The pathogenesis of the anemia of prematurity is incompletely understood. Although preterm infants possess storage iron (Fe), it is uncertain that they can mobilize Fe from storage rapidly enough for erythropoiesis to proceed at a rate commensurate with growth. Preterm infants receive protein intakes that are less than the requirements. Hence, protein deficiency may be a causative factor in anemia. The overall hypothesis of this proposal is that availability of Fe and/or protein deficiency limit RBC formation in preterm infants and that optimization of Fe availability or of protein intake will result in clinically relevant enhancement of erythropoiesis and amelioration of anemia which, in turn, may lead to a reduction of the need for RBC transfusions. Of the six Specific Aims, four will be pursued in clinical and two in animal studies. Aim #1 will test the hypothesis that, by raising oral Fe intake from the current routine level to one of two higher levels, enhancement of erythropoiesis and improvement of Fe nutritional status will be achieved, without incurring toxic effects. To determine quantitatively Fe absorption and utilization at these higher Fe intakes, absorption and RBC incorporation of Fe will be determined with use of the non-radioactive isotope, 58Fe. In another subset, we will examine the effect of RBC transfusion on Fe absorption and/or excretion. Evidence of free radical damage will be obtained at all levels of Fe intake. Aim #2 will test the hypothesis that provision of protein intakes that match estimated requirements more closely than current routine feedings will enhance erythropoiesis, leading to improved growth. Aim #3 will test the hypothesis that treatment with r- HuEpo and Fe leads to a shortening of RBC life span and/or to an expansion of RBC mass or blood volume. This aim intends to clarify observation made during a previous study suggesting that such adverse effects do occur during r-HuEPO and Fe treatment. Parental administration of Fe may offer substantial advantages over enteral administration, but is potentially more toxic. Specific Aim #4 will test the hypothesis that slow infusion of Fe to preterm infants leads to greater RBC incorporation of 58Fe than more rapid infusion. Newborn anemic lambs will be used in Aim #5 to test the hypothesis that intravenous Fe stimulates erythropoiesis when given alone or in combination with r-HuEPO. Aim #6 will test the hypothesis that intravenous Fe stimulates erythropoiesis. Aims #4,5 and 6, if successful, will provide the basis for future clinical trials of parenterally administered Fe in human infants.