Mild Hereditary Spherocytosis Research Articles

Congenital Dyserythropoietic Anemia Type II: An Update from the Congenital Dyseryhtropoietic Anemia Registry of North America (CDAR)

Congenital Dyserythropoietic Anemias (CDAs) are a heterogeneous group of rare anemias characterized by ineffective erythropoiesis and hemolysis. The CDA registry in North America (CDAR; NCT02964494) was established to facilitate natural history studies. Participants who do not have a genetic diagnosis are offered family-trio whole genome sequencing (WGS) and elect to provide samples to CDAR biorepository to support collaborative mechanistic studies. CDA type II (CDA-II), the most common CDA type, is an autosomal recessive disease caused by biallelic SEC23Bvariants. CDA-II presents with hemolytic anemia with suboptimal reticulocytosis, iron overload, splenomegaly, and characteristic binucleation in 10-30% of the bone marrow (BM) erythroblasts. Although CDA-II is a recessive disorder, only one SEC23B variant is found in some patients. Here, we present the genetic and clinical data of CDA-II patients enrolled in CDAR so far. As of July 2023, 169 individuals (85 patients; 84 unaffected family members) enrolled in CDAR. Nine have a confirmed CDA-II diagnosis. Six patients have biallelic SEC23B pathogenic variants and 3 have only one variant identified. WGS for the 3 patients with a single variant revealed rare deep-intronic candidate variants. Since SEC23Bis ubiquitously expressed, we studied SEC23B protein levels in patient-derived lymphoblastoid cell lines generated for the CDAR biorepository. Patients with a single SEC23B variant exhibited an expression level comparable to those with biallelic mutations (Figure). The candidate deep intronic variants are currently being investigated for pathogenicity. Seven patients presented with hemolytic anemia during infancy, but the age of diagnosis was delayed by a mean of 12.5 yrs (0.1 to 55 y.o). Blood smears showed mild macrocytosis, poikilocytosis, and several spherocytes congruent with an osmotic gradient ektacytometry resembling mild hereditary spherocytosis. BM examination, when performed, exhibited erythroid hyperplasia, prominent dyserythropoiesis, and binucleation in ≥ 10% of erythroblasts. Anemia severity was variable: 6 patients were transfusion-dependent, 1 received intermittent transfusions, and 2 did not need blood transfusions. Two became transfusion-independent after splenectomy at ages 6 and 7 y.o, and two underwent successful matched-sibling BMT at age 10 and 26 y.o, after adequate chelation (liver iron concentration [LIC]<7 mg/g) Iron overload, disproportionate to transfusions, was present in 6 patients; 5 received iron chelation. All 5 patients with available MRI results had hepatic hemosiderosis (LIC range 3.4 to 21.8 mg/g) and evidence of myocardial hemosiderosis (T2* range 6.3 to 20 ms). Ferritin values were variable (range 58 to 1631.5 ng/ml) and discordant from LIC and T2* measurements. Notably, ferritin underestimated iron overload burden: 2 patients with normal ferritin (58 and 160 ng/mL) had hepatic hemosiderosis (LIC 6.1 and 3.4 mg/g respectively), and patients with severe hepatic and myocardial hemosiderosis (LIC 16.6-21.8 mg/g; T2* 6.3-7.8 ms) had modest ferritin values 1475-1631 ng/mL. Iron overload was present even in a non-transfusion-dependent patient and worsened in a patient post-splenectomy, suggesting a role for ineffective erythropoiesis. To further investigate this possibility, we measured erythroferrone (ERFE), GDF-15, and hepcidin levels in 2 patients so far. ERFE and GDF-15 levels were remarkably higher than controls, and hepcidin-to-ferritin ratios were significantly lower, suggesting that high ERFE, caused by ineffective erythropoiesis, inappropriately suppressed hepcidin for the body's iron levels. In summary, CDA-II diagnosis is often delayed despite early onset of anemia. Genetic testing may expedite the diagnosis. The severity of anemia is variable and there are several patients requiring transfusions. The risk of iron overload, including myocardial overload, is high and appears to be related to ineffective erythropoiesis. While frequency of transfusions may improve post-splenectomy, iron overload is not determined by blood transfusions alone and does not resolve post-splenectomy. Serum ferritin underestimated the burden of tissue iron in CDA-II, suggesting that MRI scans should be employed early. Future studies may identify a role for SEC23B deep intronic variants in CDA-II patients with heterozygous variants.

Elevated Bilirubin Level Increase the Risk of Gallstone Disease in Pediatric Hereditary Spherocytosis Patients: A Case Report

Disorders of gall bladder are common and remain as one of the main patients’ chief complaints that might lead to surgical interventions. High level of bilirubin causes gallstones in patients with hereditary spherocytosis (HS). It is often misdiagnosed, because these deposits of bilirubin can be easily broken down in the gallbladder. Gallstones develop in less than half of patients with mild HS and do not always cause symptomatic biliary tract disease. HS is a genetic defect in red blood cells, caused by defects in structural membrane proteins. It is characterized by a decrease the surface area to volume ratio in erythrocytes due to the heterogeneous changes in the various genes that encode for proteins. A 14-year old male was brought to Medical Park hospital/Turki at the emergency room with a sudden onset of pain in the right upper quadrant, vomiting, anemia and jaundice. The patient was diagnosed with gallstone due to obstruction in common bile duct (CBD),with elevated bilirubin as a result for mild HS. This case report study highlight on the role of hereditary spherocytosis in forming many abdominal problem such as gallstone due to high level of heamolysis and elevated of bilirubin. How to Cite this Article Pubmed Style Najlaa Abdulameer Ali Al-Dahhan, Hawraa Abdulameer Ali Al-Dahhan, Bayan Jebur Hussein. Bilirubin Level Increase the Risk of Gallstone Disease in Pediatric Hereditary Spherocytosis Patients: A Case SRP. 2020; 11(6): 341-346. doi:10.31838/srp.2020.6.54 Web Style Najlaa Abdulameer Ali Al-Dahhan, Hawraa Abdulameer Ali Al-Dahhan, Bayan Jebur Hussein. Bilirubin Level Increase the Risk of Gallstone Disease in Pediatric Hereditary Spherocytosis Patients: A Case http://www.sysrevpharm.org/?mno=116258 [Access: March 29, 2021]. doi:10.31838/srp.2020.6.54 AMA (American Medical Association) Style Najlaa Abdulameer Ali Al-Dahhan, Hawraa Abdulameer Ali Al-Dahhan, Bayan Jebur Hussein. Bilirubin Level Increase the Risk of Gallstone Disease in Pediatric Hereditary Spherocytosis Patients: A Case SRP. 2020; 11(6): 341-346. doi:10.31838/srp.2020.6.54 Vancouver/ICMJE Style Najlaa Abdulameer Ali Al-Dahhan, Hawraa Abdulameer Ali Al-Dahhan, Bayan Jebur Hussein. Bilirubin Level Increase the Risk of Gallstone Disease in Pediatric Hereditary Spherocytosis Patients: A Case SRP. (2020), [cited March 29, 2021]; 11(6): 341-346. doi:10.31838/srp.2020.6.54 Harvard Style Najlaa Abdulameer Ali Al-Dahhan, Hawraa Abdulameer Ali Al-Dahhan, Bayan Jebur Hussein (2020) Bilirubin Level Increase the Risk of Gallstone Disease in Pediatric Hereditary Spherocytosis Patients: A Case SRP, 11 (6), 341-346. doi:10.31838/srp.2020.6.54 Turabian Style Najlaa Abdulameer Ali Al-Dahhan, Hawraa Abdulameer Ali Al-Dahhan, Bayan Jebur Hussein. 2020. Bilirubin Level Increase the Risk of Gallstone Disease in Pediatric Hereditary Spherocytosis Patients: A Case Systematic Reviews in Pharmacy, 11 (6), 341-346. doi:10.31838/srp.2020.6.54 Chicago Style Najlaa Abdulameer Ali Al-Dahhan, Hawraa Abdulameer Ali Al-Dahhan, Bayan Jebur Hussein. Elevated Bilirubin Level Increase the Risk of Gallstone Disease in Pediatric Hereditary Spherocytosis Patients: A Case Report. Systematic Reviews in Pharmacy 11 (2020), 341-346. doi:10.31838/srp.2020.6.54 MLA (The Modern Language Association) Style Najlaa Abdulameer Ali Al-Dahhan, Hawraa Abdulameer Ali Al-Dahhan, Bayan Jebur Hussein. Elevated Bilirubin Level Increase the Risk of Gallstone Disease in Pediatric Hereditary Spherocytosis Patients: A Case Report. Systematic Reviews in Pharmacy 11.6 (2020), 341-346. Print. doi:10.31838/srp.2020.6.54 APA (American Psychological Association) Style Najlaa Abdulameer Ali Al-Dahhan, Hawraa Abdulameer Ali Al-Dahhan, Bayan Jebur Hussein (2020) Bilirubin Level Increase the Risk of Gallstone Disease in Pediatric Hereditary Spherocytosis Patients: A Case Systematic Reviews in Pharmacy, 11 (6), 341-346. doi:10.31838/srp.2020.6.54

A clinical and experimental study of adult hereditary spherocytosis in the Chinese population.

Hereditary spherocytosis (HS) is often misdiagnosed due to lack of specific diagnostic methods. Our study summarized clinical characteristics and described the diagnostic workflow for mild and moderate HS in Chinese individuals, using data from 20 adults, 8 of whom presented a familial history for HS. We used scanning electron microscopy (SEM) to diagnose HS. We observed reduced eosin maleimide fluorescence activity (5.50 mean channel fluorescence (MCF) units) in the 10 cases of HS, which differed significantly when compared with 10 normal adults (15.50 units), iron deficiency anemia (15.50 MCF units), and megaloblastic anemia (12.00 MCF units) values (P < .05). Next generation sequencing results revealed that 9 out of 10 patients were found to have mutations in the spectrin alpha chain (SPTB), anchor protein (ANK1), and SLC4A1 genes. These mutations were not reported in the Human Gene Mutation Database (HGMD), 1000 human genome, ExAC, and dbSNP147 databases. Splenectomy proved to be beneficial in alleviating HS symptoms in 10 cases. It was found that for the diagnosis of HS, SEM and next generation gene sequencing method proved to be more ideal than red blood cell membrane protein analysis using sodium dodecyl sulfate polyacrylamide gel electrophoresis and western blotting.

Cholelitiasis in an adult patient with mild hereditary spherocytosis – a case report

Cholelitiasis in an adult patient with mild hereditary spherocytosis – a case report

Hereditary Spherocytosis Complicated by Choledocholithiasis: Compounded Hyperbilirubinemia

Hereditary Spherocytosis (HS) has an estimated incidence of 1 in 5000 individuals in the United States and is inherited in an autosomal recessive pattern. Hyperbilirubinemia occurs as spherical RBCs are produced, which are culled rapidly from circulation by the spleen and hemolysis occurs, thereby leading to hyperbilirubinemia. Cholelithiasis, which manifests as pigmented bilirubin stones, is a common finding and may be a signal of an RBC disorder in mild HS. The incidence of choledocholithiasis in these patients is unclear. Jaundice can worsen many reasons including fatigue, cold exposure, emotional distress, or pregnancy. We present a case of choledocholithiasis in a patient with complicated HS. Our patient is a 20-year-old female with HS diagnosed as a child. Patient was recommended for splenectomy 4 years prior; however, mother refused due to risk of complications. U/S six years prior was negative for cholelithiasis. Patient presented to the hospital in winter with worsening jaundice, abdominal pain, nausea and vomiting for 5 days. Patient was noted to have an elevated bilirubin of 27.2 on admission, which trended up to 50.4. Fractionated bilirubin showed direct bilirubin of 36.4. Right upper quadrant ultrasound showed cholelithiasis with mildly dilated CBD and no evidence of choledocholithiasis. Given primarily direct hyperbilirubinemia, MRCP was obtained showing dilated CBD to 8mm with a large stone seen in the CBD. ERCP was performed with evidence of a filling defect in the distal CBD. Sphincterotomy with balloon sweeps were performed and yielded a large pigmented stone. Patient bilirubin improved to 17.9 prior to cholecystectomy. Unfortunately, patient had a gastric perforation during splenectomy and cholecystectomy given multiple accessory spleens and complicated surgical procedure and subsequently passed away. Our case is a rare example of complicated HS as splenectomy was previously recommended and is the definitive treatment for HS. Choledocholithiasis in patients with HS has not been clearly defined. One study in India on 70 patients showed that 36.8% of patients with HS diagnosed had cholelithiasis or choledocholithiasis but did not clearly differentiate between the prevalence of choledocholithiasis. Literature review only yielded case reports of choledocholithiasis in patients with HS. Our case serves as an example of the difficulty in diagnosing choledocholithiasis as our patient had multiple risk factors for worsening jaundice.2109_A Figure 1. ERCP demonstrating filling defect at the distal CBD (arrow) suggestive of choledoholithiasis2109_B Figure 2. Endoscopic view during ERCP demonstrating large bilirubin pigmented stone extracted from balloon sweeps

Hereditary spherocytosis: Retrospective evaluation of 65 children.

Güngör A, Yaralı N, Fettah A, Ok-Bozkaya İ, Özbek N, Kara A. Hereditary spherocytosis: Retrospective evaluation of 65 children. Turk J Pediatr 2018; 60: 264-269. Hereditary spherocytosis (HS) is a common cause of congenital hemolytic anemia in Caucasians and it could be diagnosed at any age. The aim of this study is to examine the demographic characteristics, clinical features and laboratory findings of children with HS and their complications observed during follow up. Sixty-five patients, with hereditary spherocytosis between January 2008 and September 2013, were enrolled into this retrospective study. The age of patients at the time of diagnosis varied between 15 days and 17 years. The median age of patients at diagnosis was 48 months (IQR 2-78). The female/male ratio was 1.1. Forty-seven patients (72.3%) had a family history of HS. The patients were classified according to laboratory findings: 13 of them (20%) were diagnosed as mild HS, 36 (55.4%) as moderate HS and of 16 (24.6%) as severe HS. During follow-up, nine patients (13.8%) experienced an aplastic crisis. Megaloblastic crisis was not observed in any patient. Twenty patients (30.8%) had cholelithiasis. Splenectomy was performed in 20% of patients and the mean age for splenectomy was 8.3 years. Complications such as sepsis or thrombosis were not detected after splenectomy. Hereditary spherocytosis should be kept in mind in patients with anemia, jaundice and splenomegaly and the family history must be questioned. The most common complication was gallstone; even patients without severe hemolysis should be followed intermittently by abdominal ultrasonography in order to control the development of gallstone.

Long-Term Follow up of the Beneficial Effects and of Issues in Subtotal Splenectomy for Hereditary Spherocytosis

▪Total splenectomy is recommended in symptomatic cases of hereditary spherocytosis (HS) to reduce the severity of anemia but exposes patients to long-term infectious or thrombotic complications. Alternative strategies such as subtotal splenectomy (STS) have been developed, principally for children under the age of 6 with severe HS, who are not eligible to total splenectomy because of the high infectious risk and for older patients with mild HS complaining of chronic discomfort. Since our original report, several groups have shown that STS reduced the hemolytic rate, increased the red cell lifespan while maintaining an efficient splenic phagocytic function but the extent of follow-up has been limited. In order to define the long-term benefits of STS, we report here an update of our series that includes 90 patients who underwent STS at the Bicêtre hospital between 1985 and 2013, with a median-follow-up of 9.3 years (range 1-23 years). Two groups were defined on the basis of the disease phenotype: Group A included 42 patients for whom STS was performed because of severe/intermediate HS (transfusion requirement or hemoglobin (Hb) level <95 g/dL and Group B included 48 patients with a milder HS but presenting marked icterus, gallstones requiring cholecystectomy or chronic fatigue. At the time of STS, the mean hemoglobin level was 82 g/L in Group A vs. 110 g/L in Group B. Mean age at surgery was 7.6 ± 5.4 years (0.5-25 years), significantly lower in Group A than in Group B (4.3 vs. 10 years, p<0.0001). In all cases, STS was performed by laparotomy. Preoperative mean spleen volume was 513 ± 241% of normal. Surgery reduced by 88% the initial splenic volume. A functional assessment of the splenic remnant was performed by Howell jolly body counts and by splenic scintigraphy. Splenic function was retained in 87 of 90 patients except for 3 individuals (2 post-operative necrosis of the remnant, 1 non-functional remnant). No severe infection or thrombotic event was documented during the entire period of follow-up. A sustainable hematological response was observed in 81 patients (90%) at the end of the evaluation period. The mean increase in the hemoglobin level was 27 g/L, with a hemoglobin level after STS being still higher in group B than in group A (130 ± 3.3g/L vs. 110 ± 3.3 g/L (p<0.0001). For the whole population, the mean number of transfusions per year and per patient was 0.097 after versus 1.7 before STS (for Group A only: 0.21 after vs. 3.65 before STS, p<0.0001). Of note, STS decreased the hemolytic rate but did not abrogate it: the reticulocyte count decreased from 399 ±195 G/L to 290 ±170 G/L (p<0.001), but the bilirubin level remained unchanged. Seventeen patients (33%) developed cholelithiasis during the follow-up period. Gallstones were slightly more frequent in Group A than in Group B (47% vs. 15%, p=0.03). After an initial improvement, 8 patients (19% of patients from Group A) experienced a relapse with transfusion-dependant anemia and/or decrease of the hemoglobin level below 95g/L. Relapses occurred with a mean delay of 4.6 ± 1.2 years after STS. Assessing the remnant volume by ultrasonography and/or scintigraphy, we noticed that it increased quickly during the first year after STS but at a slower rate during the subsequent 5 years. The growth was faster in patients from Group A but there was no clear correlation at the end of the follow-up period between the hemoglobin level and the volume of the remnant. During the follow-up period twenty patients (22%) underwent total splenectomy with a mean delay of 8 ± 5.6 years after STS (range 0-20.1 years). Eight patients underwent total splenectomy for recurrence of anemia, the others for recurrent discomfort/pain of the remnant (n=7), icterus (n=3), wandering spleen (n=1) and early post-STS hemorrhage (n=1). Requirement for total splenectomy was significantly higher in Group A than Group B (40 of the patients vs. 6% at the last endpoint, p<0.0001).In summary, our results show that in the long-term, STS resulted in a decrease in the hemolytic rate in HS. In children under the age of six years with severe HS, STS decreased the transfusion rate and increased the hemoglobin to a level compatible with normal growth while retaining splenic function. However, half of these patients will require total splenectomy, but at an age where it will be much safer. In milder HS, STS removed discomfort related to the high hemolytic rate whithout exposing the patients to the risks of a total splenectomy. DisclosuresNo relevant conflicts of interest to declare.

PS-090 Hereditary Spherocytosis And Red Cell Indices Mchc, Mcv, Rdw

Background Hereditary spherocytosis (HS) is a common inherited disorder that is characterised by anaemia, jaundice, and splenomegaly. Clinical severity is variable with most patients having a well-compensated hemolytic anaemia. The primary lesion in HS is loss of membrane surface area, leading to reduced deformability due to defects in the membrane proteins ankyrin, band 3, beta spectrin, alpha spectrin, or protein 4.2. Many isolated mutations have been identified in the genes encoding these membrane proteins; common hereditary spherocytosis-associated mutations have not been identified. The classic laboratory features of HS include minimal or no anaemia, reticulocytosis, an increased mean corpuscular haemoglobin concentration (MCHC), spherocytes on the peripheral blood smear, hyperbilirubinemia, and abnormal results on the osmotic fragility test. Aim Of the study is to evaluate the role of MCV, MCHC as a screen test to diagnose spherocytosis Methods In our study are included 60 subjects, 30 children with HS and 30 children-control groups. Our patients with anaemia, jaundice, and splenomegaly are diagnose with HS by incubated osmotic fragility test, performed after incubating RBCs for 18–24 h under sterile conditions at 37°C. Results We found that 25% of pts. have mild HS, 20% moderate HS, 30% moderate to severe HS and 25% severe HS. In peripheral blood smear 7% of pts. had 0–5 spherocites for field, 30% had 5–10 spherocites for field and 63% had 10–15 spherocites for field. 70% of pts. With HS have MCHC > 38%. There are a positive correlation between MCHC and spherocites in peripheral blood smear (r = 0,898, p Conclusion The dedication of hiperdense erythrociyte today is used as a new tool in diagnosing HS. The determination of MCHC constantly growing with other red cell index, MCV 15 obtained from an electronic cell counter usually is enough to suggest for HS. Key Words Spherocytosis, MCHC, anaemia, children.

Synchronous splenectomy during cholecystectomy for hereditary spherocytosis: Is it really necessary?

Background/PurposeExpert guidelines recommend performing synchronous splenectomy in patients with mild hereditary spherocytosis (HS) and symptoms of gallstone disease. This recommendation has not been widely explored in the literature. The aim of this study is to determine if our data support expert opinion and if different practice patterns should exist. MethodsThis is an IRB-approved retrospective study. All HS patients under 18years of age who underwent cholecystectomy for symptomatic gallstones at a single institution between 1981 and 2009 were identified. Patients who underwent cholecystectomy without concurrent splenectomy were reviewed retrospectively for future need for splenectomy and evidence of recurrent gallstone disease. ResultsOf the 32 patients identified, 27 underwent synchronous splenectomy. The remaining 5 patients underwent cholecystectomy without splenectomy and had a mean age of 9.4years. One of the 5 patients eventually required splenectomy for left upper quadrant pain. None of the remaining 4 required hospitalization for symptoms related to hemolysis or hepatobiliary disease. Median follow-up is 15.6years. ConclusionThe need for splenectomy in patients with mild HS and symptomatic cholelithiasis should be assessed on a case by case basis. Our recommendation is to not perform synchronous splenectomy in conjunction with cholecystectomy for these patients if no indication for splenectomy exists.

A new paradigm in the diagnosis of hereditary hemolytic anemia

A new paradigm in the diagnosis of hereditary hemolytic anemia

A Novel ENU-Induced Intronic Mutation of Ank1 Causing Quantitative Defect of Ankyrin-1 Models Hemolytic Hereditary Spherocytosis in Mouse

AbstractAbstract 989The ability of red blood cells (RBCs) to maintain the surface area and deformability is vital for their survival. The maintenance of membrane surface is dependent upon the strong cohesion between the lipid bilayer and the skeletal network, achieved by vertical linkages between transmembrane proteins and spectrin tetramers. Mutations causing functional deficiencies in these proteins have been identified in various hemolytic anemias. Here we reported a mild hereditary spherocytosis (HS) and hemolytic anemia phenotype in mouse, named hema6, induced by N-ethyl-N-nitrosourea (ENU) mutagenesis. Hema6 phenotype is transmitted as a semidominant trait as heterozygous mice are less severely affected than homozygotes. The causal mutation was traced to a single nucleotide transition in the deep intronic region of intron 13 of gene Ank1, encoding the anchorage protein ankyrin-1 in RBC. In vitro minigene assay revealed two abnormally spliced transcripts in addition to wild-type mRNA. The wild-type Ank1 transcript was detected in the homozygous mutant mouse with 30% reduction in expression level compared to that in wild type mouse. The aberrant transcripts presumably encoded a 509 amino acids protein, which lacks beta-spectrin binding domain and C-terminal regulatory region. The truncated protein was not detected by western blotting using currently available antibodies against full-length ankyrin-1 in homozygous hema6 erythrocyte ghosts, whereas the wild-type Ankyrin-1 are present with reduced quantity. Employing biochemical and cell biology assays, we characterized the mechanism by which Ank1hema6 mutation causes hemolytic hereditary spherocytosis in mouse. Hema6 strain provides a novel tool to study ankyrin-1 and its pathogenesis role in HS. Disclosures:No relevant conflicts of interest to declare.

Comments on 'clinical course of 63 children with hereditary spherocytosis: a retrospective study'

Comments on 'clinical course of 63 children with hereditary spherocytosis: a retrospective study'

Hemolytic Crisis as the Initial Presentation of Hereditary Spherocytosis Induced by Parvovirus B19 and Herpes Virus Infection in a Patient with the Thalassemia Trait: A Case Report

To the Editor, Human parvovirus B19 (PV-B19) causes erythema infectiosum, hydrops fetalis, and transient aplastic crisis in immunocompromised patients with chronic hemolytic anemia, arthralgia, and chronic pure red cell aplasia [1]. It may also cause autoimmune hemolytic anemia [2] (which presents as aplastic crisis with reticulocytopenia or increased erythropoiesis with reticulocytosis) [3], autoimmune thrombocytopenia/neutropenia, myelodysplastic syndrome, leukoerythroblastosis, hemophagocytic lymphohistiocytosis, and leukemia [1]. Transient aplastic crisis manifests as sudden exacerbation of anemia in patients with chronic hemolytic anemia, with severe reticulocytopenia lasting 7-10 days in the absence of erythroid precursors due to lysis of the precursors by PV-B19 [4]—the hallmark of which is giant pronormoblasts in the bone marrow [5]. Hemolytic crisis in hereditary spherocytosis (HS) is characterized by more pronounced jaundice than that in stable state of HS due to accelerated hemolysis and probable viral infection [6]. Herein we describe a patient that presented with hemolytic crisis as the initial manifestation of HS during PV-B19 infection—in contrast to the expectation of transient aplastic crisis [7]. This condition was attributed to coexistent herpes virus infection. The patient had a low mean corpuscular hemoglobin (MCH) and mean corpuscular hemoglobin concentration (MCHC) due to the thalassemia trait, which masked spherocytosis. A 15-year-old female was presented for evaluation of anemia. She had a 4-month history of progressive pallor, fatigue, scleral icterus, and dark urine. She has had intractable pallor since nearly 3 years of age Icterus was not noted until four months prior to her presentation to our hospital. Her brother also had pallor without icterus. Physical examination showed pallor, scleral icterus, and hepatosplenomegaly (spleen and liver extended the costal margins by 4 cm and 3 cm, respectively). Laboratory findings were as follows: hemoglobin level: 75 g/L; hematocrit: 24.1%; red blood cell count (RBC):4.11x1012/dL; platelet count: 279 x 109/L; white blood cell count: 7.3 x 109/L: mean corpuscular volume (MCV): 59.5 fL: mean corpuscular hemoglobin concentration MCHC 31.5 g/dl (N: 32.7-35.6 g/dl); mean corpuscular hemoglobin (MCH): 18.5 pg (normal range: 27.2-33.5 pg); red blood cell distribution width: 21% (normal range: 11.8%-14.3%); reticulocyte count: 2.2% (normal range: 0.6%-2.6%). Peripheral blood smear showed anisocytosis, poikilocytosis, hypochromia, and spherocytes. Direct and indirect Coombs tests were negative, and total/indirect bilirubin was 1.68/1.33 mg/dL, haptoglobin was <5.83 mg/dL (normal range: 36-195mg/dL), plasma hemoglobin was 7% (normal: <3), serum lactate dehydrogenase was 254 IU/L, and Hb A2 was 4.37%. Bone marrow aspiration showed erythroid hyperplasia, glucose-6-phosphate dehydrogenase, pyrimidine 5’ nucleotidase, pyruvate kinase, CD 55, and CD 59 evaluation of CD 55, and CD 59 on granulocytes by flow cytometry were normal. Collagen tissue disease markers were negative. Abdominal ultrasonography showed splenomegaly and elevated osmotic fragility. Herpes simplex virus HSV type 1-2 IgM was positive. PV-B19 PCR was 75.3 copies/mL in plasma, based on real-time PCR (normal: negative). The patient was diagnosed as hereditary spherocytosis and thalassemia trait, with coexistent PV-B19 and herpes virus infections.The hemoglobin level was found to have increased spontaneously to 88 g/L and her pallor and scleral icterus were found resolved at the end of in her evaluation made on 46th day of admission At this time blood smear findings and mild indirect hyperbilirubinemia persisted and PV-B19 DNA showed 13 copies and HSV type 1-2 IgM was still positive. Viral serological tests may cross react with each other and we couldn’t perform HSV PCR. But HSV IgG was found to have increased from 6.15 to 113.2 RU/mL on the 46th day of admission and this confirmed that PV-B19 and HSV infected the patient coexistently., The presented patient had mild HS, which manifested first as hemolytic crisis due to herpes virus infection, although infection by other viruses we didn’t test for could not be excluded. Her RBC count was within the lower limits of normal, which is unexpected in a patient with hemolytic anemia and the thalassemia trait. We think the normal RBC and reticulocyte counts, despite erythroid hyperplasia of the bone marrow, were indicative of repressed erythropoiesis due to PV-B19 infection. The presented case highlights the fact that, in the setting of chronic mild hemolytic anemia, patients in hemolytic crisis that have normal reticulocyte and RBC counts should be tested for PV-B19 infection and accompanying other viral infections . Clinicians should be aware that MCH and MCHC are not elevated in HS patients with thalassemia trait. Conflict of Interest Statement None of the authors have any conflicts of interest, including specific financial interests, relationships, and/or affiliations, relevant to the subject matter or materials included.

Guidelines for the diagnosis and management of hereditary spherocytosis – 2011 update

Guidelines on hereditary spherocytosis (HS) published in 2004 (Bolton-Maggs et al, 2004) are here replaced to reflect changes in current opinion on the surgical management, (particularly the indications for concomitant splenectomy with cholecystectomy in children with mild HS, and concomitant cholecystectomy with splenectomy in those with asymptomatic gallstones). Further potential long term hazards of splenectomy are now recognised. Advances have been made in our understanding of the biochemistry of the red cell membrane which underpins the choice of tests. Biochemical assays of membranes proteins and genetic analysis may be indicated (rarely) to diagnose atypical cases. The diagnostic value of the eosin-5-maleimide (EMA) binding test has been validated in a number of studies with understanding of its limitations.

Erythropoiesis versus inflammation in Hereditary Spherocytosis clinical outcome

ObjectivesThis study aimed to evaluate the relationship between erythropoiesis and inflammation, in Hereditary Spherocytosis (HS) clinical outcome. Design and methodsWe studied 26 controls and 82 HS patients presenting mild (n=49) and severer (n=33) HS forms. We evaluated plasma levels of EPO, sTfR, ferritin, iron, folic acid, vitamin B12, TNF-α, IFN-γ, elastase and lactoferrin; leukocyte and reticulocyte counts and RPI were determined. ResultsAll HS patients showed significantly higher EPO, sTfR, reticulocytes and RPI but only mild HS presented normal hemoglobin levels; the positive significant correlations between EPO and sTfR, reticulocytes and RPI observed in mild HS were not observed in severer HS patients. HS patients presented with higher levels of neutrophils, TNF-α, IFN-γ, elastase, lactoferrin and ferritin. ConclusionsOur data show HS as a disease linked to enhanced erythropoiesis that is disturbed in the more severe forms, to which inflammation may contribute, at least in part.

A novel ENU-generated truncation mutation lacking the spectrin-binding and C-terminal regulatory domains of Ank1 models severe hemolytic hereditary spherocytosis

Hereditary spherocytosis (HS) is a heterogeneous group of spontaneously arising and inherited red blood cell disorders ranging from very mild subclinical cases to severe and life-threatening cases, with symptoms linked directly to the severity of the mutation at the molecular level. We investigated a novel mouse model in which the heterozygotes present with the diagnostic hallmarks of mild HS and surviving homozygotes phenocopy severe hemolytic HS. We used N-ethyl-N-nitrosourea mutagenesis to generate random point mutations in the mouse genome and a dominant screen to identify mouse models of human hematopoietic disease. Gene mapping of the HS strain revealed a unique in-frame nonsense mutation arising from a single base transversion in exon 27 of Ank1 (strain designation: Ank1(E924X)). Employing conventional hematopoietic, pathological, biochemical, and cell biology assays, we characterized heterozygous and homozygous Ank1(E924X) mice at the biochemical, cellular, and pathophysiological levels. Although Ank1(E924X/E924X) red blood cell ghosts lack abundant full-length ankyrin-1 isoforms, N-terminal epitope ankyrin-1 antibodies reveal a band consistent with the theoretical size of a truncated mutant ankyrin-1. Using domain-specific antibodies, we further show that this protein lacks both a spectrin-binding domain and a C-terminal regulatory domain. Finally, using antisera that detect C-terminal residues of the products of alternative Ank1 transcripts, we find unique immunoreactive bands not observed in red blood cell ghosts from wild-type or Ank1(E924X) heterozygous mice, including a band similar in size to full-length ankyrin-1. The Ank1(E924X) strain provides a novel tool to study Ank1 and model HS.

Additional erythrocytic and reticulocytic parameters helpful for diagnosis of hereditary spherocytosis: results of a multicentre study

Hereditary spherocytosis (HS) is characterised by weakened vertical linkages between the membrane skeleton and the red blood cell's lipid bilayer, leading to the release of microparticles. All the reference tests suffer from specific limitations. The aim of this study was to develop easy to use diagnostic tool for screening of hereditary spherocytosis based on routinely acquired haematological parameters like percentage of microcytes, percentage of hypochromic cells, reticulocyte counts, and percentage of immature reticulocytes. The levels of haemoglobin, mean cell volume, mean corpuscular haemoglobin concentration, reticulocytes (Ret), immature reticulocytes fraction (IRF), hypochromic erythrocytes (Hypo-He) and microcytic erythrocytes (MicroR) were determined on EDTA samples on Sysmex instruments from a cohort of 45 confirmed SH. The HS group was then compared with haemolytical disorders, microcytic anaemia, healthy individuals and routine samples (n = 1,488). HS is characterised by a high Ret count without an equally elevated IRF. All 45 HS have Ret >80,000/μl and Ret(10(9)/L)/IRF (%) greater than 7.7 (rule 1). Trait and mild HS had a Ret/IRF ratio greater than 19. Moderate and severe HS had increased MicroR and MicroR/Hypo-He (rule 2). Combination of both rules gave predictive positive value and negative predictive value of respectively 75% and 100% (n=1,488), which is much greater than single parameters or existing rules. This simple and fast diagnostic method could be used as an excellent screening tool for HS. It is also valid for mild HS, neonates and ABO incompatibilities and overcomes the lack of sensitivity of electrophoresis in ankyrin deficiencies.

Complementary markers for the clinical severity classification of hereditary spherocytosis in unsplenectomized patients

Hereditary spherocytosis (HS) is usually classified as mild, moderate or severe using conventional features, namely, hemoglobin (Hb) concentration, reticulocyte count and bilirubin levels, which do not always contribute to an adequate clinical classification. The aim of our study was to establish the importance of some laboratory routine parameters, as markers of HS clinical outcome, by studying a control group ( n = 26) and unsplenectomized HS patients ( n = 82) presenting mild, moderate or severe HS. We performed a basic hematologic study and evaluated the reticulocyte count, bilirubin, erythropoietin (EPO) and soluble transferrin receptor (sTfR) levels; the osmotic fragility (OFT) and criohemolysis tests (CHT); the ratios Hb/MCHC (mean cell hemoglobin concentration), Hb/RDW (red cell distribution width) and MCHC/RDW, were calculated. Hb changed significantly in accordance with HS severity, but not reticulocytes or bilirubin. We found that MCHC, RDW, EPO, sTfR, OFT, CHT and the calculated ratios were significantly changed in patients, and, therefore, were valuable as complementary diagnostic tools for HS. Moreover, RDW, Hb/MCHC, Hb/RDW and MCHC/RDW changed significantly with worsening of HS; thus, they are also good markers for the clinical outcome of HS. In conclusion, we propose the use of these routine parameters as useful to complement the analysis of HS severity.

Usefulness of microparticles's Release to Improve Diagnosis of Hereditary Spherocytosis: Results of a Multicentre Study

AbstractAbstract 2041 INTRODUCTION:Hereditary Spherocytosis (HS) is characterized by weakened vertical linkages between the membrane skeleton and the red blood cells (RBC) lipid bilayer, leading to the release of microparticles (MPs). SDS-PAGE and ektacytometry are the two reference tests to establish the diagnosis. However, SDS-PAGE lacks sensitivity to very mild or asymptomatic HS carriers, and a high reticulocyte count might mask a reduction in ankyrin-1 in SDS-PAGE whereas an ankyrin-1 defect represents 40–65% of HS in the USA and Europe. Ektacytometry is only available in specialised laboratories. Consequently, diagnosis of HS is currently based on a combination of clinical and family histories, physical examination and laboratory data, especially RBC indices and sometimes a hypertonic cryohaemolysis test and eosin-5-maleimide binding in flow cytometry (EMA). Besides the release of MPs due to destabilisation of the lipid bilayer, one previous study showed that in HS, but not in AIHA, the surface area loss is already present at the circulating reticulocyte stage. AIMS:The aim of this study is to use the release of MPs at the reticulocyte stage to develop an algorithm on 2 haematological analysers: XE-2100 and XE-5000 (Sysmex) as simple, fast, accurate, sensitive, and specific diagnostic laboratory test for HS. METHODS:HgB, MCV, MCHC, Reticulocytes (Ret), Immature Reticulocytes Fraction (IRF), Hypochromic RBC (Hypo-He), Microcytic RBC (MicroR) and indices were determined using XE-2100 (n= 15) and XE-5000 (n= 30). %HYPO−He and %HYPER−He are specific to the XE−5000 (Figure 1). We first studied the efficiency of discordance between Ret and IRF on the XE-2100 and XE-5000, the MicroR/Hypo-He ratio on XE-5000 and the combination of those 2 rules forming an algorithm, to screen 45 confirmed HS (22 women, 23 men, mean age 13.1, from 1 day to 76 years old). They were classified as severe (n= 6, Hg <8g/dl), moderate (n= 27, Hg 8–12 g/dl) and mild (n= 12, Hg > 12g/dl). We then studied the efficiency of the algorithm to differentiate HS from other haemolytic disorders (n=108), microcytic RBC disorders (n=93), healthy subjects (n=61) and a routine database (n=1230). RESULTS:HS is characterized by a high reticulocyte count (Ret) without an equally elevated Immature Reticulocytes Fraction (IRF) (Figure 1). The algorithm includes a precondition to screen all cases of HS, and a second rule taking into account the degree of anaemia. All 45 HS have reticulocytes > 80 G/l and a Ret (G/l)/IRF (%) ratio higher than 7.7. The second rule is different according to HgB level. All trait and mild cases of HS have a Ret/IRF ratio greater than 19. The efficiency of the algorithm to screen trait and mild HS is very interesting, as the diagnosis of trait and mild HS is currently difficult even by SDS-PAGE. The severity of the disease shown by HgB level is due to the intensity of release of MPs, which is reflected at best by MicroR. Thus, for moderate and severe cases, MicroR and MicroR/Hypo-He were combined. The optimal cut-offs for MicroR and MicroR/Hypo-He in cases of HgB between 8 and 12 g/dl, were 3.5% and 2.5 respectively and if HgB was lower than 8 g/dl, were 3.5% and 2.0 respectively. Interestingly, the algorithm was also valid for the 4 neonates and the 4 cases of ABO incompatibility patients. We also showed that this algorithm is much more performant than single parameters (Ret/IRF index) or the existing rules (reticulocytosis, % microcytes, combination of increased Mean Corpuscular Haemoglobin Concentration (MCHC) with increased red distribution width (RDW), combination of increased RDW with hyperdense cells (Hyperchromic cells), combination of increased MCHC with increased hyperdense cells). The Area Under the Curve (AUC), sensitivity, specificity, predictive positive value (PPV) and negative predictive value (NPV) were respectively 0.997, 100%, 99.3%, 75% and 100% for the HS algorithm. Finally, preliminary data on a small group of patients indicate that HS induced the release of significantly more RBC MPs than healthy subjects, paroxysmal nocturn hemoglobinuria and heparin-induced thrombocytopenia. CONCLUSIONS:We have developed a simple and inexpensive algorithm that could be used as an excellent screening device for the diagnosis of HS. This rapid algorithm also works on mild HS, ABO incompatibilities, neonates and overcomes the lack of sensitivity of SDS-PAGE in ankyrin deficiencies. [Display omitted] Disclosures:No relevant conflicts of interest to declare.

Is cholecystectomy really an indication for concomitant splenectomy in mild hereditary spherocytosis?

Background/aimThe British Committee for Standards in Haematology currently recommends concomitant splenectomy in children with mild hereditary spherocytosis (HS) undergoing cholecystectomy for symptomatic gallstones. However, splenectomy is associated with a risk...