Erythroferrone Regulates Hepcidin Expression Independently of Matriptase 2

Introduction: The iron-regulatory hormone hepcidin regulates the body iron stores and its expression is repressed when erythropoietic activity intensifies to meet the iron requirements for erythropoiesis (e.g. during anemia). Under the influence of erythropoietin (EPO), the hormone erythroferrone (ERFE) is secreted by erythroid precursors in the bone marrow and the spleen, and suppresses hepcidin synthesis to facilitate the recovery from anemia. However, the mechanism by which ERFE suppresses hepcidin is unknown. In contrast with forms of anemia in which hepcidin is suppressed, patients with mutations in transmembrane serine protease 6 (TMPRSS6) have iron-refractory iron deficiency anemia (IRIDA) but increased hepcidin production despite a severe anemia and elevated EPO levels. Recently, it has been suggested that matriptase-2 activity facilitates ERFE-mediated suppression of hepcidin. We therefore investigated the potential crosstalk between ERFE and Matriptase 2.

Methods: We first measured serum ERFE concentration in Tmprss6^-/- mice. To assess the contribution of ERFE to the phenotype of Tmprss6^-/-mice, we next generated Tmprss6^-/-mice with disrupted Erfe (Erfe^+/- Tmprss6^-/-; Erfe^-/- Tmprss6^-/- and Erfe^+/+ Tmprss6^-/-). To determine whether ERFE requires TMPRSS6 to regulate hepcidin production, we treated freshly isolated hepatocytes from wild-type (WT) or Tmprss6^-/- mice with conditioned medium from cells expressing recombinant ERFE or not.

Results: While wild-type mice have undetectable plasma ERFE (below the 500 pg/ml limit of detection), plasma ERFE concentration was elevated in Tmprss6^-/- to levels comparable to those of WT animals 24 hours after phlebotomy (~3 ng/ml) but was lower than ERFE levels in thalassemic mice (~10 ng/ml). Ablation of Erfe in Tmprss6^-/- mice did not result in any change in hematological parameters, hepcidin expression and iron levels compared to Tmprss6^-/- animals at 6 weeks of age. However, treatment of WT and Tmprss6^-/-hepatocytes with ERFE resulted in a comparable suppression of hepcidin mRNA expression.

Conclusion: Although matriptase-2 may dampen the BMP signaling under the influence of EPO, it is not part of the ERFE signaling pathway.