Sunday, May 11, 2025
5/11/2025
The role of ferroptosis in red cell aging in vivo and in vitro - PROJECT SUMMARY Red blood cells (RBCs) are a perfect model to study oxidant stress. RBCs are the most abundant cell in the body (84% of total human cells) and play an essential role in oxygen transport and thus in the regulation of all oxygen-dependent metabolic processes. To facilitate this task, RBCs are loaded with hemoglobin (Hb) and iron. Indeed, 66% of total bodily iron is in RBCs. As a result, the mature RBC faces significant oxidant stress deriving from iron-dependent Fenton and Haber-Weiss redox chemistry. The lack of nuclei and organelles prevents RBCs from synthesizing new proteins to replace oxidatively damaged components during the average lifespan of 120 days in circulation. Every day over 200 billion RBCs are removed from the bloodstream and de novo generated via erythropoiesis, a process that relies on the uptake of circulating iron in the ferric (oxidized) state, and its reduction to the ferrous state. This process is catalyzed by the ferrireductase STEAP3 – a transcriptionally-regulated target of tumor protein p53. Both p53 and STEAP3 are critical to erythropoiesis and polymorphic in humans, whereby mutations in p53 - occurring in >50% of all cancers, 0.2% of all healthy humans - are not just inherited, but commonly accumulate during organismal aging or exposure to carcinogens. In genetic studies on murine models of in vitro aging of RBCs (i.e., under conditions that mimic blood storage in the blood bank) we have documented that hypermorphic STEAP3 is associated with poor blood storage quality, owing to increased oxidant stress and elevated lipid peroxidation. Iron-mediated non-apoptotic cell death via lipid peroxidation is a hallmark of ferroptosis, a novel process of cell death investigated extensively in nucleated cells, but hitherto ignored in iron-loaded RBCs. More than other cells, RBCs rely on antioxidant systems to keep oxidant stress in check. A key antioxidant system is represented by the soluble tripeptide glutathione, glutathione-dependent detoxification systems (e.g., glutathione-peroxidase 4 - GPX4 to counteract lipid peroxidation) and oxidized glutathione recycling via NADPH-dependent enzymes. The main pathway that sustains NADPH synthesis in RBCs is the pentose phosphate pathway. Glucose 6-phoshate dehydrogenase (G6PD) is the rate-limiting enzyme of this pathway, an X-linked gene that is mutated in ~500 million people. Oxidant damage to protein triggers formation of isoaspartyl damage, a process counteracted by the enzyme PIMT. Relevant to this proposal, in other cell types p53 promotes ferroptosis by up-regulation of STEAP3 and down-regulation of G6PD, while being itself negatively regulated by PIMT. Even though genetic and pharmacological tools are available to regulate ferroptosis, these approaches are untested in mature RBCs, which is the focus of this project. Relevance to public health: RBC responses to hypoxia and oxidant stress regulate hemolysis, an etiological contributor to physio/pathological adaptations to pro-oxidant challenges in vivo (e.g., high altitude, exercise) or ex vivo, such as blood bank storage of RBCs for transfusion purposes, ranked 1st in the list of breakthroughs in the history of mankind that are credited with saving the most lives.
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