Biomedical subjects

M B Grisham

Publications and source records attributed to M B Grisham.

At least 73 records · Page 4Linked to original sources

PubMed1999

Indomethacin induced gastropathy in CD18, intercellular adhesion molecule 1, or P-selectin deficient mice.

BACKGROUND: Neutrophil-endothelial cell interactions are thought to play a critical role in the pathophysiology of non-steroidal anti-inflammatory drug (NSAID) induced gastropathy. AIMS: To optimise a mouse model of NSAID induced gastropathy and to evaluate the importance of adhesion molecules using adhesion molecule deficient mice. METHODS: Gastropathy was induced in C57BL/6 mice or their adhesion molecule deficient counterparts via oral administration of indomethacin (20 mg/kg). Lesion scores, mucosal permeability, and histopathology were used to assess gastric mucosal injury. RESULTS: Intragastric administration of indomethacin induced linear haemorrhagic mucosal lesions, primarily in the corpus of the stomach that were first observed at six hours. These lesions continued to develop over the next six hours with maximal lesion scores and mucosal permeabilities at 12 hours. When indomethacin was administered to mice deficient in CD18, intercellular adhesion molecule 1 (ICAM-1), or P-selectin, there were significant decreases in lesion scores compared with their C57BL/6 controls. In addition, mucosal permeabilities were found to be significantly lower in CD18 or ICAM-1 deficient mice observed at 12 hours. CONCLUSION: Certain leucocyte and endothelial cell adhesion molecules are important determinants for full expression of indomethacin induced gastropathy. It is proposed that this modification of the mouse model may be useful for the investigation of other pathophysiological mechanisms of NSAID induced gastropathy.

Animals↗

PubMed1999

Effect of selective proteasome inhibitors on TNF-induced activation of primary and transformed endothelial cells.

The objective of this study was to assess the effects of two structurally distinct yet selective proteasome inhibitors (PS-341 and lactacystin) on leukocyte adhesion, endothelial cell adhesion molecule (ECAM) expression, and nuclear factor-kappaB (NF-kappaB) activation in tumor necrosis factor (TNF)-alpha-stimulated human umbilical vein endothelial cells (HUVEC) and the transformed, HUVEC-derived, ECV cell line. We found that TNF (10 ng/ml) significantly enhanced U-937 and polymorphonuclear neutrophil (PMN) adhesion to HUVEC but not to ECV; TNF also significantly enhanced surface expression of vascular cell adhesion molecule 1 and E-selectin (in HUVEC only), as well as intercellular adhesion molecule 1 (ICAM-1; in HUVEC and ECV). Pretreatment of HUVEC with lactacystin completely blocked TNF-stimulated PMN adhesion, partially blocked U-937 adhesion, and completely blocked TNF-stimulated ECAM expression. Lactacystin attenuated TNF-stimulated ICAM-1 expression in ECV. Pretreatment of HUVEC with PS-341 partially blocked TNF-stimulated leukocyte adhesion and ECAM expression. These effects of lactacystin and PS-341 were associated with inhibitory effects on TNF-stimulated NF-kappaB activation in both HUVEC and ECV. Our results demonstrate the importance of the 26S proteasome in TNF-induced activation of NF-kappaB, ECAM expression, and leukocyte-endothelial adhesive interactions in vitro.

Acetylcysteine↗

PubMed1999

Nitric oxide. I. Physiological chemistry of nitric oxide and its metabolites:implications in inflammation.

The role of nitric oxide (NO) in inflammation represents one of the most studied yet controversial subjects in physiology. A number of reports have demonstrated that NO possesses potent anti-inflammatory properties, whereas an equally impressive number of studies suggest that NO may promote inflammation-induced cell and tissue dysfunction. The reasons for these apparent paradoxical observations are not entirely clear; however, we propose that understanding the physiological chemistry of NO and its metabolites will provide a blueprint by which one may distinguish the regulatory/anti-inflammatory properties of NO from its deleterious/proinflammatory effects. The physiological chemistry of NO is complex and encompasses numerous potential reactions. In an attempt to simplify the understanding of this chemistry, the physiological aspects of NO chemistry may be categorized into direct and indirect effects. This type of classification allows for consideration of timing, location, and rate of production of NO and the relevant targets likely to be affected. Direct effects are those reactions in which NO interacts directly with a biological molecule or target and are thought to occur under normal physiological conditions when the rates of NO production are low. Generally, these types of reactions may serve regulatory and/or anti-inflammatory functions. Indirect effects, on the other hand, are those reactions mediated by NO-derived intermediates such as reactive nitrogen oxide species derived from the reaction of NO with oxygen or superoxide and are produced when fluxes of NO are enhanced. We postulate that these types of reactions may predominate during times of active inflammation. Consideration of the physiological chemistry of NO and its metabolites will hopefully allow one to identify which of the many NO-dependent reactions are important in modulating the inflammatory response and may help in the design of new therapeutic strategies for the treatment of inflammatory tissue injury.

Chemical Phenomena↗

PubMed1999

Cardiac impairment and nitric oxide synthase activity in the chronic portal vein-stenosed rat.

Decreased cardiac contractility and beta-adrenergic responses have been observed in the chronic portal vein-stenosed (PVS) rat. Because nitric oxide (NO) may be increased in PVS and has been recognized as a negative inotropic agent, we investigated the induction of NO synthase (NOS2) and/or changes in constitutive NOS (NOS3) as factors in the cardiac dysfunction of the PVS rat. Ten to twelve days after portal vein stenosis or sham operation, cardiac function was evaluated in paced left ventricular papillary muscles (LVPM) and right ventricular strips (RV). To determine if NO modulation of contractile function was altered in PVS, we examined the increase in developed tension produced by the effect of Nomega-nitro-L-arginine (L-NNA) on the myocardial force-frequency relationship. Cardiac tissue NOS2 and NOS3 activities were assayed, Western blot analyses of NOS2 and NOS3 expression were performed, and circulating nitrate-nitrite (NOX) levels (an indicator of in vivo NOS activity) were assayed. Basal LVPM and RV contractile indexes were significantly reduced in PVS (30-50%), without a change in the relaxation rate. No between-group differences in the cardiac NOS2 or NOS3 enzymatic activities of PVS and sham-operated (SO) rats were observed. Western blots revealed no cardiac NOS2 expression in either SO or PVS rats. In contrast, NOS3 was expressed in both SO and PVS rats, but there was no quantitative difference in expression between the two groups. Changes in the cardiac force-frequency relationship (staircase effect) after L-NNA were consistent with NOS3 modulation of contractile function in both SO and PVS rats, but there was no between-group difference in the modulation. Circulating NOX concentrations did not differ between SO and PVS rats. In conclusion, protein expression data, enzymatic assays, end-product assays, and functional data indicate that between-group differences in NOS2 and NOS3 activity are not responsible for the cardiac impairment that has been observed in the chronic PVS rat.

Animals↗

PubMed1999

Role of inducible nitric oxide synthase in the regulation of VCAM-1 expression in gut inflammation.

The objectives of this study were to assess the role of the inducible isoform of nitric oxide synthase (iNOS) on vascular cell adhesion molecule 1 (VCAM-1) expression in vivo in an acute model of inflammation induced in iNOS-deficient (iNOS-/-) mice and compare these data to those obtained by pharmacological inhibition of iNOS in a CD4+ T lymphocyte-dependent model of chronic colitis. VCAM-1 expression was quantified in vivo using the dual radiolabel monoclonal antibody technique. We found that intraperitoneal injection of 10 microg/kg tumor necrosis factor-alpha (TNF-alpha) enhanced VCAM-1 expression by approximately twofold in the colon, cecum, and stomach but not small intestine in iNOS-/- mice compared with TNF-alpha-injected wild-type mice. Injection of wild-type mice with 25 microg/kg TNF-alpha further enhanced VCAM-1 expression by approximately twofold compared with wild-type mice injected with 10 microg/kg TNF-alpha; however, VCAM-1 expression was not further enhanced in any gastrointestinal organ system in iNOS-/- mice. In a second series of experiments, we found that continuous inhibition of iNOS using oral administration of NG-iminoethyl-L-lysine did not alter the enhanced levels of VCAM-1 expression in the colon nor did it alter the severity of colonic inflammation in SCID mice reconstituted with CD4+, CD45RB(high) T cells. We conclude that iNOS may regulate VCAM-1 expression in acute inflammation; however, this effect is modest and tissue specific and occurs only when VCAM-1 expression is submaximal. iNOS does not appear to modulate VCAM-1 expression in an immune model of chronic colitis.

Animals↗

PubMed1999

Myocardial ischemia-reperfusion injury is exacerbated in absence of endothelial cell nitric oxide synthase.

Myocardial ischemia and reperfusion (MI/R) initiates a cascade of polymorphonuclear neutrophil (PMN)-mediated injury, the magnitude of which may be influenced by the bioavailability of nitric oxide (NO). We investigated the role of endothelial cell nitric oxide synthase (ecNOS) in MI/R injury by subjecting wild-type and ecNOS-deficient (-/-) mice to 20 min of coronary artery occlusion and 120 min of reperfusion. Myocardial infarct size represented 20.9 +/- 2.9% of the ischemic zone in wild-type mice, whereas the ecNOS -/- mice had significantly (P < 0.01) larger infarcts measuring 46.0 +/- 3.8% of the ischemic zone. Because P-selectin is thought to be involved with the pathogenesis of neutrophil-mediated I/R injury, we assessed the effects of MI/R on P-selectin expression in the myocardium of wild-type and ecNOS -/- mice. P-selectin expression measured with a radiolabeled monoclonal antibody (MAb) technique after MI/R in wild-type mice was 0.037 +/- 0.009 microgram MAb/g tissue, whereas ecNOS -/- coronary vasculature was characterized by significantly (P < 0.05) higher P-selectin expression (0.080 +/- 0.013 microgram MAb/g tissue). Histological examination of the postischemic myocardium revealed significantly (P < 0.01) more neutrophils in the ecNOS -/- (29.5 +/- 2.5 PMN/field) compared with wild-type (5.0 +/- 0.9 PMN/field) mice. A similar trend in infarct size and neutrophil accumulation was observed when wild-type and ecNOS -/- mice were subjected to 30 min of ischemia and 120 min of reperfusion. These novel in vivo findings demonstrate a cardioprotective role for ecNOS-derived NO in the ischemic-reperfused mouse heart.

Animals↗

PubMed1999

Leukocyte-endothelial cell interactions in nitric oxide synthase-deficient mice.

Nitric oxide (NO) is known to be an important endogenous modulator of leukocyte-endothelial cell interactions within the microcirculation. We examined leukocyte rolling and adhesion under baseline conditions and following thrombin (0.25 U/ml) superfusion in the mesentery of wild-type, inducible NOS (iNOS)-deficient (-/-), neuronal NOS (nNOS) -/-, and endothelial cell NOS (ecNOS) -/- mice to further our understanding of NO and leukocyte function. Baseline leukocyte rolling (cells/min) was significantly elevated in both the nNOS -/- (30.0 +/- 4.0) and ecNOS -/- mice (67.0 +/- 12.0) compared with wild-type mice (11.0 +/- 1.4). In addition, baseline leukocyte adherence (cells/100 micrometers of vessel) was also significantly elevated in the nNOS -/- (5.2 +/- 1.0) and ecNOS -/- (13.0 +/- 1.3) compared with wild-type animals (1.3 +/- 0.5). Deficiency of iNOS had no effect on baseline leukocyte rolling or adhesion in the mesentery. Baseline surface expression of P-selectin was observed in 68.0 +/- 9.0% of intestinal venules in ecNOS -/- mice compared with 10.0 +/- 2.0% in wild-type mice. Additional studies demonstrated that administration of an anti-P-selectin monoclonal antibody (RB40. 34) or the soluble P-selectin ligand, PSGL-1, completely inhibited the increased rolling and firm adhesion response in nNOS -/- and ecNOS -/- mice. Transmigration of neutrophils into the peritoneum following thioglycollate injection was also significantly augmented in nNOS -/- and ecNOS -/- mice. These studies clearly indicate the NO derived from both nNOS and ecNOS is critical in the regulation of leukocyte-endothelial cell interactions.

Animals↗

PubMed1999

Inhaled NO impacts vascular but not extravascular compartments in postischemic peripheral organs.

Inhaled nitric oxide (NO) reduces pulmonary hypertension and dampens various aspects of lung inflammation; however, its effects are thought to be restricted to the lung because of its short half-life in biological systems. More recently, however, NO was shown to nitrosylate hemoglobin, albumin, and other plasma molecules to form stable nitrosothiol derivatives and could have an impact on the periphery. We examined whether inhaled NO could have an impact on the two compartments of distal organs, namely, the intravascular and extravascular spaces. The feline intestine was exposed to 1 h of ischemia and 1 h of reperfusion, and intestinal blood flow and mucosal dysfunction were measured in animals ventilated with room air and inhaling 0 or 80 ppm NO. A decrease in intestinal blood flow and an increase in mucosal barrier leakiness were noted in animals not exposed to inhaled NO. The intestinal blood flow impairment was entirely reversed in animals breathing 80 ppm NO, but the mucosal dysfunction was not affected. We further examined whether inhaled NO could reach the extravascular space by simply inhibiting NO in the intestine with the NO synthase inhibitor N(G)-nitro-L-arginine methyl ester (L-NAME) that causes an increase in mucosal permeability that is rapidly reversed with NO donors. However, inhaled NO had no effect on the rise in mucosal permeability. L-NAME reduced lymph nitrosothiol concentrations, but inhaled NO could not replenish these levels. To further explore the intravascular impact of inhaled NO, we used intravital microscopy to visualize the microvasculature and demonstrated that inhaled NO could be initiated after reperfusion and still reduced microvascular disturbances, including reversing the impairment in blood flow and increasing leukocyte adhesion. The effects of inhaled NO persisted for an additional hour after termination of NO inhalation, consistent with a dramatic increase in nitrate within 1 h of NO inhalation, which persisted for 1 h after the termination of NO inhalation. These data suggest that inhaled NO can reach distal organs to dramatically improve reperfusion-induced microvascular but not extravascular dysfunction.

Administration, Inhalation↗

PubMed1999

Effects of reactive oxygen and nitrogen metabolites on MCP-1-induced monocyte chemotactic activity in vitro.

Peroxynitrite, an oxidant generated by the interaction between superoxide and nitric oxide (NO), can nitrate tyrosine residues, resulting in compromised protein function. Monocyte chemoattractant protein-1 (MCP-1) is a chemokine that attracts monocytes and has a tyrosine residue critical for function. We hypothesized that peroxynitrite would alter MCP-1 activity. Peroxynitrite attenuated MCP-1-induced monocyte chemotactic activity (MCA) in a dose-dependent manner (P < 0.05) but did not attenuate leukotriene B4 or complement-activated serum MCA. The reducing agents dithionite, deferoxamine, and dithiothreitol reversed the MCA inhibition by peroxynitrite, and exogenous L-tyrosine abrogated the inhibition by peroxynitrite. PAPA-NONOate, an NO donor, or superoxide generated by xanthine and xanthine oxidase did not show an inhibitory effect on MCA induced by MCP-1. The peroxynitrite generator 3-morpholinosydnonimine caused a concentration-dependent inhibition of MCA by MCP-1. Peroxynitrite reduced MCP-1 binding to monocytes and resulted in nitrotyrosine formation. These findings are consistent with nitration of tyrosine by peroxynitrite, with subsequent inhibition of MCP-1 binding to monocytes, and suggest that peroxynitrite may play a role in regulation of MCP-1-induced monocyte chemotaxis.

Blood Physiological Phenomena↗

PubMed1999

Selective proteasome inhibitors as anti-inflammatory agents.

Nuclear factor kappaB (NF-kappaB) is an ubiquitous transcription factor and pleiotropic regulator of numerous inflammatory and immune responses. Once activated, NF-kappaB translocates from the cytosol to the nucleus of the cell, where it binds to its consensus sequence on the promoter-enhancer region of different genes. By so doing, this activates the transcription of a variety of different pro-inflammatory cytokines, adhesion molecules and specific enzymes, such as the inducible forms of nitric oxide synthase and cyclooxygenase. A number of different cytokines, bacterial products and oxidants activate NF-kappaB via selective phosphorlyation, polyubiquitination and degradation of the inhibitor protein, IkappaB. Since the 26S proteasome complex degrades the post-translationally modified IkappaB, thereby liberating the transcriptionally active p50/p65 heterodimeric NF-kappaB, this proteolytic complex represents a critical step in the activation of NF-kappaB. This review discusses the basic biology of the ubiquitin-proteasome pathway as it relates to the inflammatory response, and highlights those studies demonstrating that selective proteasome inhibitors are effective anti-inflammatory agents in vivo.

Journal Article↗

PubMed1999

Effect of reactive oxygen metabolites on endothelial permeability: role of nitric oxide and iron.

OBJECTIVE: We evaluated the effects of the xanthine oxidase (XO)-derived reactive oxygen metabolites on the permeability of bovine pulmonary artery-endothelial monolayers and examined how iron and nitric oxide (NO) participate in these changes in permeability. METHODS: Permeability was measured using a cell-column chromatographic method in which monolayers were exposed to combinations of agents. RESULTS: Exposure of monolayers to a superoxide/peroxide generator, xanthine (X, 0.1 mM)/XO (25 mU/mL), increased solute permeability after 10 minutes, but the same dose of either X or XO alone did not. Exposure of monolayers to peroxide (0.1 mM) also increased permeability, but only after 70 minutes. This X/XO permeability was attenuated by either catalase, superoxide dismutase, methionine (1 mM), an oxy-radical scavenger, or desferrioxamine (0.1 mM), an iron chelator. Spermine NONOate (SNO), an NO donor, attenuated X/XO permeability at 0.1 mM, but this protection was not significant at 0.01 or 1 mM. Spermine NONOate (0.1 mM) did not alter the permeability produced by 0.1 mM peroxide. L-N5-(1-iminoethyl)-ornithine (10 microM), an NO synthase inhibitor, completely blocked peroxide-, and partially attenuated X/XO-mediated permeability. However, 3-morphosynodiomine (SIN-1, 1 mM) plus catalase (1,000 U/mL), a peroxynitrite generator, did not alter permeability. CONCLUSIONS: Xanthine/Xanthine Oxidase permeability involves peroxide, superoxide, oxy-radicals, and iron. Endogenous NO may regulate peroxide-, but not superoxide-mediated permeability. The protective effects of exogenous NO on the X/XO permeability may represent interactions between superoxide, peroxide, and cell surface-bound iron.

Animals↗

PubMed1998

Role of the proteasome and NF-kappaB in streptococcal cell wall-induced polyarthritis.

The transcription factor NF-kappaB activates a number of genes whose protein products are proinflammatory. In quiescent cells, NF-kappaB exists in a latent form and is activated via a signal-dependent proteolytic mechanism in which the inhibitory protein IkappaB is degraded by the ubiquitin-proteasome pathway. Consequently, inhibition of the proteasome suppresses activation of NF-kappaB. This suppression should therefore decrease transcription of many genes encoding proinflammatory proteins and should ultimately have an anti-inflammatory effect. To this end, a series of peptide boronic acid inhibitors of the proteasome, exemplified herein by PS-341, were developed. The proteasome is the large multimeric protease that catalyzes the final proteolytic step of the ubiquitin-proteasome pathway. PS-341, a potent, competitive inhibitor of the proteasome, readily entered cells and inhibited the activation of NF-kappaB and the subsequent transcription of genes that are regulated by NF-kappaB. Significantly, PS-341 displayed similar effects in vivo. Oral administration of PS-341 had anti-inflammatory effects in a model of Streptococcal cell wall-induced polyarthritis and liver inflammation in rats. The attenuation of inflammation in this model was associated with an inhibition of IkappaBalpha degradation and NF-kappaB-dependent gene expression. These experiments clearly demonstrate that the ubiquitin-proteasome pathway and NF-kappaB play important roles in regulating chronic inflammation and that, as predicted, proteasome inhibition has an anti-inflammatory effect.

Animals↗

PubMed1998

The reaction of S-nitrosoglutathione with superoxide.

The nitric oxide (NO)-dependent S-nitrosation of thiols to generate S-nitrosothiols has been proposed as an important pathway for the metabolism of NO in vivo. Although it has been suggested that these S-nitrosated compounds are resistant to decomposition by reactive oxygen metabolites (ROMs), very little information is available regarding the interaction between S-nitrosothiols and ROMs. We found that S-nitrosoglutathione (GSNO) rapidly reacted with O2- to generate glutathione disulfide and equimolar quantities of nitrite and nitrate. The reaction was second order with respect of GSNO and first order with respect of O2- with a rate equation of -d[GSNO]/dt = 2k3[GSNO]2[O2-], where k3 = 3 - 6 x 10(8) M-2s-1. In addition, the reaction of GSNO with O2- generated a strong oxidant as an intermediate capable of oxidizing dihydrorhodamine in the absence of the apparent generation of NO. We conclude that O2- may act as a physiological modulator of S-nitrosation reactions by directly promoting the decomposition of S-nitrosothiols.

Catalase↗

PubMed1998

Differential regulation of extracellular signal-regulated kinase and nuclear factor-kappa B signal transduction pathways by hydrogen peroxide and tumor necrosis factor.

Reactive oxygen metabolites are increasingly recognized for their ability to stimulate signal transduction pathways. This is important because these oxidants are frequently generated at sites of inflammation. However, little is known about the manner in which reactive oxygen species may selectively stimulate distinct signaling pathways. We have examined this question by stimulating mesothelial cells with hydrogen peroxide (H2O2) as a model oxidant stimulus. The response to H2O2 was examined by measuring the activation of the extracellular signal-regulated kinase (ERK1/2) and the nuclear factor-kappa B (NF-kappa B) signal transduction pathways. We found that H2O2 stimulated activity of the ERK1/2 pathway in a dose- and time-dependent manner. The ability of H2O2 to activate ERK1/2 was similar to that found with tumor necrosis factor (TNF) stimulation. The oxidant effect was inhibited by various reactive oxygen scavengers. An inhibitor of mitogen-activated protein kinase/extracellular signal-regulated kinase, the upstream kinase that activates ERK1/2, inhibited the oxidant effect. The superoxide anion (O2-) also stimulated ERK1/2 activity. In contrast, H2O2 did not stimulate proteolysis of I kappa B-alpha and induced only a small degree of NF-kappa B nuclear translocation. Stimulation of the cells with O2- also induced a minimal degree of NF-kappa B activation. TNF was a potent stimulus for I kappa B-alpha proteolysis and NF-kappa B activation, demonstrating that the cells did have a functional NF-kappa B pathway. These results suggest that oxidants may selectively stimulate certain pathways, thereby preserving some specificity of the signaling process. Furthermore, different cell types and distinct signaling pathways within cells may demonstrate unique profiles in the manner in which they respond to oxidant stimulation.

Animals↗

PubMed1998

The reaction of S-nitrosoglutathione with superoxide.

Catalase↗

PubMed1998

The cytotoxicity of nitroxyl: possible implications for the pathophysiological role of NO.

In addition to the broad repertoire of regulatory functions nitric oxide (NO) serves in mammalian physiology, the L-arginine:NO pathway is also involved in numerous pathophysiological mechanisms. While NO itself may actually protect cells from the toxicity of reactive oxygen radicals in some cases, it has been suggested that reactive nitrogen oxide species formed from nitric oxide synthase (NOS) can be cytotoxic. In addition to NO, the one electron reduction product NO- has been proposed to be formed from NOS. We investigated the potential cytotoxic role of nitroxyl (NO-), using the nitroxyl donor Angelis's salt, (AS; sodium trioxodinitrate, Na2N2O3) as the source of NO-. As was found to be cytotoxic to Chinese hamster V79 lung fibroblast cells over a concentration range of 2-4 mM. The presence of equimolar ferricyanide (Fe(III)-(CN6)3-), which converts NO- to NO, afforded dramatic protection against AS-mediated cytotoxicity. Treatment of V79 cells with L-buthionine sulfoximine to reduce intracellular glutathione markedly enhanced AS cytotoxicity, which suggests that GSH is critical for cellular protection against the toxicity of NO-. Further experiments showed that low molecular weight transition metal complexes associated with the formation of reactive oxygen species are not involved in AS-mediated cytotoxicity since metal chelators had no effect. However, under aerobic conditions, AS was more toxic than under hypoxic conditions, suggesting that oxygen dramatically enhanced AS-mediated cytotoxicity. At a molecular level, AS exposure resulted in DNA double strand breaks in whole cells, and this effect was completely prevented by coincubation of cells with ferricyanide or Tempol. The data in this study suggest that nitroxyl may contribute to the cytotoxicity associated with an enhanced expression of the L-arginine:NO pathway under different biological conditions.

Animals↗

PubMed1998

Effect of nitric oxide on hemoprotein-catalyzed oxidative reactions.

Hemoglobin or myoglobin-catalyzed oxidation reactions have been suggested to initiate and/or exacerbate tissue injury associated with a variety of pathological conditions including post-ischemic tissue injury, hemorrhagic disorders, and chronic inflammation. In the present study, we investigated what effect different fluxes of nitric oxide (NO) have on hemoprotein-catalyzed oxidation reactions in vitro. The hypoxanthine/xanthine oxidase system was used to generate both O2- and H2O2, whereas the spontaneous decomposition of the spermine/NO adduct was used to generate NO at a known and constant rate. We assessed the ability of myoglobin (Mb) or hemoglobin (Hb) to oxidize dihydrorhodamine (DHR) to rhodamine (RH) in the presence of O2-/H2O2 and/or NO. In the presence of a constant flux of O2- and H2O2 (1 nmol/min each), 500 nM MetMb (Fe3+) stimulated DHR oxidation from normally undetectable levels to approximately 35 microM. This oxidation reaction was inhibited by catalase but not SOD, suggesting the formation of the ferryl-hemoprotein adduct (Fe4+). Equimolar fluxes of O2-, H2O2, and NO increased further DHR oxidation to approximately 50 microM. The 15 microM increase in DHR oxidation was independent of heme concentration and was inhibited by SOD. This suggested that equal fluxes of O2- and NO interact to yield a potent oxidant such as peroxinitrite (OONO-) which together with Mb-Fe4+ oxidizes DHR. Further increases in NO fluxes significantly inhibited DHR oxidation (80%) via the NO-dependent inhibition of Mb-Fe4+ formation. Additional studies using methemoglobin (Hb-Fe3+)-catalyzed oxidative reactions yielded virtually identical results. We conclude that in the presence of a hemoprotein such as myoglobin or hemoglobin, NO may promote or inhibit oxidation reactions depending upon the relative fluxes of O2-, H2O2, and NO.

Catalase↗

PubMed1998

Modulation of leukocyte-endothelial interactions by reactive metabolites of oxygen and nitrogen: relevance to ischemic heart disease.

Ischemia and reperfusion (I/R) are thought to play an important role in the pathophysiology of ischemic diseases of the heart. It is now well appreciated that leukocyte-endothelial cell interactions are important determinants for I/R-induced microvascular injury and dysfunction. There is a growing body of experimental data to suggest that reactive metabolites of oxygen and nitrogen are important physiological modulators of leukocyte-endothelial cell interactions. A number of investigators have demonstrated that I/R enhances oxidant production within the microcirculation resulting in increases in leukocyte adhesion and transendothelial cell migration. Several other studies have shown that exogenous nitric oxide (NO) donors may attenuate leukocyte and platelet adhesion and/or aggregation in a number of different inflammatory conditions including I/R. The objective of this review is to discuss the physiological chemistry of reactive metabolites of oxygen and nitrogen with special attention given to those interactions that may modulate leukocyte-endothelial cell interactions, provide an overview of the evidence implicating reactive metabolites of oxygen and nitrogen as modulators of leukocyte-endothelial cell interactions in vivo, and discuss how these mechanisms may be involved in the pathophysiology of ischemic heart disease.

Animals↗