The in vitro red blood cell microvesiculation exerts procoagulant activity of blood cell storage in Southeast Asian ovalocytosis

Abstract

Southeast Asian ovalocytosis (SAO) is characterized by the misfolding of band 3 protein in red blood cells (RBC). The abnormal structure of the band 3 protein results in dysmorphic RBC and related functions. Previous data showed that in vitro storage under hypothermic conditions alters band 3 protein structure and function. Microvesiculation includes shedding of RBC membranes, called RBC-derived microparticles/extracellular vesicles (RMP/EVs), and storage lesions. Unfortunately, there is no evidence of RBC microvesiculation under in vitro storage conditions in heterozygous SAO individuals. This study determined the generation of REVs and procoagulant activity during the storage of SAO blood samples in southern Thailand. Venous blood was collected from eight SAO and seven healthy individuals, preserved in citrate phosphate dextrose-adenine 1 (CPDA-1) at 4 °C for 35 days. The absolute numbers of REVs and PS-expressing RBCs were analyzed using flow cytometry. The procoagulant activity of the produced extracellular vesicles was determined by a clotting time assay. The results showed a significant increase in the number of REVs and PS-expressing RBCs in the SAO blood samples. Significantly correlated PS externalization and procoagulant activity were observed in the SAO blood samples. These lines of evidence indicate that the abnormality of the Band 3 protein is possibly involved in aberrant microvesiculation, exerting procoagulant activity in vitro. Increased pools of REV production and abnormal storage lesions in SAO blood samples should be a concern. Notably, the mechanisms underlying membrane vesiculation depend on the extent of blood cell storage under hypothermic conditions.