Increased retention of functional mitochondria in mature sickle red blood cells is associated with increased sickling tendency, hemolysis and oxidative stress.

Abstract

Abnormal retention of mitochondria into mature red blood cells (RBCs) has been recently reported in sickle cell anemia (SCA) but their functionality and their role in the pathophysiology of SCA remain unknown. The presence of mitochondria within RBCs was determined by flow cytometry in 61 SCA patients and 10 healthy donors. Patients were classified according to the percentage of mature RBCs with mitochondria contained in the whole RBC population: low (0-4%), moderate (>4 and 8%). RBC rheological, hematological, RBC senescence and oxidative stress markers were compared between the three groups. RBC senescence and oxidative stress markers were also compared between mature RBC containing mitochondria and those without. The functionality of residual mitochondria in sickle RBCs was measured by high-resolution respirometry assay and showed detectable mitochondrial oxygen consumption in sickle mature RBCs but not in healthy RBCs. Increased levels of mitochondrial reactive oxygen species (ROS) were observed in mature sickle RBCs when incubated with Antimycin A vs without. In addition, mature RBCs retaining mitochondria exhibited greater levels of ROS compared to RBCs without mitochondria, as well as greater Ca2+, lower CD47 and greater phosphatidylserine exposure. Hematocrit and RBC deformability were lower, and the propensity of RBC to sickle under deoxygenation was higher, in the SCA group with a high percentage of mitochondria retention in mature RBCs. This study showed the presence of functional mitochondria in mature sickle RBCs, which could favor RBC sickling and accelerate RBC senescence, leading to increased cellular fragility and hemolysis.