Yukio Toki

Angiographic no-reflow phenomenon as a predictor of adverse long-term outcome in patients treated with percutaneous transluminal coronary angioplasty for first acute myocardial infarction

OBJECTIVES We sought to elucidate the long-term prognostic importance of angiographic no-reflow phenomenon after percutaneous transluminal coronary angioplasty (PTCA) for acute myocardial infarction (AMI). BACKGROUND Angiographic no-reflow phenomenon, a reduced coronary antegrade flow (Thrombolysis in Myocardial Infarction [TIMI] flow grade < or =2) without mechanical obstruction after recanalization, predicts poor left ventricular (LV) functional recovery and survival in the early phase of AMI. We hypothesized that angiographic no-reflow phenomenon also predicts long-term clinical outcome. METHODS We studied 120 consecutive patients with their first AMI treated by PTCA without flow-restricting lesions. The patients were classified as either no-reflow (n = 30) or reflow (TIMI-3) (n = 90) based on post-PTCA cineangiograms to follow up (5.8 +/- 1.2 years) for cardiac death and nonfatal events. RESULTS Patients with no-reflow had congestive heart failure (p < 0.0001), malignant arrhythmia (p = 0.038), and cardiac death (p = 0.002) more often than did those with reflow. Kaplan-Meier curves showed lower cardiac survival and cardiac event-free survival (p < 0.0001) in patients with no-reflow than in those with reflow. Multivariate analyses disclosed that no-reflow phenomenon was an independent predictor of long-term cardiac death (relative risk [RR] 5.25, 95% confidence interval [CI] 1.85 to 14.9, p = 0.002) and cardiac events (RR 3.71, 95% CI 1.79 to 7.69, p = 0.0004). At follow-up, survivors with no-reflow had higher end-diastolic and end-systolic LV volume indices and plasma brain natriuretic peptide levels, and lower LV ejection fractions (p = 0.0002, p < 0.0001, p = 0.002, p < 0.0001, respectively) than did those with reflow, indicating that no-reflow may be involved in LV remodeling. CONCLUSIONS Angiographic no-reflow phenomenon strongly predicts long-term cardiac complications after AMI; these complications are possibly associated with LV remodeling.

Long-term inhibition of NO synthesis promotes atherosclerosis in the hypercholesterolemic rabbit thoracic aorta. PGH2 does not contribute to impaired endothelium-dependent relaxation.

We examined whether prostaglandin (PG) H2, as an endothelium-dependent contracting factor, or the disturbed production of endothelium-derived relaxing factor, impairs endothelium-dependent relaxation and whether long-term inhibition of nitric oxide (NO) synthesis aggravates atherosclerosis in hypercholesterolemic rabbits. Male New Zealand White rabbits were fed one of the following diets: (1) standard chow; (2) 2% cholesterol-supplemented chow; (3) standard chow with 80 micrograms/mL N omega-nitro-L-arginine methylester (L-NAME), an NO synthetase inhibitor, in their drinking water; or (4) 2% cholesterol-supplemented chow with 80 or 160 micrograms/mL L-NAME in their drinking water. The rabbits were fed these diets for 8 or 12 weeks. Then aortic rings were obtained, and changes in isometric tension were recorded. Intimal atherosclerotic areas of the thoracic aortas were subsequently measured by planimetry. The cholesterol-supplemented diet significantly impaired endothelium-dependent aortic relaxation to acetylcholine. Pretreatment with the thromboxane A2/PGH2 receptor antagonist ONO-3708 did not reverse this impaired response. Vessels from both normocholesterolemic and hypercholesterolemic rabbits given L-NAME showed more impaired endothelium-dependent relaxation than those from their dietary counterparts not given L-NAME. Morphometric analysis revealed marked enlargement of intimal atherosclerotic areas in aortas from L-NAME-treated hypercholesterolemic rabbits compared with those from untreated hypercholesterolemic rabbits. These findings suggest that PGH2 does not contribute to impaired endothelium-dependent relaxation and that long-term administration of L-NAME promotes atherosclerosis by inhibition of NO synthesis in the hypercholesterolemic rabbit thoracic aorta.

Melatonin scavenges hydroxyl radical and protects isolated rat hearts from ischemic reperfusion injury.

During postischemic reperfusion, free radicals are produced and have deleterious effects in isolated rat hearts. We investigated whether melatonin (MEL) reduces the production of hydroxyl radical (*OH) in the effluent and aids in recovery of left ventricular (LV) function. Hearts were subjected to 30 min of ischemia followed by 30 min of reperfusion. Salicylic acid (SAL) was used as the probe for *OH, and its derivatives 2,5- and 2,3-dihydroxybenzoic acid (DHBA) were quantified using HPLC. In addition, thiobarbituric acid reactive substances (TBARS) in the myocardium was measured. Plateaus in the measurement of 2,5- and 2,3-DHBA were seen from 3 to 8 min after reperfusion in each group. The group that received 100 microM MEL+ SAL had significantly reduced amounts of 2,5- and 2,3-DHBA by multiple folds, compared to the SAL group. TBARS was significantly decreased in the 100 microM MEL group (1.20+/-0.36 vs 1.85+/-0.10 micromol/g of drug-free group, p<0.001). More importantly, the 100 microM MEL group significantly recovered in LV function (LV developed pressure, +dp/dt, and -dp/dt; 63.0%, 60.3%, and 59.4% in the 100 microM MEL group; 30.2%, 29.7%, and 31.5% in the drug-free group, respectively; p<0.05). Duration of ventricular tachycardia or ventricular fibrillation significantly decreased in the 100 microM MEL group (100 microM MEL, 159+/-67 sec; drug-free, 1244+/-233 sec; p<0.05). As a result of scavenging *OH and reducing the extent of lipid peroxidation, MEL is an effective agent for protection against postischemic reperfusion injury.

Improved insulin sensitivity by bezafibrate in rats: relationship to fatty acid composition of skeletal-muscle triglycerides.

We investigated the effect of the lipid-lowering agent, bezafibrate, on insulin sensitivity in a dietary model of insulin resistance. Male Sprague-Dawley rats were divided into four groups: control group, administered a standard diet; high-fructose group, given a 40% fructose diet; high-fructose plus lard group, given a 40% fructose diet with 7% lard; and bezafibrate group, given a 40% fructose plus 7% lard diet with 10 mg · kg−1 · day−1 of oral bezafibrate. Insulin action was assessed after 2 weeks with a steady-state plasma glucose (SSPG) level. The fatty acid (FA) composition of skeletal-muscle triglycerides was also determined. A higher SSPG level (20.9 ± 0.9 vs. 16.5 ±1.1 mmol/l in the control group, P < 0.05) as well as a higher systolic blood pressure (120 ± 2 vs. 101 ± 2 mmHg, P < 0.01) was observed in the high-fructose plus lard group, but not in the high-fructose group. These changes were prevented by bezafibrate administration. The FA composition of skeletal-muscle triglycerides demonstrated a higher percentage of saturated and monounsaturated FAs (P < 0.01) and a lower percentage of polyunsaturated FAs (P < 0.01) in the highfructose plus lard group versus the control group. These changes were consistent with differences in the dietary intake of FAs. Bezafibrate virtually normalized the FA composition in the high-fructose plus lard group. The ratio of C20:4 to C20:3, an index of ∆5 desaturase activity, was significantly higher in the bezafibrate group versus the high-fructose plus lard group (8.60 ± 0.76 vs. 2.04 ± 0.27, P < 0.01). In conclusion, the dietary FA composition was closely related to insulin resistance in rats fed 40% fructose. Bezafibrate increased ∆5 desaturase activity. Such action may contribute to the improvement of insulin sensitivity.

Melatonin, a pineal hormone with antioxidant property, protects against adriamycin cardiomyopathy in rats

Adriamycin (ADR) is a potent, broad-spectrum chemotherapeutic agent whose clinical use is limited by its cardiotoxicity. Since the pathogenesis of ADR-induced cardiomyopathy may involve free radicals and lipid peroxidation, the antioxidant, melatonin (MEL) may protect against toxic effects of ADR. We therefore tested this hypothesis using a rat model of ADR-induced cardiomyopathy. Sprague-Dawley rats were given ADR (cumulative dose, 15 mg/kg), MEL (cumulative dose, 84 mg/kg), ADR+MEL, ADR plus probucol (PRB, cumulative dose, 90 mg/kg), or vehicle alone, according to known regimens. The rats were maintained for 3 weeks following treatment, after which their cardiac performance was measured. Following sacrifice, their myocardial ultrastructure was examined, and their myocardial lipid peroxidation was assessed. Mortality was observed only in rats treated with ADR alone. When compared to control rats, surviving rats in the ADR group showed significant decreases in ratio of heart to body weight, arterial pressure, and left ventricular fractional shortening as well as a significant accumulation of ascites. The amount of myocardial thiobarbituric acid reactive substances was significantly higher in ADR-treated than in control rats. Both antioxidants, MEL and PRB, significantly prevented these ADR-induced changes. Electron microscopic examination revealed myocardial lesions indicative of ADR-induced cardiomyopathy in the ADR-treated rats. In contrast, treatment of these rats with MEL or PRB preserved myocardial ultrastructure. By preventing lipid peroxidation, MEL may be highly effective in protecting against ADR-induced cardiomyopathy.

Role of Prostaglandin H2 as an Endothelium-Derived Contracting Factor in Diabetic State

This study investigated the possible involvement of prostaglandin H2, an acetylcholine-induced endothelium-derived contracting factor in rat aorta, in the development of abnormality of the vasculature in diabetes. Rings of thoracic aorta were prepared from control Wistar-Kyoto and STZ-induced diabetic rats to examine the changes in isometric tension. In 10−7 M norepinephrine-precontracted rings, acetylcholine induced relaxations, which were significantly impaired in diabetic rats. Inhibition of thromboxane A2-prostaglandin H2 receptors with ONO-3708 (10−6) M) prevented the development of the impairment of relaxation in diabetic rats. Thromboxane A2 synthesis inhibition with OKY-046 (10−5 M) did not affect the acetylcholine-induced relaxation in both control and diabetic rats. In aortic rings under resting tension, acetylcholine induced a contraction that was greater in diabetic than control rats, when the nitric oxide production was inhibited by NGxs-nitro-L-arginine methylester (10−4 M). This acetylcholine-induced contraction was observed only in the rings with intact endothelium and was completely abolished by ONO-3708 (10−6 M). The concentration of 6-keto-prostaglandin F1α in the solution bathing diabetic rat aortic rings increased significantly after acetylcholine (10−5 M) administration. Prostacyclin (10−9 · 10−6 M) did not induce contractions at all. Prostacyclin is unlikely to mediate contractions because of its low contractile potency. These findings suggest that the impairment of acetylcholine-induced relaxation in the diabetic state is not caused by the diminished production of an endothelium-derived relaxing factor or nitric oxide but rather by the increased endothelium-derived contracting factor or prostaglandin H2, which may be responsible for abnormalities of the vasculature in diabetes.

Protective effects of carvedilol against doxorubicin-induced cardiomyopathy in rats.

Carvedilol (CAR) is a vasodilating beta-blocker which also has antioxidant properties. CAR produces dose-related reduction in mortality in patients with congestive heart failure. In the present study, we tested the hypothesis that CAR protects against doxorubicin (DOX)-induced cardiomyopathy in rats. Sprague-Dawley rats were treated with DOX, CAR, CAR+DOX, or atenolol (ATN)+DOX. DOX (cumulative dose, 15 mg/kg) was administered intraperitoneally, and CAR (30 mg/kg daily) or ATN (150 mg/kg daily) was administered orally. Three weeks after the completion of these treatments, cardiac performance and myocardial lipid peroxidation were assessed. Mortality was observed in the DOX (25%) and ATN+DOX (12.5%) groups. Compared with control rats, DOX significantly decreased systolic blood pressure (104+/-4 vs. 120+/-4 mmHg, P<0.05) and left ventricular fractional shortening (38.8+/-3.1 vs. 55.4+/-1.3%, P<0.01), and resulted in a significant accumulation of ascites (14.4+/-4.9 vs. 0 ml, P<0.01). CAR significantly prevented the cardiomyopathic changes caused by DOX, while ATN did not. The myocardial thiobarbituric acid reactive substances (TBARS) content was significantly higher in DOX-treated rats than in control rats (80.4+/-7.1 vs. 51.5+/-1.2 nmol/g heart, p<0.01). CAR prevented the increase in TBARS content (48.8+/-3.0 nmol/g heart, P<0.01 vs. DOX group), whereas ATN had no significant effect (74.3+/-5.2 nmol/g heart). CAR also significantly prevented the increase in both myocardial and plasma cholesterol concentrations caused by DOX. These data indicate that CAR protects against DOX-induced cardiomyopathy and that this effect may be attributed to the antioxidant and lipid-lowering properties of CAR, not to its beta-blocking property.

Prostacyclin Synthase Gene Transfer Accelerates Reendothelialization and Inhibits Neointimal Formation in Rat Carotid Arteries After Balloon Injury

Prostacyclin (PGI2), a metabolite of arachidonic acid, has the vasoprotective effects of vasodilation, anti-platelet aggregation, and inhibition of smooth muscle cell proliferation. We hypothesized that an overexpression of endogenous PGI2 may accelerate the recovery from endothelial damage and inhibit neointimal formation in the injured artery. To test this hypothesis, we investigated in vivo transfer of the PGI2 synthase (PCS) gene into balloon-injured rat carotid arteries by a nonviral lipotransfection method. Seven days after transfection, a significant regeneration of endothelium was observed in the arteries transfected with a plasmid carrying the rat PCS gene (pCMV-PCS), but little regeneration was seen in those with the control plasmid carrying the lacZ gene (pCMV-lacZ) (percent luminal circumference lined by newly regenerated endothelium: 87. 1+/-6.9% in pCMV-PCS-transfected vessels and 6.9+/-0.2% in pCMV-lacZ vessels, P<0.001). BrdU staining of arterial segments demonstrated a significantly lower incorporation in pCMV-PCS-transfected vessels (7. 5+/-0.3% positive nuclei in vessel cells) than in pCMV-lacZ (50. 7+/-9.6%, P<0.01). Moreover, 2 weeks after transfection, the PCS gene transfer resulted in a significant inhibition of neointimal formation (88% reduction in ratio of intima/media areas), whereas medial area was similar among the groups. Arterial segments transfected with pCMV-PCS produced significantly higher levels of 6-keto-PGF1alpha, the main metabolite of PGI2, compared with the segments transfected with pCMV-lacZ (10.2+/-0.55 and 2.1+/-0.32 ng/mg tissue for pCMV-PCS and pCMV-placZ, P<0.001). In conclusion, this study demonstrated that an in vivo PCS gene transfer increased the production of PGI2 and markedly inhibited neointimal formation with accelerated reendothelialization in rat carotid arteries after balloon injury.

Correlation with blood pressure of the acetylcholine-induced endothelium-derived contracting factor in the rat aorta.

To examine a relation between the production of acetylcholine-induced endothelium-derived contracting factor and an increase in blood pressure, endothelium-dependent contraction and relaxation were evaluated by measuring the isometric tension of aortic rings from spontaneously hypertensive rats and Wistar-Kyoto rats at 5, 10, 20, and 30 weeks of age. In norepinephrine-precontracted rings, acetylcholine (10(-8) to 10(-5) M)-induced relaxations diminished at the doses of 10(-6) to 10(-5) M in both strains except at 5 weeks of age. Treatment with a thromboxane A2/prostaglandin H2 antagonist (ONO-3708) prevented this reduction in acetylcholine-induced relaxations in both strains and induced dose-dependent relaxations, which were completely inhibited by treatment with a nitric oxide inhibitor, NG-nitro-L-arginine methyl ester. In aorta treated with NG-nitro-L-arginine methyl ester without precontraction, acetylcholine induced dose-dependent contractions, which were greater in spontaneously hypertensive rats than in Wistar-Kyoto rats. These acetylcholine-induced contractions, which were observed only in rings with endothelium, were completely inhibited by treatment with ONO-3708 but not with a thromboxane A2 synthetase inhibitor (OKY-046). There was a statistically significant correlation between the acetylcholine-induced contractions and blood pressure. Release of 6-ketoprostaglandin F1 alpha by acetylcholine from the aorta was greater in spontaneously hypertensive rats. In vivo administration of another thromboxane A2/prostaglandin H2 antagonist (ONO-8809) (10 or 30 micrograms per body per day) for 3 weeks (5-8 weeks of age) did not affect blood pressure in either rat strain.(ABSTRACT TRUNCATED AT 250 WORDS)

Altered gene expression of prostacyclin synthase and prostacyclin receptor in the thoracic aorta of spontaneously hypertensive rats

OBJECTIVE The aim of this study was to evaluate the possible role of prostacyclin (PGI2) in the pathogenesis of hypertension in spontaneously hypertensive rats (SHR). METHODS Measurement of mRNA and protein levels of PGH synthase (PGHS)-1, PGI2 synthase and the PGI2 receptor, in the thoracic aorta was performed in SHR aged 5, 10, 20, and 40 weeks old and in age-matched normotensive Wistar-Kyoto (WKY) rats with a competitive polymerase chain reaction method and immunoblotting. Aortic production of 6-keto-PGF1 alpha, the main metabolite of PGI2, was also measured. RESULTS Compared with age-matched WKY rats, PGHS-1 mRNA and protein levels in the thoracic aorta of SHR increased with age, reaching three- and twofold higher than WKY rats at 40 weeks old, respectively. PGI2 synthase mRNA and protein levels in SHR were significantly higher than in WKY rats at 20 and 40 weeks old. In contrast, PGI2 receptor mRNA levels in SHR were consistently lower than in WKY rats at all ages. CONCLUSIONS These results provide evidence that hypertension elicits alterations in levels of arachidonic acid metabolites, including PGH2 and PGI2. They also suggest that the decreased expression of PGI2 receptor mRNA in prehypertensive SHR could be one of the causes of hypertension in SHR.