The In Vitro and In Vivo Antioxidant Effects of Erythropoietin in Thalassemia.

Erythropoietin (EPO), a hormone produced on hypoxia mainly in the kidneys, enhances red blood cell (RBC) production (erythropoiesis) by stimulating the proliferation of erythroid progenitors and precursors in the bone marrow. This effect is mediated by the homodimeric EPO receptor, a class 1 cytokine receptor. Recombinant human EPO is widely used for the treatment of anemia, e.g., in patients on dialysis, patients with myelodysplastic syndrome and oncology patients undergoing chemotherapy. Treatment with EPO was also tried experimentally in patients with thalassemia. In these patients, in spite the state of chronic anemia, the levels of EPO is usually low relative to the degree of anemia. Administration of EPO to thalassemic patients have been shown to increase erythropoiesis and in some cases to elevate fetal hemoglobin production. In addition, EPO has been suggested to have cardio- and neuro-protecting effects and to increase RBC survival in dialysis patients. We have previously shown that RBC and platelets derived from patients with beta-thalassemia and sickle cell disease are under oxidative stress; they have elevated potential to generate reactive oxygen species (ROS) and membrane lipid peroxides, and have lower content of reduced glutathione (GSH) than normal RBC. This oxidative stress resulted in high exposure of phosphatidylserine (PS) that is considered a major factor in shortening the life span of thalassemic RBC, and in the tendency of platelets to undergo activation and thus contributes to the high incidence of thromboembolic complications in thalassemia. In the present study, we investigated the in vitro and in vivo effects of EPO as an antioxidant on RBC and platelets from beta-thalassemic patients and mice. Using flow-cytometry methodology, we showed that in vitro treatment of blood cells from beta-thalassemic patients with 1–4 U/ml of EPO for 2 hours increased the GSH content of RBC (2.1-fold) and platelets (1.7-fold) and reduced their ROS (1.5-fold), membrane lipid peroxidation and externalization of PS. Intraperitoneal injection of EPO to heterozygotes (Hbb^th3/+) beta-thalassemic mice (3,000U/kg) significantly reduced ROS and increased GSH in their RBC within 3 hours. The in vitro effects of EPO on oxidative stress resulted in reduced sensitivity of thalassemic RBC to undergo hemolysis and phagocytosis by macrophages, and reduced tendency of platelets to undergo activation, as reflected by fewer platelets carrying external PS. Our results suggest that in addition to its effect on erythropoiesis and fetal hemoglobin production, EPO might alleviate symptoms in thalassemia and other hemolytic anemias as a potent antioxidant.