The signaling cascade induced by the interaction of erythropoietin (EPO) with its receptor (EPO‐R) is a key event of erythropoiesis. We present here data indicating that Fyn, a Src‐family‐kinase, participates in the EPO signaling‐pathway, since Fyn−/− mice exhibit reduced Tyr‐phosphorylation of EPO‐R and decreased STAT5‐activity. The importance of Fyn in erythropoiesis is also supported by the blunted responsiveness of Fyn−/− mice to stress erythropoiesis. Fyn−/− mouse erythroblasts adapt to reactive oxygen species (ROS) by activating the redox‐related‐transcription‐factor Nrf2. However, since Fyn is a physiologic repressor of Nrf2, absence of Fyn resulted in persistent‐activation of Nrf2 and accumulation of nonfunctional proteins. ROS‐induced over‐activation of Jak2‐Akt‐mTOR‐pathway and repression of autophagy with perturbation of lysosomal‐clearance were also noted. Treatment with Rapamycin, a mTOR‐inhibitor and autophagy activator, ameliorates Fyn−/− mouse baseline erythropoiesis and erythropoietic response to oxidative‐stress. These findings identify a novel multimodal action of Fyn in the regulation of normal and stress erythropoiesis.

Thrombopoiesis governs the formation of blood platelets in bone marrow by converting megakaryocytes into long, branched proplatelets on which individual platelets are assembled. The megakaryocyte cytoskeleton responds to multiple microenvironmental cues, including chemical and mechanical stimuli, sustaining the platelet shedding. During the megakaryocyte's life cycle, cytoskeletal networks organize cell shape and content, connect them physically and biochemically to the bone marrow vascular niche, and enable the release of platelets into the bloodstream. While the basic building blocks of the cytoskeleton have been studied extensively, new sets of cytoskeleton regulators have emerged as critical components of the dynamic protein network that supports platelet production. Understanding how the interaction of individual molecules of the cytoskeleton governs megakaryocyte behavior is essential to improve knowledge of platelet biogenesis and develop new therapeutic strategies for inherited thrombocytopenias caused by alterations in the cytoskeletal genes.

Erythropoiesis is a complex multistep process responsible of the production of circulating mature erythrocytes and involved the production of reactive oxygen species (ROS) during erythroid differentiation. Here, we document that Fyn, a Srcfamily-kinase, participates in erythropoietin (EPO) signaling pathway, by the reducing extent of Tyr-phosphorylation of EPO-R and by decreasing STAT5 activity. The importance of Fyn in EPO cascade is also supported by the increased sensitivity of Fyn -/mice to stress erythropoiesis. Fyn -/mouse erythroblasts adapt to the induced stress by the activation of the redox-related-transcription-factor Nrf2. However, the absence of the Nrf2 physiologic repressor Fyn resulted in the persistent activation of Nrf2 and accumulation of non-functional proteins. This is paralleled by ROS induced over-activation of Jak2-Akt-mTOR pathway and repression of autophagy and perturbation of lysosomal-clearance during Fyn -/reticulocyte maturation. Treatment with Rapamycin, a mTOR inhibitor and autophagy activator, ameliorates Fyn -/-

Erythropoiesis is a dynamic and multistep process, resulting in generation of red cells from hematopoietic stem cells. Aging is characterized by increased oxidation and reduced efficiency of cytoprotective mechanisms. Nrf2 is a key transcription factor that participates in acute response to oxidative stress and controls the expression of anti-oxidant and cytoprotective systems. We previously documented activation of Nrf2 during β-thalassemic erythropoiesis. In addition, a previous study reported a mild chronic hemolytic anemia as a result of increased erythrophagocytosis in mice genetically lacking Nrf2 (PNAS, 2004). Here, we show an age-dependent worsening of anemia in Nrf2-/- mice characterized by accelerated senescence of circulating erythrocytes in association with reticulocytopenia, suggesting a perturbation of erythroblast maturation. Indeed, we found ineffective erythropoiesis in 12 months-old Nrf2-/- mice with extramedullary erythropoiesis, increased ROS levels, reduction in the expression of Nrf2 related anti-oxidant proteins and apoptosis of erythroid precursor cells. In agreement, we observed an age dependent sensitivity of Nrf2-/- mice to stress erythropoiesis induced by either phenyhydrazine (PHZ) or doxorubicine. In 12-month-old mice we observed (i) activation of the unfoldeld protein response system (UPR) due to endosomal reticulum stress and (ii) impaired autophagy. The cytoprotective processes were unable to fully counteract oxidation re-directing cells towards apoptosis as supported by the increased activity of caspase-3. To understand the impact of oxidation in this model of accelerated senescence, we treated Nrf2-/- mice with astaxanthin, a powerful anti-oxidant but with low oral bioavailability, by administrating it in the form of PLGA loaded nanoparticles (ATS-NP). Treatment of Nrf2-/- mice with ATS-NP ameliorated the age-dependent anemia and decreased ineffective erythropoiesis through (i) inactivation of UPR system; (ii) improvement of autophagy and (iii) reduction in caspase-3 activity. In future studies, we plan to evaluate the impact of ATS-NP administration in other models of pathologic erythropoiesis. In summary, we propose that Nrf2, a key transcriptional factor plays a key protective role in regulating aged related oxidation and ensures normal erythroid maturation and growth. Disclosures No relevant conflicts of interest to declare.

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