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A role for nitric oxide (NO) produced during the reduction of nitrite by deoxygenated red blood cells (RBCs) in regulating vascular dilation has been proposed. It has not, however, been satisfactorily explained how this NO is released from the RBC without first reacting with the large pools of oxyhemoglobin and deoxyhemoglobin in the cell. In this study, we have delineated a mechanism for nitrite-induced RBC vasodilation that does not require that NO be released from the cell. Instead, we show that nitrite enhances the ATP release from RBCs, which is known to produce vasodilation by several different methods including the interaction with purinergic receptors on the endothelium that stimulate the synthesis of NO by endothelial NO synthase. This mechanism was established in vivo by measuring the decrease in blood pressure when injecting nitrite-reacted RBCs into rats. The observed decrease in blood pressure was not observed if endothelial NO synthase was inhibited by [N.sup.[omega]]-nitro-L-arginine methyl ester (L-NAME) or when any released ATP was degraded by apyrase. The nitrite-enhanced ATP release was shown to involve an increased binding of nitrite-modified hemoglobin to the RBC membrane that displaces glycolytic enzymes from the membrane, resulting in the formation of a pool of ATP that is released from the RBC. These results thus provide a new mechanism to explain nitrite-induced vasodilation. nitrite reduction; nitric oxide; anerobic glycolysis; hypoxia; red blood cell; adenosine 5'-triphosphate doi: 10.1152/ajpheart.01233.2008

The centrifugation data show that the binding of hemoglobin to band 3 protein alters the protein conformation, which changes the equilibrium constants for the the second hemoglobin binding to the protein dimer. Further, the fluorescence data actually measures the binding event. Research points out both measurements are needed to understand the binding phenomenon.

Research has been conducted on the chemically modified human and bovine hemoglobins. The systematic evaluation of these hemoglobins has been carried out and the toxicity of the hemoglobin-based oxygen carriers has been investigated in establishing the importance of the secondary oxidative reactions.

Research has been conducted on the heme degradation as a result of ferrylhemoglobin reaction with hydrogen. The mechanism of heme degradation during the reaction of oxyHb wit H (sub)2 O (sub)2 has been investigated and the results are presented.

Research was conducted to examine the curvature of dinucleotide poised for formation of trinucleotide in transcription with Escherichia coli RNA polymerase. The curvature was observed from the demonstration that a metal ion bound with ta mononucleotide in the i+1 elongation site is equidistant from bases at the 5' end and 3' end of the dinucleotide. Results suggest curvature to be a critical feature in the interaction between RNA elongate and DNA template even at the very start of transcription.

The function of the beta-93 cysteine residue in the hemoglobin autoxidation mechanism was studied by electron paramagnetic resonance. Investigation of the free radical spectrum showed that beside the superoxide radical, an isotropic free radical is formed with a g(sub iso) of 2.0133. The formation of the thiyl radical was observed to reduce the rate of autoxidation for the beta-chain and decreases heme degradation caused by the reaction of superoxide with the heme.

Low temperature electron paramagnetic resonance reveals the formation of a superoxide radical from the hemoglobin-bound oxygen blocked in the hydrophobic heme pocket where dismutation is slow. The formation of the free radical at intermediate oxygen pressures occurs when the rates of hemoglobin autoxidation for partly oxygenated intermediates increases. The flexibility of the heme pocket, which is high under hypoxia, possibly contributes to the formation of the superoxide.

Abstract Background Our earlier studies have shown that red blood cell (RBC) morphology in Alzheimer's disease (AD) subjects was altered (> 15% of the RBCs were elongated as compared to 5.9% in normal controls (p < 0.0001)). These results suggested alterations in the RBC membrane architecture in AD subjects, possibly due to RBC-β-amyloid interactions and/or changes in the expression of membrane proteins. We hypothesized that the observed changes could be due to changes in the level of the protein components of the cytoskeleton and those linked to the RBC membrane. To examine this, we performed a proteomic analysis of RBC membrane proteins of AD subjects, and their age-matched controls using one pool of samples from each group, following their separation by SDS-PAGE, in-gel Tryptic digestion, LC-MS-MS of peptides generated, and a label-free approach of semi-quantitative analysis of their relative MS spectral intensities. Results The data suggest, (1) RBC shape/morphology changes in AD subjects are possibly attributed primarily to the changes (elevation or decrease) in the level of a series of membrane/cytoskeleton proteins involved in regulating the stability and elasticity of the RBC membrane, and (2) changes (elevation or decrease) in the level of a second series of proteins in the RBC membrane proteome reflect similar changes reported earlier by various investigators in AD or animal model of AD. Of particular interest, elevation of oxidative stress response proteins such as heat shock 90 kDa protein 1 alpha in AD subjects has been confirmed by western blot analysis in the RBC membrane proteome. Conclusions The results suggest that this study provides a potential link between the alterations in RBC membrane proteome in AD subjects and AD pathology.