Background Intravascular red cell hemolysis impairs NO-redox homeostasis, producing endothelial dysfunction, platelet activation and vasculopathy. Red blood cell storage under standard conditions results in reduced integrity of the erythrocyte membrane, with formation of exocytic microvesicles or “microparticles” and hemolysis, which we hypothesized could impair vascular function and contribute to the putative “storage lesion” of banked blood. Methods and Results We now find that storage of human red blood cells under standard blood banking conditions results in the accumulation of cell free and microparticle-encapsulated hemoglobin which, despite 39 days of storage, remains in the reduced ferrous oxyhemoglobin redox state and stoichiometrically reacts with and scavenges the vasodilator nitric oxide (NO). Using stopped-flow spectroscopy and laser triggered NO release from a caged NO compound we found that both free hemoglobin and microparticles react with NO about 1000 times faster than with intact erythrocytes. In complementary in vivo studies we show that hemoglobin, even at concentrations below 10 μM (in heme), produces potent vasoconstriction when infused into the rat circulation, while controlled infusions of methemoglobin and cyanomethemoglobin, which do not consume NO, have substantially reduced vasoconstrictor effects. Infusion of the plasma from stored human red cell units into the rat circulation produces significant vasoconstriction related to the magnitude of storage related hemolysis. Conclusions The results of these studies suggest new mechanisms for endothelial injury and impaired vascular function associated with the most fundamental of storage lesions, hemolysis.

. Nitrite as a vascular endocrine nitric oxide reservoir that contributes to hypoxic signaling, cytoprotection, and vasodilation. Am J Physiol Heart Circ Physiol 291: H2026 -H2035, 2006. First published June 23, 2006 doi:10.1152/ajpheart.00407.2006.-Accumulating evidence suggests that the simple and ubiquitous anion salt, nitrite (NO 2 Ϫ ), is a physiological signaling molecule with potential roles in intravascular endocrine nitric oxide (NO) transport, hypoxic vasodilation, signaling, and cytoprotection after ischemia-reperfusion. Human and animal studies of nitrite treatment and NO gas inhalation provide evidence that nitrite mediates many of the systemic therapeutic effects of NO gas inhalation, including peripheral vasodilation and prevention of ischemia-reperfusion-mediated tissue infarction. With regard to nitritedependent hypoxic signaling, biochemical and physiological studies suggest that hemoglobin possesses an allosterically regulated nitrite reductase activity that reduces nitrite to NO along the physiological oxygen gradient, potentially contributing to hypoxic vasodilation. An expanded consideration of nitrite as a hypoxiadependent intrinsic signaling molecule has opened up a new field of research and therapeutic opportunities for diseases associated with regional hypoxia and vasoconstriction.hemoglobin; hypoxia; S-nitrosated albumin; cysteine 93 HYPOXIC VASODILATION is a conserved systemic physiological response that matches blood flow and oxygen delivery to tissue metabolic demand. This hypoxic response has been appreciated for more than 100 years since the initial description by Roy and Brown in 1879 (80). This response is thought to involve feedback mechanisms that require oxygen or pH sensing of a divergence in the normal relationship between delivered blood oxygen and tissue oxygen consumption (94). This leads to the feedback generation of putative vasodilatory effectors that increase blood flow to maintain adequate tissue oxygenation. Important to the considerations of the mechanisms responsible for oxygen sensing, in mammals hypoxic vasodilation appears to occur as the hemoglobin desaturates from 60% to 40%, around a partial pressure of oxygen ranging from . Surprisingly, measurements of microcirculatory and tissue oxygen tension and hemoglobin oxygen saturation using modern methodologies suggest that much of the oxygen delivery occurs within the resistance arterioles, especially in the case of skeletal muscle (91). Thus, in these microvascular beds, the anatomical site of hypoxic sensing is proximal to the site of resistive control (arterioles and arteriolar capillaries). In other tissues, such as heart and brain, more oxygen is extracted within the capillary network. This creates a paradox as to how hypoxic sensing can modulate retrograde feedback vasodilation in these tissues. The solution to this paradox has been in part solved by the work of , who suggested that acetycholine-dependent vasodilation of the capillary or venous circulation produces retrograde intracellular propagation of ...

The anion nitrite (NO(2)(-)) constitutes a biochemical reservoir for nitric oxide (NO). Nitrite reduction to NO may be catalyzed by hemoglobin, myoglobin or other metal-containing enzymes and occurs at increasing rates under conditions of physiologic hypoxia or ischemia. A number of laboratories have now demonstrated in animal models the ability of nitrite to provide potent cytoprotection following focal ischemia-reperfusion (IR) injury of the heart, liver, brain, and kidney. While the mechanism of nitrite-mediated cytoprotection remains to be fully characterized, the release of nitrite-derived NO following IR appears to be central to this mechanism. The evidence of nitrite-mediated cytoprotection against IR injury in multiple animal models opens the door to potential therapeutic opportunities in human disease. Here we review the mechanisms for nitrite formation in blood and tissue, its metabolic equilibrium with NO, nitrate, and NO-modified proteins, the evidence supporting nitrite-mediated cytoprotection, and the potential mechanisms driving cytoprotection, and we explore the opportunities for the therapeutic application of nitrite for human disease.

This study shows that at low hematocrit, the oxygen-delivering capacity of human red blood cells stored 5-6 wks is reduced compared with fresh cells and red blood cells stored for an intermediate period. Although red blood cells stored for 2-3 wks are completely devoid of 2,3-diphosphoglycerate, their oxygen-delivering capacity to the intestines was the same as fresh red blood cells. Our study showed that red blood cell deformability was preserved during storage, suggesting that other mechanisms may account for the observed decrease in oxygen delivery by red blood cells stored 2-3 wks.