Transferrin Receptor 1 Up-Regulation Plays a Role in Iron Accumulation in a Murine Model of Sideroblastic Anemia.

Manganese superoxide dismutase (SOD2) detoxifies superoxide anion radicals generated by mitochondrial respiration (Weisiger and Fridovich, J. Biol. Chem. 1973). While SOD2-deficiency is lethal, SOD2-deficient (SOD2^−/−) hematopoietic stem cells can rescue lethally irradiated wild-type mice. SOD2^−/− hematopoietic chimeras show a persistent hemolytic anemia similar to human sideroblastic anemia (Friedman et al. J. Exp. Med. 2001). SOD2^−/− erythroid progenitor cells have increased mitochondrial mass, and reticulocytes show mitochondrial iron deposition. Mature RBC show abundant siderotic granules, evidence of a defect in iron incorporation into heme, and shortened lifespan. SOD2^−/− progenitors and mature RBC show both enhanced reactive oxygen species production and protein oxidative damage (Friedman et al. Blood 2004; Martin et al. Submitted).

To define early events in the pathogenesis of the SOD2-deficiency anemia and, in particular to identify genes involved in the response of erythroid progenitors to oxidative stress, we compared gene expression of sorted TER-119⁺ CD71⁺ erythroblasts from SOD2^−/−versus wild-type hematopoietic stem cells recipients. Using cDNA microarrays and class comparison analysis, we identified 600 transcripts as significantly discriminant between genotypes. Analysis showed that eleven transcripts encoding different subunits of the mitochondrial oxidative phosphorylation, ATP synthase, and TCA cycle were down-regulated in SOD2^−/− erythroblasts. Previous work showed similar results at the protein level in SOD2^−/− RBC (Friedman et al. Blood 2004) and at the activity level in specific tissues of SOD2^−/− neonates prior to death (Melov et al. PNAS 1999). One interpretation is that SOD2^−/− erythroblasts show metabolic decline. Of interest, a single transcript involved in iron homeostasis, Trfr, was found to be expressed at twice the levels found in wild-type erythroblasts (p<0.0007, parametric p value). Trfr encodes transferrin receptor 1; two-fold up-regulation of transferrin receptor 1 was also shown at the protein level by flow cytometry analysis of SOD2^−/− erythroblasts (p<0.0001, unpaired two-tailed t-test). Transferrin receptor 1 is the cellular gatekeeper for iron uptake whose genetic inactivation induces abnormal erythropoiesis and iron homeostasis (Levy et al. Nat. Genet. 1999). The stability of the Trfr transcript is highly regulated by iron regulatory proteins (IRPs), that are known to be controlled by numerous effectors including reactive oxygen species (Hentze et al. Cell 2004, for review). We focus our current work on investigating, in vitro and in vivo, the role that up-regulation of transferrin receptor 1, likely through oxidative stress-mediated IRPs activity regulation, plays in iron overload in our SOD2-deficiency model.

Taken together, our findings raise the possibility that increased iron delivery may be sufficient to cause sideroblastic anemia or may contribute to a self-reinforcing cycle where oxidative stress favors increased iron, and increased iron results in enhanced oxidative damage.