Differential HIF and NOS responses to acute anemia: defining organ-specific hemoglobin thresholds for tissue hypoxia

Tissue hypoxia likely contributes to anemia-induced organ injury and mortality. Severe anemia activates hypoxia-inducible factor (HIF) signaling by hypoxic-and neuronal nitric oxide (NO) synthase-(nNOS) dependent mechanisms. However, organ-specific hemoglobin (Hb) thresholds for increased HIF expression have not been defined. To assess organ-specific Hb thresholds for tissue hypoxia, HIF-alpha (oxygen-dependent degradation domain, ODD) luciferase mice were hemodiluted to mild, moderate, or severe anemia corresponding to Hb levels of 90, 70, and 50 g/l, respectively. HIF luciferase reporter activity, HIF protein, and HIF-dependent RNA levels were assessed. In the brain, HIF-1 alpha was paradoxically decreased at mild anemia, returned to baseline at moderate anemia, and then increased at severe anemia. Brain HIF-2 alpha remained unchanged at all Hb levels. Both kidney HIF-1 alpha and HIF-2 alpha increased earlier (Hb similar to 70-90 g/l) in response to anemia. Liver also exhibited an early HIF-alpha response. Carotid blood flow was increased early (Hb similar to 70, g/l), but renal blood flow remained relatively constant, only increased at Hb of 50 g/l. Anemia increased nNOS (brain and kidney) and endothelia NOS (eNOS) (kidney) levels. Whereas anemia- induced increases in brain HIF alpha were nNOS-dependent, our current data demonstrate that increased renal HIF alpha was nNOS independent. HIF-dependent RNA levels increased linearly (similar to 10-fold) in the brain. However, renal HIF-RNA responses (MCT4, EPO) increased exponentially (similar to 100-fold). Plasma EPO levels increased near Hb threshold of 90 g/l, suggesting that the EPO response is sensitive. Collectively, these observations suggest that each organ expresses a different threshold for cellular HIF/NOS hypoxia responses. This knowledge may help define the mechanism(s) by which the brain and kidney maintain oxygen homeostasis during anemia.