Hiroaki Kosaka

Reduced Plasma Concentrations of Nitrogen Oxide in Individuals With Essential Hypertension

Patients with essential hypertension exhibit blunted endothelium-dependent vasodilator responses, which may be largely attributable to reduced bioactivity of nitric oxide (NO). Therefore, we measured the end product of NO, nitrate plus nitrite (nitrogen oxide), and examined the relationship between the degree of hypertension and plasma nitrate plus nitrite levels in patients with essential hypertension. The combined plasma concentration of nitrate plus nitrite, end products of NO metabolism, was reduced in individuals with essential hypertension relative to that in control subjects (15.7+/-1.1 versus 22.8+/-1.4 mmol x L(-1), P<.001); individuals with borderline hypertension showed values that were intermediate between those of the other two groups (18.2+/-1.2 mmol x L(-1), P<.001). The plasma nitrogen oxide concentration showed significant inverse correlations with both systolic and diastolic blood pressures. The basal concentration of nitrogen oxide in the plasma was reduced, at least in the peripheral circulation, in individuals with essential hypertension.

Stoichiometry of the reaction of oxyhemoglobin with nitrite

During the reaction of oxyhemoglobin (HbO2) with nitrite, the concentration of residual nitrite, nitrate, oxygen, and methemoglobin (Hb+) was determined successively. The results obtained at various pH values indicate the following stoichiometry for the overall reaction: 4HbO2 + 4NO2- 4H+ leads to 4Hb+ + 4NO3- + O2 + 2H2 O (Hb denotes hemoglobin monomer). NO2- binds with methemoglobin noncooperatively with a binding constant of 340 M-1 at pH 7.4 and 25 degrees C. Thus, the major part of Hb+ produced is aquomethemoglobin, not methemoglobin nitrite, when less than 2 equivalents of nitrite is used for the oxidation.

Detection of nitric oxide production in lipopolysaccharide-treated rats by ESR using carbon monoxide hemoglobin

Release of nitric oxide (NO), from macrophages activated with E. coli lipopolysaccharide (LPS) and endothelial cells, has been proposed using chemiluminescence and spectrophotometry. However these methods can not distinguish NO from NO2-. The present study was aimed to prove in vivo production of NO, by ESR using CO-hemoglobin (HbCO) as a trapping agent of NO in the peritoneal cavity of rats treated with LPS. We detected a broad signal in the recovered HbCO solution. Inositol hexaphosphate induced a three-line hyperfine structure, characteristic of NO-hemoglobin (HbNO). In the arterial blood, ESR signal of HbNO with faint hyperfine structure was detected. NG-Monomethyl-L-arginine inhibited the formation of HbNO. HbNO was not detected in the peritoneal cavity of the LPS-untreated rat given i.p. both NO2- and HbCO. HbNO was, therefore, derived from NO, not from NO2-. These results show that free NO is produced in vivo by the stimulation of LPS.

Beneficial Effects of Inhibition of Angiotensin-Converting Enzyme on Ischemic Myocardium During Coronary Hypoperfusion in Dogs

BACKGROUND Angiotensin-converting enzyme (ACE) produces angiotensin II, causing vasoconstriction of coronary arteries and reduction of coronary blood flow. The present study was undertaken to test the hypothesis that an ACE inhibitor increases coronary blood flow and improves myocardial metabolic and contractile functions of ischemic myocardium. METHODS AND RESULTS In 65 open-chest dogs, the left anterior descending coronary artery was perfused through an extracorporeal bypass tube from the left carotid artery. When cilazaprilat (3 micrograms/kg per minute) was infused into the bypass tube for 10 minutes after reduction of coronary blood flow due to partial occlusion of the bypass tube, coronary blood flow increased from 30 +/- 1 to 43 +/- 2 mL/100 g per minute despite there being no changes in coronary perfusion pressure (43 +/- 1 mm Hg). The ratio of myocardial endocardial flow to epicardial flow increased during an infusion of cilazaprilat. Both fractional shortening and lactate extraction ratio of the perfused area were increased (fractional shortening: 4.1 +/- 0.6% to 8.9 +/- 0.6%, P < .001; lactate extraction ratio: -55.7 +/- 3.3% to -36.7 +/- 3.9%, P < .001). During an infusion of cilazaprilat, the bradykinin concentration of coronary venous blood was markedly increased. The increased coronary blood flow due to cilazaprilat was attenuated by HOE-140 (an inhibitor of bradykinin receptors; coronary blood flow: 35 +/- 2 mL/100 g per minute), and by N omega-nitro-L-arginine methyl ester (an inhibitor of nitric oxide synthase; coronary blood flow: 34 +/- 2 mL/100 g per minute). Intracoronary administration of bradykinin mimicked the beneficial effects of cilazaprilat. Cyclic GMP content of the coronary artery was increased by cilazaprilat compared with the untreated condition in the ischemic myocardium. In the denervated hearts, the increased coronary blood flow due to cilazaprilat was not attenuated. On the other hand, CV11974, an inhibitor of angiotensin II receptors, slightly increased coronary blood flow to 34 +/- 2 from 30 +/- 1 mL/100 g per minute. CONCLUSIONS We conclude that an inhibitor of ACE can increase coronary blood flow and ameliorate myocardial ischemia, primarily due to accumulation of bradykinin and production of nitric oxide from the ischemic myocardium. Inhibition of angiotensin II production due to inhibition of ACE partially contributes to coronary vasodilation in the ischemic myocardium.

Elevation of plasma nitric oxide end products during focal cerebral ischemia and reperfusion in the rat

We investigated the alterations in the stable end products of nitric oxide, i.e., nitrate and nitrite, in the plasma during and after rat focal cerebral ischemia by an automated procedure based on the Griess reaction. At 2 h of middle cerebral artery (MCA) occlusion, plasma nitrate/nitrite levels were significantly higher (53 ± 8 μM, mean ± SD, n = 5, p < 0.05) than in rats with sham operation (36 ± 9 μM, n = 5), and were mildly elevated at 4 h of MCA occlusion (42 ± 9 μM, n = 5, n.s.). At 30 min of reperfusion after 2 h of MCA occlusion, plasma nitrate/nitrite levels were more markedly elevated (72 ± 7 μM, n = 5, p < 0.01 vs. sham operation), but were moderately elevated at 2 h of reperfusion after 2 h of MCA occlusion (61 ± 10 μM, n = 5, p < 0.05). Plasma nitrite levels were not changed during these experimental periods. Administration of 20 mg/kg of NG-nitro-l-arginine methyl ester (l-NAME) significantly decreased plasma nitrate/nitrite as well as nitrite at 30 min of reperfusion after 2 h of MCA occlusion (n = 5), but 2 mg/kg of l-NAME did not (n = 3). The effect of 20 mg/kg of l-NAME on plasma nitric oxide end products was reversed by the simultaneous administration of 200 mg/kg of l-arginine (n = 3), but not d-arginine (n = 3). The present study suggests that the l-arginine-nitric oxide pathway is activated during acute cerebral ischemia and reperfusion.

Increased Release of NO During Ischemia Reduces Myocardial Contractility and Improves Metabolic Dysfunction

BACKGROUND We have reported that myocardial ischemia increases nitric oxide (NO) production. Several lines of evidence suggest that NO reduces myocardial contraction. Therefore, we tested whether endogenous NO decreases the inotropic response of the ischemic myocardium and whether endogenous NO is beneficial in the metabolic function of ischemic myocardium. METHODS AND RESULTS The left anterior descending coronary artery was perfused with blood from the left carotid artery in 72 dogs. An infusion of NG-nitro-L-arginine methyl ester (L-NAME), an inhibitor of NO synthase, did not affect fractional shortening (FS) under nonischemic conditions. After reduction of perfusion pressure so that coronary blood flow decreased to 60% of the control value, FS of the perfused area decreased, and intravenous infusion of isoproterenol increased FS. Before and during intravenous infusion of isoproterenol under conditions of coronary hypoperfusion, FS was significantly increased in the L-NAME group compared with the untreated group. Both lactate extraction ratio and the pH in coronary venous blood were significantly lower in the L-NAME-treated group than in the untreated group during coronary hypoperfusion. Infusion of L-arginine prevented the effects of L-NAME in the ischemic myocardium. CONCLUSIONS These results indicate that endogenous NO reduces myocardial contractile function and improves myocardial metabolic function in the ischemic heart. The myocardial energy-sparing effect as well as coronary vasodilation due to NO may be beneficial to the ischemic myocardium.

Mechanism of autocatalytic oxidation of oxyhemoglobin by nitrite an intermediate detected by electron spin resonance

Oxidation of oxyhemoglobin by nitrite is characterized by the presence of a lag phase followed by the autocatalysis. Just before the autocatalysis begins, an asymmetric ESR signal is detected which is similar to that of the methemoglobin radical generated from methemoglobin and H2O2 in shape, g value (2.005), peak-to-peak width (18 G) and other properties, except the difference in the dependence on temperature. Generation of H2O2 is indicated by the prolongation of the lag phase by the addition of catalase. On the other hand, the oxidation is modified by neither superoxide dismutase nor Nitroblue tetrazolium. The oxidation is prolonged in the presence of KCN. The present results indicate a free-radical mechanism for the oxidation in which the asymmetric radical catalyzes the formation of NO2 from NO2- by a peroxidase action and NO2 oxidizes oxyhemoglobin in the autocatalytic phase.

Roles of NO and Ca2+-activated K+ channels in coronary vasodilation induced by 17beta-estradiol in ischemic heart failure.

Estrogen induces the generation of nitric oxide (NO) and produces coronary vasodilation by opening the Ca2+‐activated K+ (KCa) channels. The hypothesis that 17β‐estradiol produces NO and activates KCa channels during coronary hypoperfusion was investigated. In open‐chest dogs, the left anterior descending coronary artery was perfused through an extracorporeal bypass tube from the left carotid artery. 17β‐Estradiol was infused into the bypass tube for 20 min after coronary blood flow was reduced by partial occlusion of the bypass tube. 17β‐Estradiol increased the difference in NO concentrations between the coronary venous and arterial blood as well as coronary blood flow. The lactate extraction ratio and pH of coronary venous blood were both also increased by 17β‐estradiol, indicating a reduction in myocardial anaerobic metabolism. Whereas the increase in the coronary arteriovenous difference in NO concentration was completely attenuated by NG‐nitro‐L‐arginine methyl ester (L‐NAME, an inhibitor of NO synthase), the increase in coronary blood flow induced by 17β‐estradiol was only partially attenuated by L‐NAME. The combination of L‐NAME and iberiotoxin (a blocker of high‐conductance KCa channels) completely abolished the coronary vasodilatory effect of 17β‐estradiol. The data indicate that during coronary hypoperfusion in canine hearts, 17β‐estradiol increases coronary blood flow and improves metabolic dysfunction by increasing NO release and opening KCa channels.—Node, K., Kitakaze, M., Kosaka, H., Minamino, T., Sato, H., Kuzuya, T., Hori, M. Roles of NO and Ca2+‐activated K+ channels in coronary vasodilation induced by 17β‐estradiol in ischemic heart failure. FASEB J. 11, 793–799 (1997)

Statins induce S1P1 receptors and enhance endothelial nitric oxide production in response to high-density lipoproteins

Sphingosine 1‐phosphate (S1P) is a serum‐borne naturally occurring sphingolipid, specifically enriched in high‐density lipoprotein (HDL) fractions. S1P binds to G‐protein‐coupled S1P1 receptors to activate endothelial NO synthase (eNOS) in vascular endothelial cells. We explored whether and how statins, 3‐hydroxy‐3‐methylglutaryl coenzyme A (HMG‐CoA) reductase inhibitors, modulate expression of S1P1 receptors and endothelial responses for subsequent stimulation with S1P or with HDL.

Reversible Reduction in Plasma Concentration of Nitric Oxide Induced by Cigarette Smoking in Young Adults

The concentration of nitrate plus nitrite, metabolic end products of nitric oxide, in serum prepared from systemic venous blood was significantly (p <0.001) decreased in both heavy (14.5 +/- 1.3 micromol/L) and moderate (17.6 +/- 2.3 micromol/L) smokers relative to that in nonsmokers (22.6 +/- 0.4 micromol/L).