Efficacy of cytochemical tests in gene analysis of hereditary spherocytosis: a case study of six patients with different disease subtypes

Abstract

Objectives: In this study, clinical and biochemical methods were utilized to predict the final diagnosis of hereditary spherocytosis (HS), correlate the diagnosis with splenectomy, and examine the usefulness of this approach. Methods: We biochemically and cytochemically analysed erythrocyte membrane proteins before making a final HS diagnosis based on gene analysis to compare diagnostic approaches. The clinical features of six patients with various subtypes of HS and symptoms were observed by blood analysis using eosin-5′-maleimide staining, biochemical analysis using sodium dodecyl sulphate–polyacrylamide gel electrophoresis with western blotting, and mass spectrometry. Finally, diagnostic membrane gene analysis was performed. Results: Five of the six patients showed mild to moderate or severe anaemia, and the other patient was non-anaemic; all six patients showed faint eosin-5′-maleimide staining. In western blotting of erythrocyte membrane proteins, all six patients (three with β-spectrin, two with ankyrin, and one with SLC4A1 anomalies) showed low-molecular-weight peptide fragments, which were confirmed by mass spectrometry in the region corresponding to the band 3 protein. The two patients with an ankyrin gene anomaly exhibited severe anaemia, and two patients with simultaneous SLC4A1, SPTB, and UGT1A1 anomalies exhibited mild anaemia and hyperbilirubinemia. Discussion: We determined the relationship among clinical features, cytochemical parameters, and gene anomalies in six patients with newly diagnosed HS while referring to previously published cases. Conclusion: These findings reveal a close relationship between clinical features and membrane characteristics in HS, which can facilitate diagnosis and inform treatment.