Apotransferrin Injection Leads to More Effective Erythropoiesis in β-Thalassemic Mice.

β-thalassemia is a disease resulting from a β-globin gene mutation which leads to less β-globin, expanded and ineffective erythropoiesis, and anemia. Additionally, mature red blood cells have a shortened survival. Although the degree of anemia varies, from severe transfusion-dependence to only an increase in iron absorption in the gut to maintain hemoglobin levels, all thalassemic patients develop some degree of iron overload. In a previous study using β-thalassemic mice, we were able to induce iron overload using iron dextran injections and demonstrated an increase in hemoglobin with increased reticulocytosis and an expansion of extramedullary erythropoiesis in the liver and spleen. The fact that iron administration reduced anemia in β-thalassemic mice was surprising. Since patients with β-thalassemia have ample iron supply, we hypothesized that part of the anemia in β-thalassemia may result from a maldistribution of iron as a consequence of insufficient circulating transferrin to deliver iron for erythropoiesis. In the present study, we analyzed the effect of intraperitoneal human apotransferrin injections on markers of hematopoiesis and iron metabolism in thalassemic mice. We used three different doses of apotransferrin - 5mg, 10mg, and 30mg - daily, for a 10 day course. Mice with β-thalassemia intermedia (Hbb^th1/th1) were compared with age and gender matched control C57BL/6J mice. Although no increase in hemoglobin was observed, the reticulocyte count decreased after apotransferrin injections (2975±125 x 10⁹ vs. 1636±130 x 10⁹ cells/L, P=2.29 x 10⁻⁵). The efficiency of erythropoiesis, as measured by red cell to reticulocyte ratio, increased after apotransferrin injections (0.029±0.002 vs. 0.007±0.0007, P=0.0002), confirming that more red cells circulate as a result of each maturing erythroid precursor. We were able to demonstrate that apotransferrin is effective in increasing transferrin iron binding capacity (TIBC) (739.5±98.4 vs. 419.1±18.3 mg/dL, P=0.002) without changing the transferrin saturation (23.0±7.1 vs. 34.9±4.2%, P=0.15) in our mice. Apotransferrin injections also resulted in a reduction of iron deposition in the liver (5.49±0.48 vs. 11.08±1.24 mg/g dry weight, P=0.003) and heart (1.25±0.18 vs. 2.26±0.26 mg/g dry weight, P=5.7 x 10⁻⁵) of Hbb^th1/th1 mice without changes in labile plasma iron levels. Using flow cytometry, we demonstrated an increase in erythroid precursors in the bone marrow of Hbb^th1/th1 mice (68.7±1.5 vs. 56.5±3.96% ter119+ precursors, P=0.01) but a decrease in the spleen (41.05±3.16 vs. 60.95±8.3% ter119+ precursors, P=0.03) compared to baseline. Lastly, liver hepcidin expression was progressively suppressed with increasing transferrin dose in our mice. Taken together, this data strongly suggests that exogenous apotransferrin is able to mobilize stored iron for production of erythroid precursors in the bone marrow; this process leads to hepcidin suppression. Diseases of ineffective erythropoiesis, in which expanding erythropoiesis may be limited by the iron delivery system to maintain hemoglobin production, may be a result of insufficient transferrin and relative iron deficiency. The significance of our current findings has potential broad implications for the mobilization of stored iron for use in erythropoiesis in many diseases in which iron overload co-exists with anemia such as β-thalassemia, sideroblastic anemia, and the myelodysplastic syndromes.