Background: To date, there is no safe and effective hemoglobin (Hb)-based oxygen carrier (HBOC) to substitute for red blood cell transfusion.Clinical application of HBOCs has been stymied by the adverse side effects of nitric oxide (NO) scavenging.The precise mechanisms remain unclear.We hypothesized that reduced NO bioavailability due to endothelial dysfunction would enhance vasospasm after HBOC challenge.Materials and Methods: PolyHeme(R) was provided by Northfield Laboratories (Evanston, IL), MP4-OX and MP4-CO were provided by Sangart (San Diego, CA) and Hb-vesicles (HbV) were obtained from Dr.Hiromi Sakai (Waseda Bioscience Research Institute, Singapore).All Hb solutions were given as a top load (16% of blood volume) via a tail vein.Systolic blood pressure (SBP) and heart rate was measured in awake standard-diet (SD) fed wild-type mice (SD-fed WT, C57BL6), WT mice fed a high-fat diet for 4-6 weeks (HFD-fed WT), and diabetic (db/db) mice by noninvasive tail-cuff.Invasive hemodynamics measurements were obtained in anesthetized mice.Blood and tissue samples were collected 2 h after HBOCs infusion for inflammation analysis.Results: In SD-fed WT mice, infusing MP4-OX or MP4-CO produced systemic vasoconstriction, but infusing PolyHeme or HbV did not.In HFD-fed WT mice or db/db mice (with endothelial dysfunction), infusing PolyHeme induced systemic hypertension, but there was none after infusing HbV.These findings were confirmed with invasive hemodynamic measurements obtained in anesthetized mice.Infusion of PolyHeme, MP4 or HbV at 2 h did not change liver and lung levels of mRNAs encoding IL-6, TNF-, TF or HO-1.At 2 h, no changes were observed in plasma levels of IL6, IL-1 0, or MCP-1 after PolyHeme or MP4 infusion.Conclusions: Our results demonstrate that some HBOCs (e.g.MP4-OX or-CO) produce more hypertension (due to systemic vasoconstriction) than others (PolyHeme) in SD-fed WT mice.However, mice with endothelial dysfunction (db/db) are more sensitive to NO scavenging effects, and even PolyHeme produces vasoconstriction.HbV did not produce vasoconstriction in db/db mice.None of the agents produced elevated cytokine levels in tissues or plasma at 2 h.Future HBOCs should be studied in models with endothelial dysfunction.