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A prospective study to assess the predictive value for hereditary spherocytosis using five laboratory tests (cryohemolysis test, eosin-5′-maleimide flow cytometry, osmotic fragility test, autohemolysis test, and SDS-PAGE) on 50 hereditary spherocytosis families in Argentina

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Abstract

This prospective study was carried out to assess the usefulness of five laboratory tests in the diagnosis of hereditary spherocytosis (HS), based on the correlation of erythrocyte membrane protein defects with clinical and laboratory features, and also to determine the membrane protein deficiencies detected in Argentina. Of 116 patients and their family members tested, 62 of them were diagnosed to have HS. The specificity of cryohemolysis (CH) test was 95.2%, and its cut-off value to distinguish HS from normal was 2.8%. For flow cytometry, cut-off points of 17% for mean channel fluorescence (MCF) decrease and 14% coefficient of variation (CV) increase showed 95.9% and 92.2% specificity, respectively. Both tests showed the highest percentages of positive results for diagnosis. Either CH or flow cytometry was positive in 93.5% of patients. In eight patients, flow cytometry was positive only through CV increase. Protein defects were detected in 72.3% of patients; ankyrin and spectrin were the most frequently found deficiencies. The CV of the fluorescence showed significantly higher increases in moderate and severe anemia than in mild anemia (p = 0.003). Severity of anemia showed no other correlation with tests results, type of deficient protein, inheritance pattern, or neonatal jaundice. CH and flow cytometry are easy methods with the highest diagnostic accuracy. Simultaneous reading of mean channel fluorescence (MCF) decrease and CV increase improve diagnostic usefulness of flow cytometry. This test seems to be a reliable predictor of severity. The type of detected protein deficiency has no predictive value for outcome. Predominant ankyrin and spectrin deficiencies agree with reports from other Latin American countries.

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References

  1. Grace RF, Lux SE (2009) Disorders of the red cell membrane. In: Orkin SH, Nathan DG, Ginsburg D, Fisher DE, Lux SE (eds) Hematology of infancy and childhood. Saunders, Philadelphia, pp 659–838

    Google Scholar 

  2. Hassoun H, Palek J (1996) Hereditary spherocytosis: a review of the clinical and molecular aspects of the disease. Blood Rev 10:129–147

    Article  PubMed  CAS  Google Scholar 

  3. Bolton-Maggs PHB, Stevens RF, Dodd NJ, Lamont G, Tittensor P (2004) On behalf of the General Haematology Task Force of the British Committee for Standards in Haematology. Guidelines for the diagnosis and management of hereditary spherocytosis. Br J Haematol 126:455–474

    Article  PubMed  CAS  Google Scholar 

  4. Gallagher PG (2005) Red cell membrane disorders. Hematology. Am Soc Hematol Educ Program

  5. Mariani M, Barcellini W, Vercellati C, Marcello AP, Fermo E, Pedotti P et al (2008) Clinical and hematologic features of 300 patients affected by hereditary spherocytosis grouped according to the type of the membrane protein defect. Haematologica 93:1310–1317

    Article  PubMed  CAS  Google Scholar 

  6. Iolascon A, Avvisati RA (2008) Genotype/phenotype correlation in hereditary spherocytosis. Haematologica 93:1283–1288

    Article  PubMed  CAS  Google Scholar 

  7. King MJ, Telfer P, MacKinnon H, Langabeer L, McMahon C, Darbyshire P et al (2008) Using the eosin-5-maleimide binding test in the differential diagnosis of hereditary spherocytosis and hereditary pyropoikilocytosis. Cytometry Part B Clin Cytom 74B:244–250

    Article  CAS  Google Scholar 

  8. Dacie JV, Lewis SM, Luzzato L (1991) Investigation of the hereditary haemolytic anaemias: membrane and enzyme abnormalities. In: Dacie JV, Lewis SM (eds) Practical haematology. Churchill Livingstone, Edinburgh, pp 195–225

    Google Scholar 

  9. Korones D, Pearson HA (1989) Normal erythrocyte osmotic fragility in hereditary spherocytosis. J Pediatr 114:264–266

    Article  PubMed  CAS  Google Scholar 

  10. Perrotta S, Gallagher PG, Mohandas N (2008) Hereditary spherocytosis. Lancet 372:1411–1426

    Article  PubMed  CAS  Google Scholar 

  11. King MJ, Behrens J, Rogers C, Flynn C, Greenwood D, Chambers K (2000) Rapid flow cytometric test for the diagnosis of membrane cytoskeleton-associated haemolytic anaemia. Br J Haematol 111:924–933

    Article  PubMed  CAS  Google Scholar 

  12. Streichman S, Gesheidt Y, Tatarsky I (1990) Hypertonic cryohemolysis: a diagnostic test for hereditary spherocytosis. Am J Hematol 35:104–109

    Article  PubMed  CAS  Google Scholar 

  13. Streichman S, Gescheidt Y (1998) Cryohemolysis for the detection of hereditary spherocytosis: correlation studies with osmotic fragility and autohemolysis. Am J Hematol 58:206–212

    Article  PubMed  CAS  Google Scholar 

  14. Stoya G, Gruhn B, Vogelsang H, Baumann E, Linss W (2006) Flow cytometry as a diagnostic tool for hereditary spherocytosis. Acta Haematol 116:186–191

    Article  PubMed  Google Scholar 

  15. Tachavanich K, Tanphaichitr VS, Utto W, Viprakasit V (2009) Rapid flow cytometric test using eosin-5-maleimide for diagnosis of red blood cell membrane disorders. Southeast Asian J Trop Med Public Health 40:570–574

    PubMed  CAS  Google Scholar 

  16. Parpart AK, Lorenz PB, Papart ER, Gregg JR, Chase AM (1947) The osmotic resistance (fragility) of human red cells. J Clin Invest 26:636–640

    Article  CAS  Google Scholar 

  17. Selwyn JG, Dacie JV (1954) Autohemolysis and other changes resulting from the incubation in vitro of red cells from patients with congenital hemolytic anemia. Blood 9:414–438

    PubMed  CAS  Google Scholar 

  18. Vittori D, Garbossa G, Lafourcade C, Pérez G, Nesse A (2002) Human erytroid cells are affected by aluminium. Alteration of membrane band 3 protein. Biochem Biophys Acta 1558:142–150

    Article  PubMed  CAS  Google Scholar 

  19. Lowry OH, Roseborough NJ, Farr AL, Randall RL (1951) Protein measurements with Folin reagent. J Biol Chem 193:267–275

    Google Scholar 

  20. Streichman S, Kahana E, Tatarsky I (1985) Hypertonic cryohemolysis of pathologic red blood cells. Am J Hematol 20:373–381

    Article  PubMed  CAS  Google Scholar 

  21. Romero RR, Poo JL, Robles JA, Uriostegui A, Vargas F, Majluf-Cruz A (1997) Usefulness of cryohemolysis test in the diagnosis of hereditary spherocytosis. Arch Med Res 28:247–251

    PubMed  CAS  Google Scholar 

  22. Iglauer A, Reinhardt D, Schröter W, Pekrun A (1999) Cryohemolysis test as a diagnostic tool for hereditary spherocytosis. Ann Hematol 78:555–557

    Article  PubMed  CAS  Google Scholar 

  23. Girodon F, Garçon L, Bergoin E, Largier M, Delaunay J, Feneant-Thibault M et al (2008) Usefulness of the eosin-5′-maleimide cytometric method as a first-line screening test for the diagnosis of hereditary spherocytosis: comparison with ektacytometry and protein electrophoresis. Br J Haematol 140:468–470

    Article  PubMed  CAS  Google Scholar 

  24. Kar R, Rao S, Srinivas UM, Mishra P, Pati HP (2009) Clinico-hematological profile of hereditary spherocytosis: experience from a tertiary care center in North India. Hematology 14:164–167

    Article  PubMed  Google Scholar 

  25. Kedar PS, Colah RB, Kullkarni S, Ghosh K, Mohanty D (2003) Experience with eosin-5-maleimide as a diagnostic tool for red cell membrane cytoskeleton disorders. Clin Lab Haematol 25:373–376

    Article  PubMed  CAS  Google Scholar 

  26. Miraglia del Giudice F, Iolascon A, Pinto L, Nobili B, Perrotta S (1994) Erythrocyte membrana protein alterations underlying clinical heterogeneity in hereditary spherocytosis. Br J Haematol 88:52–55

    Article  PubMed  CAS  Google Scholar 

  27. Yawata Y, Kanzaki A, Yawata A, Doerfler W, Ozcan R, Eber SW (2000) Characteristic features of the genotype and phenotype of hereditary spherocytosis in the Japanese population. Int J Hematol 7:118–135

    Google Scholar 

  28. Lee YK, Cho HI, Park SS, Lee YJ, Ra E, Chang YH et al (2000) Abnormalities of erythrocyte membrane proteins in Korean patients with hereditary spherocytosis. J Korean Med Sci 15:284–288

    PubMed  CAS  Google Scholar 

  29. Sánchez-López JY, Camacho AL, Magaña MT, Ibarra B, Perea FJ (2003) Red cell membrane protein deficiencias in Mexican patients with hereditary spherocytosis. Blood Cells Mol Dis 31:357–359

    Article  PubMed  Google Scholar 

  30. Costa FF, Agre P, Watkins PC, Winkelmann JC, Tang TK, John KM et al (1990) Linkage of dominant hereditary spherocytosis to the gene for the erythrocyte membrane-skeleton protein ankyrin. N Engl J Med 323:1046–1050

    Article  PubMed  CAS  Google Scholar 

  31. Saad ST, Costa FF, Vicentin DL, Salles TS, Pranke PH (1994) Red cell membrane protein abnormalities in hereditary spherocytosis in Brazil. Br J Haematol 88:295–299

    Article  PubMed  CAS  Google Scholar 

  32. Lanciotti M, Perutelli P, Valetto A, Di Martino D, Mori PG (1997) Ankyrin deficiency is the most common defect in dominant and non dominant hereditary spherocytosis. Haematologica 82:460–462

    PubMed  CAS  Google Scholar 

  33. Dhermy D, Bournier O, Bougeois M, Grandchamp B (1999) The red blood cell band 3 variant (band 3 Bicetrel: R 490 C) association with dominant hereditary spherocytosis causes defective membrane targeting of the molecule and a dominant negative effect. Mol Membr Biol 16:305–312

    Article  PubMed  CAS  Google Scholar 

  34. Premetis E, Stamoulakatou A, Loukopoulos D (1999) Hereditary spherocytosis in Greece: collective data on a large number of patients. Hematology 4:361–366

    PubMed  Google Scholar 

  35. Ricard MP, Gilsanz F, Millan I (2000) Erythroid membrane protein defects in hereditary spherocytosis. A study of 62 Spanish cases. Haematologica 85:994–995

    PubMed  CAS  Google Scholar 

  36. Cynober T, Mohandas N, Tchernia G (1996) Red cell abnormalities in hereditary spherocytosis: relevance to diagnosis and understanding of the variable expression of clinical severity. J Lab Clin Med 12:259–269

    Article  Google Scholar 

  37. Rocha S, Rebelo I, Costa E, Catarino C, Belo L, Castro EBM et al (2005) Protein deficiency balance as a predictor of clinical outcome in hereditary spherocytosis. Eur J Haematol 74:374–380

    Article  PubMed  CAS  Google Scholar 

  38. Shi Z, Afzal V, Coller B, Patel D, Chasis JA, Parra M et al (1999) Protein 4.1R-deficient mice are viable but have erythroid membrane skeleton abnormalities. J Clin Invest 103:331–340

    Article  PubMed  CAS  Google Scholar 

  39. Delaunay J (2007) The molecular basis of hereditary red cell membrane disorders. Blood 21:1–20

    Article  CAS  Google Scholar 

  40. Rivera A, De Franceschi L, Peters LL, Gascard P, Mohandas N, Brugnara C (2006) Effect of complete protein 4.1R deficiency on ion transport properties of murine erythrocytes. Am J Physiol Cell Physiol 291:C880–C886

    Article  PubMed  CAS  Google Scholar 

  41. Goodman SR, Shiffer KA, Casoria LA, Eyster ME (1982) Identification of the molecular defect in the erythrocyte membrane skeleton of some kindreds with hereditary spherocytosis. Blood 60:772–784

    PubMed  CAS  Google Scholar 

  42. Becker PS, Morrow JS, Lux SE (1987) Abnormal oxidant sensitivity and beta-chain structure of spectrin in hereditary spherocytosis associated with defective spectrin-protein 4.1 binding. J Clin Invest 80:557–565

    Article  PubMed  CAS  Google Scholar 

  43. Agre P, Asimos A, Casella JF, McMillan C (1986) Inheritance pattern and clinical response to splenectomy as a reflection of erythrocyte spectrin deficiency in hereditary spherocytosis. N Engl J Med 315:1579–1583

    Article  PubMed  CAS  Google Scholar 

  44. Jarolim P, Murray JL, Rubin HL, Taylor WM, Prchal JT, Ballas SK et al (1996) Characterization of 13 novel band 3 gene defects in hereditary spherocytosis with band 3 deficiency. Blood 88:4366–4374

    PubMed  CAS  Google Scholar 

  45. Nakanishi H, Kanzaki A, Yawata A, Yamada O, Yawata Y (2001) Ankyrin gene mutations in Japanese patients with hereditary spherocytosis. Int J Hematol 73:54–63

    Article  PubMed  CAS  Google Scholar 

  46. Morton NE, MacKinney AA, Kosower N, Schilling RF, Gray MP (1963) Genetics of spherocytosis. Am J Hum Genet 14:170–184

    Google Scholar 

  47. Eber SW, Armbrust R, Schröter W (1990) Variable clinical severity of hereditary spherocytosis: relation to erythrocyte spectrin concentration, osmotic fragility and autohemolysis. J Pediatr 117:409–416

    Article  PubMed  CAS  Google Scholar 

  48. Gallagher PG, Forget BG (2001) Hereditary spherocytosis, elliptocytosis and related disorders. In: Beutler E, Lichtman MA, Coller BS, Kipps TJ, Seligsohn U (eds) Williams hematology. McGraw-Hill, New York, pp 503–518

    Google Scholar 

  49. Eber SW, Gonzalez JM, Dornwell M et al (1996) Ankyrin-1 mutations are a major cause of dominant and recessive hereditary spherocytosis. Nat Genet 13:214–218

    Article  PubMed  CAS  Google Scholar 

  50. Gallagher PJ, Ferreira JD, Costa FF, Saad ST, Forget BG (2000) A recurrent frameshift mutation of the ankyrin gene associated with severe hereditary spherocytosis. Br J Haematol 111:1190–1193

    Article  PubMed  CAS  Google Scholar 

  51. Miraglia del Giudice E, Nobili B, Francese M, D’Urso L, Iolascon A, Eber S et al (2001) Clinical and molecular evaluation of non-dominant hereditary spherocytosis. Br J Haematol 112:42–47

    Article  PubMed  CAS  Google Scholar 

  52. Pinto L, Iolascon A, MiragliadelGiudice E, Matarese MRS, Nobili B, Perrotta S (1995) The Italian pediatric survey on hereditary spherocytosis. Int J Pediatr Hematol Oncol 2:43–47

    Google Scholar 

  53. Aramburu Arriaga N, Fernández Cuesta MA, Martínez Gonzalez MJ, Astigarraga Aguirre I, Fernandez-Teijeiro Alvarez A, Navajas Gutierrez A et al (2000) Hereditary spherocytosis in neonates. Review of our casuistics. Ann Esp Pediatr 52:569–572

    CAS  Google Scholar 

  54. Delhommeau F, Cynober T, Schischmanoff PO, Rohrlich P, Delaunay N, Mohandas N et al (2000) Natural history of hereditary spherocytosis during the first year of life. Blood 95:393–397

    PubMed  CAS  Google Scholar 

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Acknowledgment

The authors would like to thank Viviana Sanchez for her technical assistance in the preparation of the manuscript and Laura Marangunich for statistical advice.

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Authors and Affiliations

  1. División Hematología Clínica, Departamento de Medicina, Hospital Nacional Alejandro Posadas, Buenos Aires, Argentina

    Renée L. Crisp & Graciela Alfonso

  2. Laboratorio de Análisis Biológicos, Departamento de Química Biológica, Facultad de Ciencias Exactas y Naturales, Universidad de Buenos Aires, Buenos Aires, Argentina

    Renée L. Crisp, Daiana Vota, Maria E. Chamorro, Daniela Vittori & Alcira Nesse

  3. Consultorios de Hematología Infantil, Buenos Aires, Argentina

    Renée L. Crisp, Gabriel A. Schvartzman & Hugo Donato

  4. Laboratorio de Citometría, Departamento de Diagnóstico, Hospital Nacional Alejandro Posadas, Buenos Aires, Argentina

    Liliana Solari, Daniel Gammella & Belen Venegas

  5. División Hematología y Oncología Pediátrica, Departamento de Pediatría, Hospital Nacional Alejandro Posadas, Buenos Aires, Argentina

    Eliana García & Cecilia Riccheri

  6. Servicio de Medicina Transfusional, Departamento de Diagnóstico, Hospital Nacional Alejandro Posadas, Buenos Aires, Argentina

    Gabriela Miguez & Sergio Caldarola

  7. División Hematología Clínica, Departamento de Medicina, Hospital Nacional Alejandro Posadas, Fernandez de Enciso 4602, 1419, Buenos Aires, Argentina

    Renée L. Crisp

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  1. Renée L. Crisp
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Correspondence to Renée L. Crisp.

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Crisp, R.L., Solari, L., Vota, D. et al. A prospective study to assess the predictive value for hereditary spherocytosis using five laboratory tests (cryohemolysis test, eosin-5′-maleimide flow cytometry, osmotic fragility test, autohemolysis test, and SDS-PAGE) on 50 hereditary spherocytosis families in Argentina. Ann Hematol 90, 625–634 (2011). https://doi.org/10.1007/s00277-010-1112-0

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