Clinical and gene mutation characteristics of patients with hereditary ellipsocytosis: nine cases report and literature review / 中华血液学杂志

Objective: To report gene mutations in nine patients with hereditary elliptocytosis (HE) and analyze the characteristics of pathogenic gene mutations in HE. Methods: The clinical and gene mutations of nine patients clinically diagnosed with HE at Institute of Hematology & Blood Diseases Hospital from June 2018 to February 2022 were reported and verified by next-generation sequencing to analyze the relationship between gene mutations and clinical phenotypes. Results: Erythrocyte membrane protein gene mutations were detected among nine patients with HE, including six with SPTA1 mutation, one with SPTB mutation, one with EPB41 mutation, and one with chromosome 20 copy deletion. A total of 11 gene mutation sites were involved, including 6 known mutations and 5 novel mutations. The five novel mutations included SPTA1: c.1247A>C (p. K416T) in exon 9, c.1891delG (p. A631fs*17) in exon 15, E6-E12 Del; SPTB: c.154C>T (p. R52W) ; and EPB41: c.1636A>G (p. I546V) . Three of the six patients with the SPTA1 mutation were SPTA1 exon 9 mutation. Conclusion: SPTA1 is the most common mutant gene in patients with HE.

Genetic Analysis of a Chinese Pedigree with Hereditary Spherocytosis Caused by Copy Number Variation Deletion of SPTB Gene / 中国实验血液学杂志

OBJECTIVE@#To investigate the molecular mechanism of the disease based on the clinical characterization and genetic mutation analysis in a family with hereditary spherocytosis.@*METHODS@#The proband with jaundice and anemia was referred to Yidu Central Hospital of Weifang in May 2021. Peripheral blood samples were collected from six members of the family. Second-generation sequencing was used to screen the pathological mutations, and the clinically significant variant sites were selected. Then the relevant databases were used to analyze the variant sites, and RT-qPCR was used to detect the relative mRNA levels of candidate gene. The structure and function of SPTB protein were analyzed by UniProt and SMART databases.@*RESULTS@#We infer that the SPTB gene copy number variation (CNV) deletion was co-segregated with the phenotype of the patients in this family based on the results of second-generation sequencing (about 700 target genes). The UCSC Genome Browser demonstrated that the deleted region was mainly located in exon2-3 of SPTB gene. The results of RT-qPCR showed that the relative SPTB mRNA levels of all patients were lower than the healthy control. UniProt and SMART databases analysis showed that SPTB protein without CH1 and CH2 domains could not bind to erythrocyte membrane actin.@*CONCLUSION@#The CNV deletion of SPTB gene may be the reason for the hereditary spherocytosis in this family.

Analysis of the characteristics of SPTB gene variants among 16 children with Hereditary spherocytosis / 中华医学遗传学杂志

OBJECTIVE@#To analyze the clinical characteristics and spectrum of SPTB gene variants among 16 Chinese children with Hereditary spherocytosis (HS) and explore their genotype-phenotype correlation.@*METHODS@#Sixteen children who were diagnosed with HS at the Affiliated Hospital of Capital Institute of Pediatrics from November 2018 to July 2022 were selected as the research subjects. Genetic testing was carried out by whole exome sequencing. Candidate variants were verified by Sanger sequencing and subjected to bioinformatic analysis and prediction of 3D structure of the protein. Correlation between the SPTB genotypes and clinical phenotypes was analyzed using Chi-squared test.@*RESULTS@#The male-to-female ratio of the HS patients was 6 : 10, with the median age being 7-year-and-10-month. Clinical features of the patients have included anemia, reticulocytosis and gradual onset of splenomegaly. Mild, moderate and severe anemia have respectively occurred in 56.25% (9/16), 31.25% (5/16) and 12.50% (2/16) of the patients. SPTB gene variants were detected in all patients, among which 10 were unreported previously and 7 were de novo in origin. Loss of function (LOF) variants accounted for 93.75% (15/16). Only one missense variant was detected. Eleven, 4 and 1 of the variants had occurred in the repeat domain, CH1 domain, and dimerization domain, respectively. There was no significant correlation between the type or domain of the SPTB gene variants with the clinical features such as severity of anemia (x² = 3.345, P > 0.05). All of the variants were predicted to be pathogenic or likely pathogenic based on the guidelines from the American College of Medical Genetics and Genomics.@*CONCLUSION@#Mild to moderate anemia are predominant clinical features of the HS children harboring a SPTB gene variant, for which LOF variants are the main mutational type. The clinical feature of HS is unaffected by the type of the variants.

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.

Genetic Analysis and Prenatal Diagnosis of a Family with Hereditary Spherocytosis Caused by a Novel Compound Heterozygous Mutation of SPTB Gene / 中国实验血液学杂志

OBJECTIVE@#To investigate the clinical and genetic characteristics of a family with hereditary spherocytosis (HS), to clarify the cause of the disease, and to provide the basis for genetic counseling and prenatal diagnosis.@*METHODS@#The clinical data of proband and his parents were collected, and HS-related pathogenic genovariation of the proband was detected by high throughput sequencing. Suspected pathogenic mutation sites were verified by PCR-Sanger sequencing, and the fetus were conceived by a proband mother underwent prenatal diagnosis.@*RESULTS@#Clinical manifestations of the proband showed moderate anemia, mild splenomegaly, and jaundice (an indirect increase of bilirubin). The gene detection showed that the proband showed compound heterozygous mutations of SPTB gene c. 6095T > C (p.Leu2032Pro) and c. 6224A > G (p.Glu2075Gly), which was inherited from the asymptomatic mother and father, respectively. Both mutations were detected rarely in the common population. Prenatal diagnosis revealed that the fetus inherited a mutant gene of the mother.@*CONCLUSION@#The compound heterozygous mutations of SPTB genes c.6095T>C (p.Leu2032Pro) and c.6224A>G (p.Glu2075Gly) were the causes of the family disease, which provides a basis for family genetic counseling and prenatal diagnosis. This report is the first one found in the HGMD,1000G and EXAC database, which provides an addition to the mutation profile of the SPTB gene.

Hiperplasia eritroblástica secundaria a infección aguda por parvovirus B19

Resumen La infección aguda por parvovirus B19 es una enfermedad autolimitada en pacientes sin trastornos inmunitarios. Sin embargo, en pacientes con discrasias sanguíneas pueden manifestarse con una crisis aplásica. Presentamos el caso de un varón de 48 años, con una esferocitosis hereditaria no diagnosticada previamente, la cual debutó con una crisis aplásica inducida por una infección aguda de parvovirus B19. La sospecha clínica se planteó luego del análisis histopatológico de la médula ósea, en el que se observó una hiperplasia eritroblástica, con precursores eritroides gigantes e inclusiones nucleares virales, y cuyo análisis inmunohistoquímico fue positivo para la proteína de la cápside viral VP1 y VP2 de parvovirus B19 en células infectadas. Se confirmó la sospecha diagnóstica con la detección de anticuerpos IgM de parvovirus B19. De acuerdo a nuestra revisión, este es el primer reporte de un adulto en Latinoamérica que debutó con una crisis aplásica inducida por una infección aguda por parvovirus B19, como primera manifestación de una esferocitosis hereditaria.

Abstract Acute parvovirus B19 infection is a self-limiting disease in patients with normal immune response. However, in patients with blood dyscrasias, it is possible to present with an aplastic crisis. We present the case of a 48-year-old man who had developed an aplastic crisis as a result of an acute parvovirus B19 infection with an undiagnosed hereditary spherocytosis. Suspicions of the parvovirus infection began to arise after a routine bone marrow histopathological analysis which showed erythroblastic hyperplasia with giant erythroid precursor and viral inclusions. A subsequent immunohistochemical analysis tested positive for VP1 and VP2 capsid proteins of parvovirus B19 in infected cells. The diagnostic suspicion was later confirmed with the presence of anti-parvovirus B19 IgM. According to our review, this is the first published case in Latin America that documents an adult patient with normal immune response whose first symptom of hereditary spherocytosis was an aplastic crisis induced by an acute parvovirus B19 infection.

Conducción anestésica de un paciente pediátrico con micro esferocitosis hereditaria / Anesthetic approach on a pediatric patient with hereditary microspherocytosis

Introducción: La esferocitosis hereditaria (ESH) es una anemia hemolítica de observación frecuente, en la cual existen defectos cualitativos o cuantitativos de algunas proteínas de la membrana eritrocitaria que llevan a la formación de hematíes de forma esférica, osmóticamente frágiles, que son atrapados de formas selectiva y destruidos en el bazo, con incidencia variable y más frecuente en pacientes con descendencia europea. Objetivo: Describir la conducta clínica y anestesiológica de un paciente pediátrico con diagnóstico de micro esferocitosis hereditaria programado de forma electiva para procedimiento quirúrgico. Desarrollo: Se presenta un caso clínico de un paciente escolar con diagnóstico de micro esferocitosis hereditaria al cual se le realizó esplenectomía total electiva convencional. Con principal signo dolor a la palpación en hipocondrio izquierdo. Se condujo con anestesia total intravenosa con buenos resultados clínicos quirúrgicos, utilizando propofol a razón de 3 mcg/mL y ketamina a 0,2 mg/mL. La estrategia estuvo basada en cinco aspectos claves: evitar la hipoxia, la hipotermia, la acidosis, reducir la pérdida de sangre, así como un correcto control del dolor postoperatorio. Asociado a lo anterior es indispensable una estrecha vigilancia ya que estos pacientes pueden manifestar crisis hemolítica y aplásica. Conclusiones: El manejo perioperatorio del paciente con esferocitosis hereditaria depende de la severidad del cuadro clínico, de la anemia y su repercusión y del grado de hemólisis. La anestesia total intravenosa es una técnica segura para el tratamiento de pacientes con esferocitosis hereditaria(AU)

Introduction: Hereditary spherocytosis (HS) is a hemolytic anemia of frequent occurrence, in which there are qualitative or quantitative defects of some erythrocyte membrane proteins that lead to the formation of sphere-shaped red blood cells, which are osmotically fragile, and that are selectively trapped and destroyed in the spleen, with variable and more frequent incidence in patients with European descent. Objective: To describe the clinical and anesthesiological behavior of a pediatric patient with a diagnosis of hereditary microspherocytosis electively programmed for a surgical procedure. Development: A clinical case of a school-age patient with a diagnosis of hereditary microspherocytosis was presented. The patient underwent conventional elective total splenectomy. Pain was as the main sign on palpation to the left hypochondrium. The case was conducted with total intravenous anesthesia, with good surgical clinical results, using propofol at a rate of 3 mcg/mL and ketamine at 0.2 mg/mL. The strategy was based on five key aspects: avoid hypoxia, hypothermia, acidosis, reduce blood loss, as well as proper control of postoperative pain. Associated with the above-mentioned, close monitoring is essential, as these patients may manifest hemolytic and aplastic crisis. Conclusions: The perioperative management of the patient with hereditary spherocytosis depends on the severity of the clinical status, the anemia and its repercussion, and the degree of hemolysis. Total intravenous anesthesia is a safe technique for the treatment of patients with hereditary spherocytosis(AU)

Hereditary spherocytosis due to a novel c.5798+1G>A variant of the SPTB gene / 中华医学遗传学杂志

OBJECTIVE@#To explore the genetic basis of a pedigree affected with hereditary spherocytosis.@*METHODS@#Peripheral blood samples were collected from 17 members of the pedigree. Genomic DNA of the proband was subjected to next generation sequencing. Candidate variant was validated by co-segregation analysis. pCAS2(c.5798+1G) and pCAS2(c.5798+1A) plasmids were constructed by homologous recombination and transfected into 293T cells. Reverse transcription PCR, TA cloning and Sanger sequencing were used to analyze the effect of candidate variant on splicing. Meanwhile, peripheral blood RNAs were extracted to analyze the effect of candidate variant on splicing in vivo.@*RESULTS@#The proband was found to carry a c.5798+1G>A variant of the SPTB gene. The variant has co-segregated with the phenotype in the pedigree. In vitro and in vivo splicing experiments confirmed that the mutation has significantly affected the splicing, resulting in shift of reading frame and produced a premature termination codon.@*CONCLUSION@#The novel c.5798+1G>A variant of the SPTB gene probably underlies the pathogenesis of hereditary spherocytosis in this pedigree.

Analysis of ANK1 gene mutation in a family with hereditary spherocytosis type Ⅰ / 中华医学遗传学杂志

OBJECTIVE@#To detect the disease-causing mutation in a family with hereditary spherocytosis type Ⅰ.@*METHODS@#Genomic DNA was extracted from peripheral blood samples of the proband and his relatives. Next-generation sequencing was used to detect the mutations of relevant genes. Suspected pathogenic mutation was verified by Sanger sequencing.@*RESULTS@#The proband was found to harbor a novel frameshifting mutation in the coding region of ANK1 gene, which has resulted in abnormal structure or function of the protein. The mutation was confirmed by Sanger sequencing, with both his father and brother found to have carried the same mutation.@*CONCLUSION@#The c.247delG mutation of proband hereditary spherocytosis typeⅠin this family due to mutation of the ANK1 gene．.

Clinical and genetic features of children with hereditary spherocytosis: an analysis of 4 cases / 中国当代儿科杂志

Four children (two boys and two girls), aged from 3 years and 7 months to 5 years, had mild or moderate anemia, mild hepatosplenomegaly, jaundice (mainly an increase in indirect bilirubin), an increase in the percentages of reticulocytes and spherical erythrocytes in peripheral blood smear and an increase in erythrocyte osmotic brittleness. High-throughput sequencing found two novel mutations in the SLC4A1 gene, c.37G>A and c.340T>C, in case 1 and case 2 respectively, and these two mutations were predicted to be pathogenic by Mutation Taster. The Polyphen2 scores of these two mutations were 0.87 and 0.83 respectively, which suggested that these mutations were probably damaging. The SIFT scores of these two mutations were 0.008 and 0.09 respectively, suggesting that these mutations were probably damaging. No abnormality in this gene was found in their parents. Two reported heterozygous mutations in the ANK1 gene, c.830A>G and c.985G>C, were found in case 3 and case 4 respectively. Gene detection was not performed for the parents of case 3. The mother of case 4 was diagnosed with hereditary spherocytosis and had a heterozygous mutation of c.985G>C in the ANK1 gene. All four children were diagnosed with hereditary spherocytosis. Case 3 had a hemoglobin level of 105 g/L. Hereditary spherocytosis is a hereditary hemolytic disease caused by abnormality in erythrocyte membrane protein, and gene detection helps to make a confirmed diagnosis.

Clinical characteristics and genetic analysis of hereditary spherocytosis caused by mutations of ANK1 and SPTB genes / 中国当代儿科杂志

This study analyzed the clinical features of 5 children with hereditary spherocytosis (HS) and the characteristics of ANK1 and SPTB gene mutations. All 5 children were confirmed with HS by peripheral blood genetic detection. Anemia, jaundice and splenomegaly were observed in all 5 children. Three children had an increase in erythrocyte osmotic fragility. All 5 children had negative results of the Coombs test, glucose 6 phosphate dehydrogenase test, sucrose hemolysis test, acidified-serum hemolysis test and thalassemia gene test. Peripheral blood smear showed an increase in spherocyte count in one child. High-throughput sequencing revealed ANK1 gene mutations in patients 1 to 3, namely c.3398(exon29)delA, c.4306C>T and c.957(exon9)_c.961(exon9)delAATCT, among which c.3398(exon29)delA had not been reported before. Patient 4 had c.318delGExon3 mutation in the SPTB gene. Patient 5 had mutations in the SPTB and SLC4A1 genes, among which c.3484delC in the SPTB gene was a spontaneous mutation; the mutation site of the SLCA4A1 gene was inherited from the father and was a non-pathogenic gene. This study suggests that anemia, jaundice and splenomegaly are major clinical manifestations of HS children. Most children with HS do not have the typical spherocytic changes. Genetic detection may help with the accurate diagnosis of HS.

Associação entre eliptocitose e esferocitose hereditária em um idoso. Relato de caso / Association between elliptocytosis and hereditary spherocytosis in an elderly patient. Case report

A anemia é muito prevalente em idosos; todavia, a eliptocitose hereditária, uma anemia hereditária caracterizada pela presença de eritrócitos em forma elíptica no sangue periférico; raramente causa anemia sintomática em pacientes idosos. A eliptocitose esferocítica é uma anemia hereditária com caráter autossômico dominante. A associação entre essas duas eritroenzimopatias hereditárias é rara e seu curso varia com alterações da forma eritrocitária, observando-se a presença simultânea de eliptocitose e esferocitose no sangue periférico. A hemólise pode variar de média a moderada intensidade. Relatamos o caso de uma paciente adulta idosa com eliptocitose esferocítica revelada após 26 anos, sendo realizada a esplenectomia para resolução do quadro de hemólise.

Anemia is very prevalent in the elderly; however, hereditary elliptocytosis, an inherited anemia characterized by the presence of elliptical erythrocytes in the peripheral blood; rarely causes symptomatic anemia in elderly patients. Spherocytic elliptocytosis is an inherited anemia with an autosomal dominant character. The association between these two hereditary erythroenzyopathies is rare and its course varies with alterations of the erythrocyte form, observing the simultaneous presence of elliptocytosis and spherocytosis in the peripheral blood. Hemolysis can range from medium to moderate intensity. We report the case of a patient with spherocytic elliptocytosis revealed after 26 years, and splenectomy was performed to resolve the hemolysis.

Application of High Resolution Melting Curve Analysis in Detection of SLC4A1 Gene Mutation in Patients with Hereditary Spherocytosis / 中国实验血液学杂志

OBJECTIVE@#To investigate the feasibility and clinical significance of high resolution melting(HRM) curve analysis to detect SLC4A1 gene D38A and K56E mutations in the patients with hereditary spherocytosis(HS).@*METHODS@#Peripheral blood was collected from 23 cases of HS for routine tests and their genomic DNA was extracted by routine technique. Specific primers of mutation sites D38A and K56E of SLC4A1 gene were designed. The HRM method was used to analyze all the samples, and then the results of HRM were verified with DNA sequencing technology.@*RESULTS@#Among 23 specimens of HS patients, 6 cases of heterozygous mutant gene were detected by HRM technology, including 3 cases of D38A mutation and 3 cases of K56E mutation, which were confirmed by DNA sequencing.@*CONCLUSION@#The HRM technology can correctly detect 2 common mutation sites including D38A and K56E in SLC4A1 gene in an efficient, fast, and reliable way, which not only can be used for clinical diagnosis, but also expected to be a new method for clinical researchers to define gene mutation spectrum in HS patients.

Esferocitosis hereditaria: experiencia clínica y diagnóstica en Argentina / Hereditary spherocytosis: clinical and diagnostic experience in Argentina / Esferocitose hereditária: experiência clínica e diagnóstica na Argentina

Los objetivos del presente estudio fueron: a) Analizar las características demográficas y clínicas de nuestra población al diagnóstico; b) Evaluar si las pruebas más recientes presentan ventajas sobre las tradicionales; c) Confirmar la frecuencia de las distintas deficiencias de proteínas de membrana; d) Establecer la relación entre severidad y resultado de las pruebas o tipo de deficiencia. Se analizaron 359 individuos estudiados desde 2007, cuando se incorporaron criohemólisis hipertónica (CH), citometría de flujo con eosina-5'- maleimida (5'EMA-CF), FOE por citometría de flujo (FOE-CF) y electroforesis de proteínas de membrana (SDS-PAGE) al estudio de laboratorio clásico, fragilidad osmótica eritrocitaria (FOE) y autohemólisis (AH). Criterios diagnósticos para Esferocitosis Hereditaria (ESH): esferocitos en frotis y dos pruebas positivas. Se identificaron 174 pacientes con ESH y 22 portadores sanos. El 74,9% eran menores de 12 años. La transmisión fue dominante en el 83,1% de los casos. Tuvieron manifestaciones neonatales 89,1%. Las pruebas con mayor sensibilidad fueron CH (92,0%), FOE diferida (91,1%) y 5'EMA-CF (88,5%). En los 125 pacientes en quienes se realizaron CH, 5'EMA-CF y FOE-CF se observó que todos tenían al menos una prueba positiva; 122 (97,6%) tuvieron dos o tres positivas. Las deficiencias más frecuentes fueron ankirina y espectrina. No hubo diferencia en el resultado de las pruebas entre los subgrupos de severidad. Se concluye que las deficiencias más frecuentes en Argentina son ankirina y espectrina, coincidiendo con otras poblaciones latinoamericanas. El uso simultáneo de CH, 5'EMA-CF y FOE-CF permite diagnosticar más del 97% de los casos. La incidencia de manifestaciones neonatales es elevada.

The aims of this study were (a) to assess demographic and clinical aspects of our population at diagnosis; (b) to evaluate diagnostic accuracy of hypertonic cryohemolysis (HC), eosin-5'-maleimide flow cytometry (EMA-FC) and flow cytometric osmotic fragility (OF-FC) in relation to standard screening tests osmotic fragility (OF) and autohemolysis (AH); (c) to confirm the previously reported prevalence of membrane proteins defects; and (d) to assess the relationship between severity of anemia and results of confirmatory tests. Since 2007, the following tests were available in our laboratory: OF, AH, HC, EMA-FC, OF-FC and SDS-PAGE of membrane proteins. Diagnostic criteria for hereditary spherocytosis were spherocytes in blood smear plus ≥2 positive tests. Data from 359 individuals were analyzed: 174 HS patients and 22 silent carriers were detected; 74.9% of patients were less than 12 years old; 83.1% of them showed a dominant inheritance pattern; antecedent of neonatal jaundice/anemia was registered in 89.1%. Tests with higher sensitivity were: HC (92.0%), incubated OF (91.1%), and EMA-FC (88.5%). HC, EMA-FC and OF-FC were simultaneously performed on 125 patients: each of them had at least 1 positive test; 122 (97.6%) had 2 or 3 positive tests. Ankyrin and spectrin were the most frequently found protein deficiencies. Comparison of test results in relation to severity of anemia showed no difference between groups. It can be concluded that compared toother Latin American countries, ankyrin and spectrin were the most frequent protein deficiencies. Simultaneous performing of HC, EMA-FC and OF-FC enabled diagnosing HS in more than 97% of patients. A high incidence of neonatal jaundice/anemia was observed.

Os objetivos do presente estudo foram: a) analisar as características demográficas e clínicas de nossa população ao diagnóstico; b) Avaliar se as provas mais recentes apresentam vantagens sobre as tradicionais; c) Confirmar a frequência das diversas deficiências de proteínas de membrana; d) Establecer a relação entre severidade e resultado das provas ou tipo de deficiência. Foram analisados 359 indivíduos estudados desde 2007, quando se incorporaram crio-hemólise hipertônica (CH), citometria de fluxo com eosina-5'-maleimida (5'EMA-CF), FOE por citometria de fluxo (FOE-CF) e eletroforese de proteínas de membrana (SDS-PAGE) ao estudo de laboratório clássico - fragilidade osmótica eritrocitária (FOE) e auto-hemólise (AH). Critérios diagnósticos para ESH: esferócitos em esfregaço e duas provas positivas. Foram identificados 174 pacientes com ESH e 22 portadores sadios. 74,9% eram menores de 12 anos. A transmissão foi dominante em 83,1%. Tiveram manifestações neonatais 89,1%. As provas com maior sensibilidade foram CH (92,0%), FOE diferida (91,1%) e 5'EMA-CF (88,5%). Nos 125 pacientes aos quais lhes realizaram CH, 5'EMA-CF e FOE-CF se observou que todos tinham no mínimo uma prova positiva; 122 (97,6%) tiveram duas ou três positivas. As deficiências mais frequentes foram anquirina e espectrina. Não houve diferença no resultado das provas entre os subgrupos de severidade. Conclui-se que as deficiências mais frequentes na Argentina são anquirina e espectrina, as quais coincidem com outras populações latinoamericanas. O uso simultâneo de CH, 5'EMA-CF e FOE-CF permite diagnosticar mais de 97% dos casos. A incidência de manifestações neonatais é elevada.

Identification of a novel ANK1 gene mutation in a newborn with hereditary spherocytosis / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To determine the disease-causing mutation in a newborn with hereditary spherocytosis.</p><p><b>METHODS</b>Genomic DNA was extracted from peripheral blood samples of the patient and her parents. Next-generation sequencing was used to analyze the related genes. Suspected pathogenic mutation was verified with polymerase chain reaction and Sanger sequencing.</p><p><b>RESULTS</b>An insertional mutation g.834_833insC was identified in the coding region of ankyrin-1 (ANK1) gene, which has caused a frame shift, resulting premature termination of protein translation.</p><p><b>CONCLUSION</b>The hereditary spherocytosis in the neonate was probably due to the g.834_833insC mutation of the ANK1 gene.</p>

Establishment of Method for Detecting Red Blood Cell Osmotic Fragility by Flow Cytometry / 中国实验血液学杂志

<p><b>OBJECTIVE</b>To establish a new method for detection of red blood cell osmotic fragility by using flow cytometry.</p><p><b>METHODS</b>The hypotension salt solution of different concentrations (0.70 ml normal saline+0.3 ml deionized water, 0.60 ml normal saline+0.40 ml deionized water and 0.55 ml normal saline+0.45 ml deionized water) were prepared with normal saline and deionized water, in which the red blood cells were suspended, and the residual red blood cells were detected by flow cytometer.</p><p><b>RESULTS</b>There was no significant difference in percentage of residual red blood cells between different time points detected by flow cytometer in 3 different hypotonic salt solutions. The percentage of residual red blood cells in B+C+D+E+F+G detected time region was different among 3 NaCl dilution groups. The percentage of residual red blood cells in normal control was lower than that in hemoglobinopathy group. The percentage of residual red blood cells in hereditary spherocytosis (HS) group was obviously lower than that in hemoglobinopathy and normal control groups. The comparison of 3 different dilution concentrations found that the second concentration (0.60 ml normal saline+0.40 ml deionized water) is more suitable to screen HS by FC500 flow cytometer.</p><p><b>CONCLUSION</b>The detection of red cell osmotic fragility by using flow cytometry is a simple, rapid, objective and economic way that can be an effective screening method for diagnose the HS.</p>

Esferocitosis hereditaria: Revisión. Parte I. Historia, demografía, etiopatogenia y diagnóstico / Hereditary spherocytosis. Review. Part I. History, demographics, pathogenesis, and diagnosis

La esferocitosis hereditaria es la anemia hereditaria más frecuente en nuestro país luego de la talasemia menor. En este artículo, se revisan aspectos históricos, demográficos, genéticos y etiopatogénicos de la enfermedad, y se describen las pruebas de laboratorio para su diagnóstico. Se remarca el comportamiento de la enfermedad en nuestra población y se detallan las deficiencias proteicas predominantes en nuestro país. Se enfatiza sobre las nuevas técnicas de laboratorio actualmente disponibles, con alta sensibilidad y especificidad, que permiten realizar un diagnóstico más temprano con volúmenes de muestra mucho menores que los necesarios para las pruebas convencionales.

Hereditary spherocytosis is the most frequent hereditary anemia excluding beta thalassemia in Argentina. Historical, demographic, genetic and pathogenic aspects of the disease are reviewed, and confirmatory laboratory tests are described. Special characteristics on the outcome of the disease in our population and prevalent protein deficiencies in our country are described. Emphasis is given on new available laboratory tests, which allow an earlier diagnosis using volume of blood samples significantly smaller than required for conventional tests.

Flow cytometric test using eosin-5'-maleimide (EMA) labelling of red blood for diagnosis of hereditary spherocytosis / 中华血液学杂志

<p><b>OBJECTIVE</b>To investigate the sensitivity and specificity of eosin-5'-maleimide (EMA)assay for the diagnosis of hereditary spherocytosis (HS), and to verify the stability of reagent and samples.</p><p><b>METHODS</b>EMA flow cytometry test, NaCl-osmotic fragility test and acidified glycerol lysis test were performed using peripheral blood samples from 80 patients with HS and 44 patients with other blood diseases, the sensitivity and specificity of the three methods were compared, and the feasibility of EMA binding test was estimated. The stability of EMA reagent and HS samples stored at different temperatures were tested.</p><p><b>RESULTS</b>Among the 124 tested samples, the sensitivity and specificity of EMA binding test was 0.925 and 0.954, that of NaCl-osmotic fragility test was 0.950 and 0.455, and that of acidified glycerol lysis test was 1.000 and 0.318, respectively. Although the sensitivity of NaCl-osmotic fragility test and acidified glycerol lysis test was a little higher than that of EMA binding test, the specificity of the former two methods was poor, they couldn't clearly distinguish whether spherocytosis is hereditary spherocytosis. The experiment results showed that EMA was sensitive to the temperature and should not be stored in a small aliquots at -80 ℃ over a period of 6 months. The stability of the HS sample was better, 6 days storage at 4 ℃ and 3 days storage at room temperature had no influence on the results.</p><p><b>CONCLUSION</b>EMA binding test by flow cytometry showed good sensitivity and specificity for HS diagnosis. EMA reagent should be stored at-80 ℃ and the HS samples should be tested within 6 days storage at 4 ℃ and 3 days at room temperature.</p>

Hematopoyesis extramedular en paciente con esferocitosis hereditaria asintomática / Extramedullary hematopoiesis in patients with asymptomatic hereditry spherocytosis

En diversas patologías con alteraciones en la producción de los elementos formes de la sangre pueden desarrollarse focos de hematopoyesis extramedular en diferentes sitios. Los más frecuentes son: bazo, hígado, ganglios linfáticos, y más raramente otros órganos como: glándulas adrenales, hillios renales, cartílagos, ligamentos, tejido adiposo, timo, pulmón, mediastino y duramadre de cráneo y columna. Generalmente el proceso es difuso pero pueden formarse grandes tumores de tejido hematopoyético. Las condiciones patológicas de la médula ósea más frecuentemente asociadas a hematopoyesis extramedular son: esferocitosis hereditaria, talasemia, síndromes mieloproliferativos con fibrosis medular, ocupación medular por patologías neoplásicas. presentamos un paciente de 45 años con esferocitosis hereditaria con masas de tejido hematopoyético extramedular paravertebral mediastinal que respondieron favorablemente a la esplenectomía.

Hereditary spherocytosis (HS) is a relatively common inherited hemolytic disorder in northern Europe and in the US. The reported prevalence of HS in Western countries is 1:5000. We describe a patient 45 years old, with hereditary spherocytosis with masses of mediastinal paravertebral extramedullary hematopoietic tissue, with a favorable response to splenectomy. The medical lieterature refers some cases of extramedullary hematopoiesis as a clinical expression of hereditary spherocytosis, mainly as thoracic masses with usually paravertebral localization. HS should be distinguished from other spherocytic hermolytic anemias. Diagnosis is usually made uring infancy or in young adults, but it can be at any moment of their life, until the seventh decade of life. Ocasionally, the diagnosis is first made in old age. The clinical expression of HS is highly variable, ranging from asymptomatic condition to a severe life-threatening hemolytic anemia. Laboratory features include spherocytosis, osmotic fragility, manifestations of hemolytic disease, elevated unconjugated bilirubin and reticulocytosis. The principal diagnostic test, RBC osmotic fragility, measures the surface/volume Ratio of the cells. The treatment of choice of HS in patients with inherited spherocytosis is splenectomy, which corrects hemolytic anemia. According to he literature, cases of failure following splenectomy have been reported.

Extramedullary Hematopoiesis at the Posterior Mediastinum in Patient with Hereditary Spherocytosis: A Case Report

Thoracic extramedullary hematopoiesis (EMH) is a rare disease entity that is usually associated with hematologic disorders, such as myelodysplastic or hemolytic disease. Because thoracic EMH is usually encountered as a mass during radiologic examinations, it should be differentiated from posterior mediastinal neurogenic tumors. Here, the authors report a case of EMH associated with hereditary spherocytosis. The patient underwent a complete excision by thoracoscopic surgery to differentiate it from other mediastinal tumors.