#819 Erythropoietin modulates redox homeostasis and immunomodulatory functions of monocytes in kidney transplant recipients

Abstract Background and Aims Erythropoietin (EPO) has been shown to exert immunomodulatory effects beyond its role in erythropoiesis. Immune cells, particularly monocytes, undergo metabolic reprogramming during activation, which affects their function. This study aims to investigate the immunomodulatory effects of EPO in human kidney transplantation, with a focus on elucidating the effects of EPO on the metabolic state of immune cell subsets using redox homeostasis as a proxy. Method Healthy donor (HD) peripheral blood mononuclear cells (PBMC) and kidney transplant recipients (KTR) PBMC (without EPO treatment in vivo) were treated with EPO in vitro to assess direct EPO effects. Additionally, a cohort of stable KTR (n = 34) with anemia (Hb < 10.0 g/dL) was enrolled and studied before and after 4 weeks of EPO therapy. As a proxy for redox set point, glutathione (GSH) levels were monitored with FJF375, a novel small molecule selective GSH fluorescent sensor developed in the group. Immune phenotyping, cytokine quantification, and bulk RNA sequencing were performed to explore molecular mechanisms of EPO's effects. Results EPO treatment in vitro induced metabolic alterations in monocytes, as detected by our GSH sensor. While no significant changes were observed in T cells or B cells, monocytes significantly decreased GSH baseline levels after EPO treatment in vitro (P = 0.005). KTR treated with EPO in vivo exhibited immune modulation associated with monocyte phenotypes. A significant decrease in GSH levels in monocytes (P = 0.017) was observed, correlating with a marked reduction of TNF-α release (P = 0.0037). RNA sequencing deconvolution analysis further revealed a tendency toward increased IL-4 levels, an anti-inflammatory cytokine associated with M2 macrophages. Conclusion EPO treatment induces cell-specific changes in GSH levels in monocytes, but not in T cells or B cells. EPO may influence immune cell function in KTR by promoting a shift in monocyte phenotyping from a pro-inflammatory M1 profile to an immunomodulatory M2 profile through alterations in redox homeostasis. Future studies are planned to investigate the functional consequences of these metabolic alterations in immune cells and their potential implications for transplantation outcomes.