Sysmex KX-21N), Mentzer and Shine&Lal indexes, RORST, iron, transferrin and transferrin saturation (Wiener Lab), reticulocytes (brilliant cresyl blue), sickling and hemoglobin electrophoresis at alkaline and acid pH were performed. BTTG showed hypochromic microcytic anemia, and SHBG and HSG, normochromic normocytic anemia. Iron was normal. BTTG and SHBG were resistant in RORST 4.0 g/L, but BTTG showed more resistance (p<0.05). SHG was less resistant than NG at RORST 5.5 g/L (p<0.05). RORST at values of 4.0 and 5.5 g/L would be recommended for the presumptive diagnosis of BTT and SH, respectively.

As anemias hereditárias mais comuns em Tucumán (Argentina) são o traço beta talassemia minor (BTM), as hemoglobinopatias estruturais (HBP) e esferocitose hereditária (EH). A resistência osmótica dos eritrócitos imediata e 24 horas pós-incubação é o método de diagnóstico da EH, e como um único tubo (ROETU) é usado para a detecção de BTM. O objectivo deste estudo foi determinar o comportamento de ROETU (4,0 e 5,5 g/L de NaCl) para o diagnóstico de anemias hereditárias. Foram estudados 125 pacientes: 34 normais (GN), 59 com BTM (GBTM), 21 com HBP (GHBP) e 11 com EH (GEH), que foram reunidos em crianças (≤12 anos), mulheres e homens (>12 anos). Foi realizado hemograma (Coulter AcT10 e Sysmex KX-21N), índices de Mentzer e Shine&Lal, ROETU, ferro, transferrina e saturação de transferrina (Wiener Lab), reticulócitos (azul de cresil brilhante), teste de falcização e eletroforese de hemoglobina em pH alcalino e ácido. GBTM mostrou anemia microcítica hipocrômica, e GEH e GHBP, anemia normocítica normocrômica. O ferro foi normal. GRBT e GHBP foram resistentes em ROETU 4,0 g/L, mas GBTM mostrou maior resistência (p<0,05). GEH foi menos resistente que GN em ROETU 5,5 g/L (p<0,05). ROETU 4,0 e 5,5 g/L seria recomendado para o diagnóstico presuntivo da BTM e EH, respectivamente.

Índices eritrocitarios en la esferocitosis hereditaria / Erythrocyte indexes in hereditary spherocytosis

La esferocitosis hereditaria es un grupo heterogéneo de desórdenes caracterizados por la variabilidad en la clínica, en los defectos proteicos del citoesqueleto eritrocitario y en el tipo de herencia. Se estudió la sensibilidad y especificidad de la concentración de hemoglobina corpuscular media (CHCM) y el índice de amplitud de distribución eritrocitaria (ADE) en el screening diagnóstico de la esferocitosis hereditaria. Noventa y cuatro pacientes fueron comparados con niños sanos de igual sexo y edad. Todos los índices se obtuvieron por impedancia eléctrica (autoanalizador hematológico Coulter JT). En los pacientes con esferocitosis hereditaria, la CHCM (35.67±1.33 g/dl) y el ADE (20.60±4.5%), fueron significativamente más elevados que en el grupo control (CHCM 33.48±0.68 g/dl, p 0.000; ADE 13.22±0.9%, p 0.000). Con los valores de corte utilizados en nuestro laboratorio (CHCM ≥ 34.5 g/dl; ADE ≥ 14.5%) ambos índices elevados mostraron una sensibilidad de 81% y una especificidad de 98.9% en el screening de esferocitosis hereditaria. La combinación de ambos índices es un excelente predictor para el diagnóstico de esferocitosis hereditaria.

Hereditary spherocytosis is a group of heterogenous disorders characterized by variability in its clinical manifestations, membrane protein defects and inheritance. We analysed the sensitivity and specificity of mean corpuscular hemoglobin concentration (MCHC) and red cell distribution width (RDW) in the diagnostic screening of hereditary spherocytosis. Ninetyfour patients were compared to equal number of healthy, age-matched children. All indexes were derived from measurements obtained by aperture impedance (Coulter Counter Model JT). In patients with hereditary spherocytosis, MCHC (35.67±1.33 g/dl) and RDW (20.60±4.5%) were significantly higher than in normal control subjects (MCHC 33.48±0.68 g/dl, p: 0.000; RDW 13.22±0.9%, p: 0.000). By using a cutoff for the MCHC of 34.5 g/dl and for the RDW of 14.5%, both indexes showed a sensitivity of 81% and a specificity of 98.9%. The combination of the two test is an excellent predictor for the diagnosis of hereditary spherocytosis.

Microesferocitosis: perfil eritroide y su relación con distintas pruebas de laboratorio / Microspherocytosis: erythroid profile and its relation with different laboratory tests

La esferocitosis hereditaria (EH) es una anemia hemolítica congénita con microesferocitos en sangre periférica y Coombs negativa. Hay heterogeneidad en la clínica, herencia y defectos moleculares (alteración de proteínas de la membrana), pero la EH típica es autosómica dominante, con anemia, ictericia y esplenomegalia. El objetivo de este estudio fue presentar nuestra experiencia y establecer relación entre parámetros hematológicos y bioquímicos, y pruebas de fragilidad osmótica y autohemólisis tradicionalmente diagnósticas. En nuestro medio, la EH ocupa el segundo lugar dentro de las anemias hemolíticas hereditarias, después de la b talasemia menor. El diagnóstico lo basamos en la microesferocitosis, el análisis de RGO y la autohemólisis. Se estudiaron 47 pacientes (45 de origen latino y 2, sajón). Doce pacientes no tenían antecedentes familiares. Valores promedios: hematíes (x1012/L) / hemoglobina (g/dL): niños (22): 3.84/10.5; adultos (25): mujeres (13): 3.54/10.59; hombres (12): 4.65/13.15. La autohemólisis (promedio %) estuvo muy aumentada (15.54) y corrigió con el agregado de glucosa (4.07). La fragilidad corpuscular media (promedio g/dL) estuvo incrementada tanto en sangre fresca (0.48) e incubada (0.65). Haptoglobina ausente se encontró en 76.5% de los niños y 26.7% de los adultos Reticulocitos, bilirrubina indirecta y LDH estuvieron aumentadas (medias 336.35 x109/L, 36.25 mmol/L, 236.48 UI/L, respectivamente). Microcitosis, esferocitosis, policromatofilia, acantocitosis, punteado basófilo, "células pinzadas" fueron observados en 43, 41, 41, 12, 11, 1 pacientes respectivamente. Las alteraciones morfológicas estuvieron más marcadas en los niños. Los resultados del laboratorio podrían ser consecuencia de la microcitosis más que de la esferocitosis

Abnormalities of erythrocyte membrane proteins in Korean patients with hereditary spherocytosis

Hereditary spherocytosis (HS) is a common inherited erythrocyte membrane disorder characterized by chronic hemolytic anemia. Clinical manifestations and biochemical abnormalities of HS are heterogeneous. In this study, we investigated erythrocyte membrane protein defects in 27 Korean HS cases. Utilizing both the Fairbanks system and the Laemmli system, sodium dodecyl sulfate polyacrylamide gel electrophoresis of erythrocyte membrane proteins was performed. Proteins were stained with Coomassie brilliant blue and gels were scanned using a densitometer. We detected spectrin deficiency in 7.4% of cases (2/27), ankyrin deficiency in 29.6% (8/27), combined spectrin and ankyrin deficiency in 3.7% (1/27), band 3 deficiency in 11.1% (3/27) and protein 4.2 deficiency in 14.8% (4/27). Membrane protein deficiencies were not observed in nine cases (33.3%, 9/27). Members of two of seven families tested showed the same protein defects as the proband. Ankyrin deficiency alone and combined with spectrin deficiency accounted for 33.3% of cases (9/27), and they were the most common biochemical defects in Korean HS cases. Protein 4.2 deficiency caused HS more frequently in Koreans than in Caucasians.

Abnormalities of erythrocyte membrane proteins in Korean patients with hereditary spherocytosis

Hereditary spherocytosis (HS) is a common inherited erythrocyte membrane disorder characterized by chronic hemolytic anemia. Clinical manifestations and biochemical abnormalities of HS are heterogeneous. In this study, we investigated erythrocyte membrane protein defects in 27 Korean HS cases. Utilizing both the Fairbanks system and the Laemmli system, sodium dodecyl sulfate polyacrylamide gel electrophoresis of erythrocyte membrane proteins was performed. Proteins were stained with Coomassie brilliant blue and gels were scanned using a densitometer. We detected spectrin deficiency in 7.4% of cases (2/27), ankyrin deficiency in 29.6% (8/27), combined spectrin and ankyrin deficiency in 3.7% (1/27), band 3 deficiency in 11.1% (3/27) and protein 4.2 deficiency in 14.8% (4/27). Membrane protein deficiencies were not observed in nine cases (33.3%, 9/27). Members of two of seven families tested showed the same protein defects as the proband. Ankyrin deficiency alone and combined with spectrin deficiency accounted for 33.3% of cases (9/27), and they were the most common biochemical defects in Korean HS cases. Protein 4.2 deficiency caused HS more frequently in Koreans than in Caucasians.

Usefulness of cryohemolysis test in the diagnosis of hereditary spherocytosis

The clinical suspicion of hereditary spherocytosis (HS) must be confirmed at the clinical laboratory. The osotic fragility test (OFT) and the autohemolysis test (AHT) are the worlwide accepted assays to establish a definitive diagnosis of HS; however, they have some disadvantages. We describe herein our experience with the cryohemolysis test (CHT) as a tool to confirm the HS diagnosis. We included four groups of subjects, namely, patients with clinical HS, patients with mechanical heart valve prosthesis, malignant hematological diseases and healthy blood donors. CHT was carried out in all the groups, while OFT and AHT only in the HS patients and healthy individuals. OFT and AHT were performed according to previously described techniques. CHT was performed using red blood cells incubated in a hypertonic solution, preheated for 10 min and then tranferred to an ice bath for an additional 10 min. The resulting cryohemolysis was determined mesuring the free hemoglobin in the sample. There were no differences among the groups in terms of general characteristics. All HS suspicious patients had a positive OFT and AHT. CHT was positive in all patients from the HS group but in none of the subjects from the control groups (p<0.001). We found that CHT is a faster and easier-to-perform assay compared with OFT and AHT. Moreover, using CHT, the zone between normal and abnormal results is wider than OFT or AHT. We propose 0.7 to 11 percent hemolysis as reference values for CHT

Esferocitosis hereditaria en el período de recién nacido / Clinical manifestations of heredity sphecocytosis in the neonatal period

Se analizan las características clínicas y hematológicas de 7 RN portadores de EH, 5 de los cuales tenían antecedentes familiares positivos. Los 7 casos presentaron ictericia precoz con ascenso importante de la bilirrubinemia que obligó a usar fototerapia en todos y en 3 a realizar exsanguinotransfusión. Los 7 enfermos presentaron anemia importante que debió tratarse con transfusión de glóbulos rojos. La mayoría de los casos presentó como evolución una forma leve o moderada tal como sucede en la EH diagnosticada más tardíamente. Se acusan los datos de la literatura que coinciden con los hallazgos anotados. Como referencia se analizan otros 29 casos de EH diagnosticados en edades más tardías encontrando el antecedente de ictericia neonatal patológica en el 44,8% lo que sugiere que la hemolisis con manifestaciones clínicas está presente en el período neonatal con mucho más frecuencia de la diagnosticada. Se pone énfasis en que debe investigarse la EH en todos los casos de ictericia neonatal de causa no explicada y especialmente si se acompaña de anemia. Deben investigarse los antecedentes familiares de EH y hacer un estudio hematológico completo que incluya el examen de fragilidad osmótica. El diagnóstico diferencial más importante es con la enfermedad hemolítica por incompatibilidad ABO