Endothelial Hemoglobin Alpha Mediates Iron Regulation of Endothelial Nitric Oxide

Abstract

Iron deficiency anemia (IDA) is a common comorbidity in cardiovascular disease. Previous clinical studies have found iron deficiency anemia (IDA) increases endothelial NO signaling and attributed this change to the reduction in circulating hemoglobin. However, it has been shown in ex vivo preparations that endothelial NO signaling is unaffected by the presence of erythrocytes suggesting changes in circulating hemoglobin do not alter NO signaling. Our lab has previously shown the hemoglobin-α (Hbα) is expressed in the endothelium of small arteries and scavenges NO. We hypothesized endothelial Hbα, rather than blood hemoglobin is responsible for the increased NO signaling in IDA. We first tested this hypothesis in a mouse model of IDA in which we were able to replete vascular iron stores. Male and female C57BL/6 mice were fed an iron deficient (2-6 ppm Fe) or nutrient matched control diet (48 ppm Fe) beginning at weaning. IDA mice underwent phlebotomies (10% blood volume) at 9 and 11 weeks old. To restore vascular iron stores, a subset of IDA mice received a single i.p. injection of iron dextran (FeDex; 20 mg/kg) at 12 weeks. All measurements were taken one week later. As expected, IDA mice had low blood hemoglobin, and FeDex did not rescue the anemia but did rescue vascular iron stores as measured by ferritin light chain protein. To investigate NO signaling, we used laser speckle contrast imaging to measure changes in in vivo blood flow in response to the NO synthase inhibitor L-NAME. The magnitude of the response to L-NAME is an indicator of NO signaling. IDA mice exhibited an exaggerated response to L-NAME indicating increased NO signaling. FeDex treatment restored the L-NAME response back to control levels suggesting reduced vascular iron stores, rather than the anemia itself increased NO signaling. In order to investigate the role of endothelial Hbα, we repeated these studies in endothelial specific Hbα knockout mice (Hba1fl/fl-Cdh5-CreERT2+). Lack of endothelial Hbα did not prevent the increased response to L-NAME in IDA mice but did prevent the rescue by FeDex. This suggests a model in which endothelial Hba participates in regulating NO in response to iron but not the increased NO in IDA. Because Hbα does not explain the initial increase in NO, we have begun investigating other anemia-associated NO stimulators. Erythropoetin (EPO) is known to stimulate NO production in endothelial cells and is elevated in our anemia model regardless of FeDex treatment. Future work will investigate the role of EPO in regulating NO in IDA. University of Virginia Basic and Translational Research Training Grant T32 007284 (LSD, MAL, SAL); NIH HL088554; LaunchPad; Lipedema Foundation; AHA Predoctoral Fellowship (MAL). This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.