Yi-Ping Sun

Passive smoking increases experimental atherosclerosis in cholesterol-fed rabbits.

OBJECTIVES We evaluated the influence of passive smoking on experimental atherosclerosis in cholesterol-fed rabbits. BACKGROUND Exposure to environmental tobacco smoke (ETS) has been epidemiologically linked to death from ischemic heart disease in nonsmokers. METHODS New Zealand male rabbits were randomly divided into three groups after 2 weeks of a 0.3% cholesterol diet. Sixteen rabbits were exposed to a high and 16 rabbits to a low dose of ETS; 32 rabbits located in another room served as an unexposed control group. After 10 weeks of ETS exposure, all rabbits were killed, and the percent of aortic and pulmonary artery endothelial surfaces covered by lipid lesions was measured by staining and planimetry. RESULTS Average air nicotine, carbon monoxide and total particulate concentrations were 1,040 micrograms/m3, 60.2 ppm and 32.8 mg/m3 for the high dose ETS group, 30 micrograms/m3, 18.8 ppm and 4.0 mg/m3 for the low dose ETS group and < 1 microgram/m3, 3.1 ppm and 0.13 mg/m3 for the control group. The percent atherosclerotic involvement of the aorta and pulmonary artery increased significantly with ETS exposure (for the aorta, 30 +/- 19% [mean +/- SD] for the control group, 36 +/- 14% for the low dose ETS group and 52 +/- 21% for the high dose ETS group, p < 0.001; for the pulmonary artery, 22 +/- 15% for the control group, 29 +/- 25% for the low dose ETS group, and 45 +/- 12% for the high dose ETS group, p < 0.001). Bleeding time was significantly shorter in the two ETS groups than in the control group (86 +/- 17 vs. 68 +/- 15, 68 +/- 18 s, p < 0.001). There were no significant differences in serum triglycerides, cholesterol and high density lipoprotein cholesterol at the end of the study. CONCLUSIONS Environmental tobacco smoke affects platelet function and increases aortic and pulmonary artery atherosclerosis. This increase of atherosclerosis was independent of changes in serum lipids and exhibited a dose-response relation. These results are consistent with data from epidemiologic studies demonstrating that ETS increases the risk of death due to heart disease.

Exposure to environmental tobacco smoke increases myocardial infarct size in rats.

BACKGROUND Exposure to environmental tobacco smoke (ETS) has been epidemiologically linked to death from ischemic heart disease in nonsmokers. In this study, we evaluated the influence of 3 days, 3 weeks, and 6 weeks of ETS exposure on myocardial infarct size in a rat ischemia/reperfusion model. METHODS AND RESULTS Sprague-Dawley rats exposed to ETS (four Marlboro cigarettes per 15 minutes, 6 hours per day, 5 days per week) for 3 days (n = 24), 3 weeks (n = 21), or 6 weeks (n = 12) and control rats (n = 24, n = 21, and n = 12, respectively) were subjected to 35 minutes of left coronary artery occlusion and 2 hours of reperfusion. Infarct size and risk area were determined by triphenyltetrazolium chloride and phthalocyanine blue staining, respectively. Air nicotine, carbon monoxide, and total particulates were measured during ETS exposure. Serum lipids, plasma carbon monoxide hemoglobin (COHb), nicotine, and cotinine concentrations were measured in additional groups (6 to 13 rats each) exposed to 3 days, 3 weeks, or 6 weeks of ETS and controls. Average air nicotine, carbon monoxide, and total particulate concentrations were 1103 micrograms/m3, 92 ppm, and 60 mg/m3 for the ETS-exposed rats. Infarct size (infarct mass/risk area x 100%) increased significantly in the ETS groups compared with the control groups in a dose-dependent manner (P = .023), with longer exposure associated with larger infarct size. Infarct size nearly doubled with 6 weeks of ETS exposure (61 +/- 5% versus 34 +/- 3% for control, mean +/- SEM). Plasma COHb, nicotine, and cotinine levels increased significantly in the ETS groups in a dose-dependent manner (all P < .001). CONCLUSIONS Exposure to passive smoking increases myocardial infarct size in a rat model of ischemia and reperfusion. This increase of infarct size exhibited a dose-response relation. These results are consistent with epidemiological studies demonstrating that ETS increases the risk of heart death.

Testosterone Worsens Endothelial Dysfunction Associated With Hypercholesterolemia and Environmental Tobacco Smoke Exposure in Male Rabbit Aorta

OBJECTIVES To assess the effects of interaction of sex hormones, hypercholesterolemia (HC) and environmental tobacco smoke (ETS) exposure on endothelium-dependent relaxation, we examined vascular reactivity in vitro in an animal model of atherogenesis. BACKGROUND Animal and human studies indicate the presence of interactions between classic coronary artery disease risk factors and endothelium-dependent relaxation. Sex hormones have also been shown to influence release of endothelium-derived relaxing factor. METHODS New Zealand White rabbits were randomized to receive either an HC diet (n = 8) or ETS exposure plus HC diet (n = 8). Eight rabbits receiving a normal diet, without exposure to ETS, served as the control group. The HC diet consisted of 3% soybean oil and 0.3% cholesterol by weight over 13 weeks. The source of ETS was sidestream smoke of 4 cigarettes/15 min, 6 h/day, 5 days/week over 10 weeks in a smoking chamber. Rabbits were killed, and fresh aortic rings were harvested and maintained in oxygenated Krebs solution in an organ bath at 37 degrees C. Rings were precontracted with norepinephrine and exposed to acetylcholine in increasing doses, and isometric tension was recorded. Rings were also exposed to physiologic concentrations (1 nmol/liter) of either 17-beta-estradiol, testosterone or progesterone before pre-contraction with norepinephrine and relaxation with acetylcholine. Endothelium-independent relaxation was studied using nitroglycerin. The surface area of the ring covered by lipids was measured by Sudan IV staining. RESULTS HC and ETS significantly reduced endothelium-dependent relaxation (p = 0.01 and p < 0.0005, respectively) and caused atherogenesis (p < 0.0005 and p = 0.047, respectively) but did not affect endothelium-independent relaxation. Incubation with estradiol and estradiol plus progesterone did not influence endothelium-dependent relaxation. Testosterone reduced endothelium-dependent relaxation (p = 0.049) and augmented the endothelial dysfunction associated with ETS exposure and HC (p = 0.03). CONCLUSIONS Both HC and ETS are atherogenic and impair endothelial function but do not affect endothelium-independent relaxation. Physiologic levels of estradiol and estradiol plus progesterone do not affect endothelium-dependent relaxation. Physiologic levels of testosterone impair relaxation and augment the endothelial dysfunction associated with ETS exposure and HC.

Comparative effects of pretreatment with captopril and losartan on cardiovascular protection in a rat model of ischemia-reperfusion.

OBJECTIVES We sought to assess the comparative effects of pretreatment with captopril and losartan on myocardial infarct size and arrhythmias in a rat model of ischemia-reperfusion. BACKGROUND Angiotensin-converting enzyme (ACE) inhibitors and angiotensin II receptor blockers (ARBs) inhibit the renin-angiotensin system in different ways. However, the comparative effects of pretreatment with ACE inhibitors or ARBs on acute myocardial infarct size and arrhythmias are unknown. METHODS We randomly assigned 117 female Sprague-Dawley rats into three groups: group N was the normal control; group C was given 40 mg/kg body weight per day of captopril in drinking water; and group L was given 40 mg/kg per day of losartan in drinking water. After 10 weeks of pretreatment, 25 rats in each group were subjected to 17 min of left anterior descending coronary artery occlusion and 2 h of reperfusion with hemodynamic and electrocardiographic monitoring. Fourteen rats in each group had blood samples drawn and aortic rings removed to study vascular reactivity. RESULTS Mortality during ischemia and reperfusion was lower in combined groups L and C than in group N (4.2% vs. 19.2%, p = 0.042). Rats treated with losartan had significantly higher levels of angiotensin II in their plasma. Hemodynamic variables were not significantly different among the three groups. The thresholds of ventricular fibrillation (VF) before occlusion and after reperfusion were significantly higher in groups L and C than in group N (1.99 +/- 0.24 and 1.93 +/- 0.27 vs. 1.23 + 0.17 mA, p = 0.04; 2.13 +/- 0.25 and 1.78 +/- 0.22 vs. 0.95 +/- 0.11 mA, p = 0.001). The average episodes of ventricular tachycardia (VT) and VF per rat were significantly less in groups L and C than in group N (0.96 +/- 0.2 and 1.2 +/- 0.3 vs. 2.8 + 0.4 mA, p < 0.001). Myocardial infarct size was significantly smaller in groups L and C than in group N (34 +/- 3% and 35 +/- 3% vs. 44 +/- 3%, p = 0.031, 0.043). Endothelium-dependent vasorelaxation induced by a calcium ionophore (A23187) was increased in both groups but was only statistically significant in group C (p = 0.020). CONCLUSIONS Losartan and captopril have similar cardiovascular protective effects in a rat model of ischemia-reperfusion. They increased the threshold of VF, decreased mortality and decreased episodes of VT and VF, as well as decreased myocardial infarct size.

Effect of Lovastatin on Suppression and Regression of Atherosclerosis in Lipid-fed Rabbits

Summary The present study was designed to evaluate the effects of lovastatin on suppression and regression of atherosclerosis in the lipid-fed rabbit. Fifty-seven New Zealand rabbits in six groups were fed a 0.3% cholesterol diet for 10 weeks. In the progression phase of the study, group CIO served as a control and received 1 ml of DMSO daily by gavage. Two other groups, L10 and H10, received low (L)-dose (10 mg/day) or high (H)-dose (20 mg/ day) lovastatin dissolved in 1 ml of DMSO for 10 weeks. In the regression phase of the study, three groups of rabbits received the high lipid diet for 10 weeks and were then shifted to a normal diet for the second 10 weeks. During the second 10 weeks, the control group C20 received 1 ml of DMSO daily, and groups L20 and H20 received 10 and 20 mg/day of lovastatin by gavage, respectively. In the progression phase of the study, lovastatin significantly attenuated the percent of aortic lesions in groups L10 (8 ± 7%) and H10 (9 ± 14%) vs. the control group C10 (31 ± 17%; p < 0.01). There was a similar reduction in pulmonary lesions in groups L10 (10 ± 6%) and H10 (4 ± 5%) compared to the control group C10 (30 ± 16%; p < 0.01). There was also a reduction in plaque thickness in both the aorta and pulmonary artery, and hence an even greater reduction in estimated plaque volume. In the regression phase of the study, lovastatin also significantly reduced the percent of aortic lesions (groups L20 and H20 vs. C20: 27 ± 18 and 22 ± 7% vs. 40 ± 17%; p < 0.05) and pulmonary lesions (21 ± 10 and 17 ± 6% vs. 26 ± 9%; p > 0.05 and p < 0.05, respectively); the average maximum plaque thickness of aorta (0.24 and 0.26 mm vs. 0.49 mm; p < 0.01); and the standardized plaque volume per unit area of aorta (4.5 and 3.4 vs. 12.1 mm-%; p < 0.05 and p < 0.01, respectively). However, there was no significant difference in the percent of aortic lesions between groups L20 and H20 and group C10 (27 ± 18 and 22 ± 7 vs. 31 ± 17%; p > 0.05). In the progression phase of the study, the total serum cholesterol in groups L10 and H10 was lower than that in group C10 (715 ± 465 and 832 ± 461 vs. 1,187 ± 427 mg/dl), suggesting that the beneficial effect seen with lovastatin was due to its cholesterol-lowering effect. In the regression phase of the study, however, there were no differences in cholesterol levels in the control and treated groups. These data demonstrate that lovastatin can effectively suppress aortic and pulmonary atherosclerosis in cholesterol-fed rabbits. In addition to its cholesterol-lowering action, other mechanisms of lovastatin may contribute to this effect.

In-utero and neonatal exposure to secondhand smoke causes vascular dysfunction in newborn rats

OBJECTIVES We sought to determine the effects of secondhand smoke (SHS) exposure on vascular reactivity in newborn and infant rats. BACKGROUND Secondhand smoke exposure increases cardiovascular risk. Secondhand smoke-induced endothelial dysfunction has been demonstrated in older teenagers and young adults. We have previously shown in adult rabbits that SHS induces atherogenesis and endothelial dysfunction. The effects of SHS on vascular function in the offspring of SHS-exposed mothers and in infants are unknown. METHODS In this study the effects of in-utero (21 days) and neonatal (28 days) exposure to SHS were examined in 80 rats, 4 weeks of age, in a 2-by-2 design study. Rats were exposed to sidestream smoke in smoking chambers. Aortic rings were excised and isometric force responses to phenylephrine, acetylcholine, A23187 and nitroglycerin were studied in organ baths. RESULTS Neonatal SHS exposure reduced animal weight (p=0.009). In-utero exposure increased the sensitivity (decreased the EC50) of aortic rings to phenylephrine (p < 0.0005), as did neonatal exposure (p=0.01). Maximal contraction to phenylephrine was reduced by in-utero exposure (p=0.04). In-utero SHS exposure reduced maximal endothelium-dependent relaxation to acetylcholine (p=0.04) and increased the EC50 (p=0.05), suggesting impaired sensitivity to acetylcholine. In-utero exposure decreased the sensitivity (increased the EC50) to the endothelium-independent vasodilator nitroglycerin (p=0.003). CONCLUSIONS Secondhand smoke has detrimental effects on vascular smooth muscle function in the newborn.

Mechanisms of vasorelaxation induced by eicosapentaenoic acid (20:5n-3) in WKY rat aorta

The vasorelaxant activity of eicosapentaenoic acid (EPA, 20:5n‐3), the omega‐3 polyunsaturated fatty acid, was investigated in isolated Wistar Kyoto (WKY) rat aortae by measuring isometric tension. Eicosapentaenoic acid (1–100 μM) relaxed rat aortae contracted with high K+ (80 mM) or noradrenaline (NA, 1 μM) in a concentration‐dependent manner. Contractions induced by Bay K 8644 or increasing concentrations of calcium were unaffected by EPA. The relaxant effect of EPA (3–100 μM) was significantly inhibited by indomethacin (10 μM), the cyclo‐oxygenase inhibitor, but not by the nitric oxide (NO) synthesis inhibitor, Nω‐nitro‐L‐arginine methyl ester hydrochloride (L‐NAME, 100 μM). Removal of the endothelium did not alter EPA‐induced relaxations. In Ca2+‐free, EGTA 2 mM solution, EPA (10–30 μM significantly inhibited NA‐sustained contractions. Incubation with EPA (5, 10 μM) diminished both NA‐induced (1 μM) phasic and sustained contractions. The vasorelaxant effects of EPA (30 μM) on NA‐induced (1 μM) contractions were significantly inhibited by the K+ channel blocker, glibenclamide (10 μM), but not tetraethylammonium (1 mM). Moreover, indomethacin and glibenclamide combined significantly inhibited EPA‐induced (1–100 μM) responses. These results indicate EPA exerts its endothelium‐independent vasorelaxant effects in WKY rat aortae through production of prostanoids which activate K+ATP channels. Inhibition of Ca2+ mobilization from intracellular pools and influx through the non‐L‐type, but not the L‐type, Ca2+ channel are also possible mechanisms action of EPAs.

Is the reduction of myocardial infarct size by dietary fish oil the result of altered platelet function

Sprague-Dawley rats fed a diet containing 12% fish oil (18% eicosapentaenoic acid [EPA] and 12% docosahexaenoic acid [DHA]), for 1 week (group I, n = 9) or 8 weeks (group III, n = 42) and controls (group II, n = 8; group IV, n = 36, respectively) were subjected to 35 minutes of left coronary artery occlusion followed by 120 minutes of reperfusion. Compared to the controls, infarct size was significantly reduced in group III (15% +/- 2%, n = 42 vs 34% +/- 4%, n = 36; p < 0.001; infarct mass/risk area x 100%), but no change in group I (39% +/- 5%, n = 9 vs 35% +/- 5%, n = 8; p = not significant). Bleeding time was prolonged in group III (290 +/- 73 sec) compared to group IV (99 +/- 10 sec, p = 0.015). Omega-3 fatty acid (EPA and DHA) levels in platelets were significantly higher in the rats fed 8 weeks of fish oil (group III) compared to the controls (group IV) and the rats fed 8 weeks of fish oil and then a regular diet until bleeding time normalized (group V) (7.2% +/- 0.6% vs 1.2% +/- 0.2% and 4.9% +/- 0.5%; 3.8% +/- 0.7% vs 1.8% +/- 0.3% and 2.8% +/- 0.6%, p < 0.001 and 0.05, respectively). These data indicate that long-term (8 weeks) dietary fish oil supplementation significantly reduces infarct size; short-term (1 week) does not. This reduction of infarct size appears to correlate with altered platelet function and EPA and DHA levels in platelets.

Comparative effects of ACE inhibitors and an angiotensin receptor blocker on atherosclerosis and vascular function.

Background: Both angiotensin-converting enzyme inhibitors (ACE-IS) and angiotensin receptor blockers (ARBS) provide vascular protection. This study was designed to compare ACE-IS with widely differing tissue affinity (captopril and quinapril) and an ARB (losartan) on vascular protection against the adverse effects of high cholesterol. Methods and Results: Forty-two New Zealand rabbits on a 0.5% cholesterol diet were ran-domized into control, captopril (10 mg/kg/d), quinapril (0.3 mg/kg/d), and losartan (8 mg/kg/d) groups for 14 weeks. Captopril, quinapril, and losartan significantly attenuated aortic lipid lesions (P = 0.001). Captopril and quinapril were more effective than losartan in preserving vascular relaxation. Conclusions: Captopril, quinapril, and losartan had similar protective effects against atherogenesis. Captopril and quinapril were more effective than losartan in preserving vascular function. Increased bradykinin by ACE inhibition may be responsible for this improved vascular endothelial function.

Chronic Dietary l-Arginine Prevents Endothelial Dysfunction Secondary to Environmental Tobacco Smoke in Normocholesterolemic Rabbits

Our goal was to determine whether environmental tobacco smoke causes endothelial dysfunction in the absence of hypercholesterolemia and whether such an effect can be prevented by supplementation with L-arginine. Environmental tobacco smoke exposure is associated with an increase in coronary artery disease events and mortality. We have previously demonstrated that environmental tobacco smoke causes endothelial dysfunction and atherosclerosis in rabbits with diet-induced hypercholesterolemia and atherosclerosis and that chronic dietary L-arginine supplementation prevents this. The effects of L-arginine supplementation (2.25% solution ad libitum) and environmental tobacco smoke (smoking chambers for 10 weeks) were examined with a 2 x 2 design in 32 rabbits fed a normal diet. Acetylcholine, calcium ionophore A23187, and nitroglycerin-induced vasorelaxation were assessed in aortic rings precontracted with phenylephrine. Endothelial L-arginine levels were measured by chromatography. Chronic L-arginine supplementation increased serum (P < .001) and endothelial (P = .003) L-arginine levels. Environmental tobacco smoke reduced endothelium-dependent acetylcholine-induced relaxation, and L-arginine blocked this adverse effect (P = .04). Environmental tobacco smoke tended to increase phenylephrine-induced contraction (P = .06). Neither environmental tobacco smoke nor L-arginine influenced A23187-induced relaxation nor endothelium-independent nitroglycerin-induced relaxation. Endothelial dysfunction secondary to environmental tobacco smoke may occur in the absence of diet-induced hypercholesterolemia and atherosclerosis. Chronic dietary supplementation with a nitric oxide donor such as L-arginine offsets the endothelial dysfunction associated with environmental tobacco smoke in normocholesterolemic rabbits, possibly through substrate loading of the nitric oxide pathway.