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.

[Hereditary stomatocytoses--diagnostic problems and their molecular basis]. / Dziedziczne stomatocytozy--problemy diagnostyczne i podstawy molekularne.

Hereditary stomatocytosis (HSt) is a group of haemolytic anaemias in which the common symptom is an increased permeability of the red cell membrane for monovalent cations. HSt is diagnosed really seldom and the difficulties in diagnosing are connected to the fact that the clinical presentation of individual subtypes of HSt is very diverse. Many cases are characterised by unique phenotypes. Nevertheless, the number of diagnosed HSt cases is increasing each year. The aim of this review was the presentation of current information and an attempt to systematize it, what might be helpful in clinical diagnostic of the new cases of this anaemia. The most frequent mistake is to classify a case of HSt as the most common haemolytic anaemia--hereditary spherocytosis (HS), in which to improve patient condition a splenectomy is often recommended. Most cases of HSt no positive response to splenectomy and often thromboembolic complications are observed. It is interesting that commonly present in blood film stomatocytes and in many cases absent or severely reduced stomatin in HSt red cell membrane are not correlated with nucleotide sequence changes of the gene encoding stomatin. Many diagnosed cases are related to mutations in SLC4A1 and RHAG genes. Extensive research carried out on HSt in the entire world will certainly permit to know the molecular basis of the disease, the diversify of its subtypes and to estimate the real incidence of HSt.

Clinical and hematologic features of 300 patients affected by hereditary spherocytosis grouped according to the type of the membrane protein defect.

BACKGROUND: Hereditary spherocytosis is a very heterogeneous form of hemolytic anemia. The aim of this study was to relate the type of molecular defect with clinical and hematologic features and response to splenectomy using information from a large database of patients. DESIGN AND METHODS: Data from 300 consecutive patients with hereditary spherocytosis, grouped according to the results of sodium dodecyl sulphate-polyacrylamide gel electrophoresis, were analyzed and the sensitivity of red cell osmotic fragility tests was compared in various subsets of patients. RESULTS: Band 3 and spectrin deficiencies were the most common protein abnormalities (54% and 31%, respectively); 11% of cases were not classified by the electrophoretic analysis. Spectrin deficiency was more frequently diagnosed in childhood and band 3 deficiency in adulthood. Hemoglobin concentration was slightly lower, spherocyte number and hemolysis markers higher in spectrin deficiency than in band 3 deficiency. The sensitivity of the osmotic fragility tests ranged from 48% to 95%, and was independent of the type and amount of the membrane defect. The association of the acidified glycerol lysis test and the NaCl test on incubated blood reached a sensitivity of 99%. Splenectomy corrected the anemia in patients with all subtypes of hereditary spherocytosis although spectrin-deficient patients still showed increased reticulocyte numbers and levels of unconjugated bilirubin. Splenectomy allowed the identification of the membrane defect in all the previously unclassified patients, most of whom had spectrin and/or ankyrin deficiency. CONCLUSIONS: The definition of the red cell membrane defect in hereditary spherocytosis has no major clinical implications, but may be useful for a differential diagnosis from other hematologic disorders that mimic this hemolytic anemia.

Variable clinical severity of hereditary spherocytosis: relation to erythrocytic spectrin concentration, osmotic fragility, and autohemolysis.

To determine whether stratifying hereditary spherocytosis by degree of severity could provide guidelines regarding which patients would benefit from splenectomy, we evaluated the clinical characteristics of 80 patients (63 children) and 27 healthy relatives. In addition to routine hematologic determinations, osmotic fragility, autohemolysis, erythrocyte spectrin content, and erythrocyte membrane lipid phosphorus were measured and correlated with the disease severity. Four categories were identified: (1) spherocytosis as a trait in symptom-free relatives of patients with recessively inherited disease; (2) mild and (3) moderate spherocytosis, largely observed in patients with dominantly inherited disease; and (4) severe spherocytosis, observed in only two patients, who were characterized by recessive inheritance and transfusion dependence. By the identification of carriers, a recessive mode of inheritance could be demonstrated in 20% of the families with spherocytosis. The erythrocyte spectrin concentration was normal in carriers and patients with mild spherocytosis, and was significantly reduced in the moderate and severe states of the disease. This difference was not accounted for by reduced membrane area of the cells, as measured by the phospholipid concentration per cell. We conclude that patients with mild spherocytosis usually do not require splenectomy during childhood and adolescence; patients with moderate or severe disease should have splenectomy. Patients with severe spherocytosis have a partial response to splenectomy but a considerable degree of increased hemolysis persists. Most patients with less than 80% of normal spectrin content require splenectomy.

Characterization of 13 novel band 3 gene defects in hereditary spherocytosis with band 3 deficiency.

Hereditary spherocytosis (HS) is a common hemolytic anemia of variable clinical expression. Pathogenesis of HS has been associated with defects of several red cell membrane proteins including erythroid band 3. We have studied erythrocyte membrane proteins in 166 families with autosomal dominant HS. We have detected relative deficiency of band 3 in 38 kindred (23%). Band 3 deficiency was invariably associated with mild autosomal dominant spherocytosis and with the presence of pincered red cells in the peripheral blood smears of unsplenectomized patients. We hypothesized that this phenotype is caused by band 3 gene defects. Therefore, we screened band 3 DNA from these 38 kindred for single strand conformational polymorphisms (SSCP). In addition to five mutations detected previously by SSCP screening of cDNA, we detected 13 new band 3 gene mutations in 14 kindred coinherited with HS. These novel mutations consisted of two distinct subsets. The first subset included seven nonsense and frameshift mutations that were all associated with the absence of the mutant mRNA allele from reticulocyte RNA, implicating decreased production and/or stability of mutant mRNA as the cause of decreased band 3 synthesis. The second group included five substitutions of highly conserved amino acids and one in-frame deletion. These six mutations were associated with the presence of comparable levels of normal and mutant band 3 mRNA. We suggest that these mutations interfere with band 3 biosynthesis leading thus to the decreased accumulation of the mutant band 3 allele in the plasma membrane.