Down regulation of aortic nitric oxide and antioxidant systems in chronic alcohol-induced hypertension in rats

The aim of this study was to investigate the relationship between chronic ethanol-induced increase in blood pressure (BP) and alterations in the aortic nitric oxide (NO) and the antioxidant systems in rats. Male Fisher rats (200-250 g) were divided into two groups of six animals each and treated as follows: 1) control (5% sucrose, orally) daily for 12 weeks and 2) 20% ethanol (4 g/kg, orally) daily for 12 weeks. The BP (systolic, diastolic and mean) was recorded every week through tail-cuff method. The animals were sacrificed 12 weeks after treatments and thoracic aorta was collected and analysed. The results show that systolic, diastolic and mean BP was significantly elevated 12 weeks after ethanol ingestion in rats compared to control. The endothelial nitric oxide synthase (eNOS) and vascular endothelial growth factor (VEGF) expressions were down-regulated (50-55% of control) leading to depletion of aortic NO levels (69% of control) in ethanol treated rats compared to control. The ratio of reduced to oxidized glutathione (GSH/GSSG) was significantly depleted (58% of control) in the aorta of ethanol-treated rats compared to control. The decrease in aortic GSH/GSSG ratio was good correlated with increase in BP (r = 0.69). The antioxidant enzymes: copper/zinc-superoxide dismutase (CuZn-SOD) and manganese (Mn)-SOD, catalase (CAT) and glutathione peroxidase (GSH-Px) activities were significantly depressed (36-53% of control) in the aorta of ethanol-treated rats compared to control. The study concluded that chronic ethanol ingestion induces hypertension which relates to the vascular endothelial dysfunction on account of the down-regulation of aortic endothelial antioxidants and NO generating system in rats.     

Inducible nitric oxide synthase attenuates adrenergic signaling in alcohol fed rats

Chronic, excessive alcohol consumption is associated with myocardial dysfunction in humans. The molecular mechanisms and cellular signaling pathways contributing to this cardiac dysfunction remain largely unknown. This study examined the effects of chronic alcohol consumption on myocardial function and cardiac myocyte signaling pathways. Adult male rats were fed a commercially prepared diet containing either ethanol (13 g/kg/d) or isocaloric control diet for 1 month. In vivo hemodynamics were measured in awake rats after inserting a catheter tip in the left ventricle under general anesthesia. Ventricular dysfunction was evidenced in awake, alcohol-fed rats by increased left ventricular end diastolic pressure, decreased systolic developed left ventricular pressure, and decreases in both positive and negative dp/dt compared with controls. Cardiac myocytes isolated from alcohol-fed rats also demonstrated an attenuated response to the beta-adrenergic agonist, isoproterenol, compared to controls. This response was significantly reversed by the nitric oxide synthase (NOS) inhibitor, N-monomethyl-L-arginine (L-NMMA). Western analyses confirmed inducible nitric oxide synthase (iNOS) protein synthesis in cardiac myocytes isolated from alcohol fed rats. It is therefore concluded that chronic alcohol ingestion results in iNOS-mediated attenuation of adrenergic signaling and depression in both systolic and diastolic function in rats.     

Chronic ethanol ingestion induces aortic inflammation/oxidative endothelial injury and hypertension in rats

The study aim was to investigate the relationship of chronic ethanol-induced inflammation leading to vascular endothelial injury and elevation of blood pressure (BP) in a rat model. Male Fisher rats were divided into two groups of six animals each and treated as follows: (1) Control (5% sucrose, orally) daily for 12 weeks and (2) 20% ethanol (4 g kg(-1), orally) daily for 12 weeks. The mean arterial blood pressure was recorded every week. The animals were anesthetized with pentobarbital after 12 weeks; thoracic aorta were isolated and analyzed for aortic reactivity response, inflammatory mediators, oxidant/antioxidant enzyme protein expression and endothelial nitric oxide-generating system. The results show that the mean BP was significantly elevated 12 weeks after ethanol ingestion. The increased BP was related to increased aortic inflammation (tumor necrosis factor [TNF]-α; nitric oxide synthase [iNOS], COX-2 and MCP-1 protein expression) and elevated angiotensin II levels in alcohol-treated group compared to control. Aortic Nicotinamide adenine dinucleotide phosphate reduced (NADPH) oxidase activity, membrane and cytosolic subunits p22(phox) and p47(phox) expression and Mn-SOD activity and protein expression significantly increased, whereas nitric oxide (NO), endothelial NO synthase (eNOS), vascular endothelial growth factor (VEGF)-A and CuZn-SOD activity and protein expression significantly decreased in alcohol-treated group compared to control. The acetylcholine-mediated vasorelaxation response was depressed in the aorta of ethanol-treated rats compared to control. In conclusion, chronic ethanol-induced elevation in BP is related to increased aortic inflammation, elevated angiotensin II levels, induction of NADPH oxidase causing endothelial injury, depletion of CuZn-SOD, down-regulation of endothelial NO generating system and impaired vascular relaxation in rats.     

Chronic β-AR activation-induced calpain activation and impaired eNOS–Akt signaling mediates cardiac injury in ovariectomized female rats

Objective: To address the pathophysiological relevance of ovarian hormones in chronic β-adrenergic stimulation-induced myocardial injury, we assessed impairments of Ca²⁺-mediated cell signaling in the left ventricle of ovariectomized female rats. Research design/methods: Female Wistar rats were subjected to bilateral ovariectomy and sham operation. Six weeks after ovariectomy (OVX), both OVX and sham rats were treated with isoproterenol (5mg/kg, intraperitoneally), a nonselective β-adrenergic agonist, once a day for 28 days. Results: We found that chronic β-adrenergic stimulation caused enhanced breakdown of sarcolemmal proteins such as dystrophin and utrophin in OVX rats compared to sham-operated rats. Generation of calpain-mediated 150 kDa-breakdown product of spectrin confirmed calpain activation following isoproterenol treatment. Marked breakdown of endogenous calpain inhibitor, calpastatin, in OVX rats was consistent with the calpain activation following chronic β-adrenergic stimulation. In addition to calpain activation, we also found marked reduction of endothelial nitric oxide synthase (eNOS) activity with concomitant deregulation by heat shock proteins 90 kDa and caveolin 3, both of which are eNOS-associated proteins. Finally, we documented decreased Akt phosphorylation with concomitant increased glycogen synthase kinase 3β phosphorylation underlying cell injury following chronic β-adrenergic stimulation. Conclusion: Taken together chronic β-adrenergic stimulation caused severe cardiac injury in OVX rats through calpain activation and impairments of Akt and eNOS signaling pathways.     

二苯乙烯苷预防性干预大鼠动脉硬化对其一氧化氮合酶表达的影响

目的探讨二苯乙烯苷(TSG)对动脉粥样硬化(AS)大鼠一氧化氮(NO)含量及主动脉一氧化氮合酶(NOS)表达的影响。方法SD大鼠60只,♂,随机分为6组:①正常对照组;②模型组;③TSG低剂量组;④TSG中剂量组;⑤TSG高剂量组;⑥辛伐他汀阳性对照组。造模12wk后,颈动脉取血,检测血清NO含量。取血后分离主动脉,保存于-70℃,检测主动脉组织中NOS含量,RT-PCR检测大鼠主动脉eNOS和iNOS mRNA表达。实验数据采用STATA8.0软件进行统计分析。结果与模型组相比,辛伐他汀组和TSG各剂量组均能增加血清及主动脉组织NO的含量、主动脉组织NOS的水平、主动脉组织eNOS mRNA的表达及降低主动脉组织iNOS mRNA的表达,并呈剂量依赖性。结论TSG能上调AS大鼠动脉壁eNOS mRNA的表达,抑制AS大鼠动脉壁iNOS mRNA的表达,这可能是TSG抗AS的机制之一。     

Propofol protects human umbilical vein endothelial cells from cisplatin-induced injury

The anticancer drug cisplatin can up-regulate endothelial adhesion molecule expression, and trigger vascular endothelial injury. Propofol, an intravenous anesthetic, can inhibit endothelial adhesion molecule expression in some situations. Here, we explored whether and how propofol improved cisplatin-induced up-regulation of endothelial adhesion molecules in human umbilical vein endothelial cells. Compared with control group, cisplatin reduced endothelial nitric oxide synthase dimer/monomer ratio, activated protein kinase C and enhanced endothelial nitric oxide synthase-Thr⁴⁹⁵ phosphorylation, decreased nitric oxide production, augmented intercellular adhesion molecule 1 expression and monocyte-endothelial adhesion. These cisplatin-mediated effects were attenuated by propofol treatment. Nω-Nitro-L-arginine methyl ester hydrochloride, a nitric oxide synthase inhibitor, inhibited the effect of propofol on cisplatin-induced intercellular adhesion molecule 1 expression. Propofol improved cisplatin-mediated tetrahydrobiopterin reduction and nitrotyrosine overexpression. Compared with control group, cisplatin and PMA, a protein kinase C activator, both increased endothelial nitric oxide synthase-Thr⁴⁹⁵ phosphorylation, while propofol and GFX, a protein kinase C inhibitor, both decreased cisplatin-induced endothelial nitric oxide synthase-Thr⁴⁹⁵ phosphorylation. Our data indicated that propofol, via reducing cisplatin-induced endothelial nitric oxide synthase uncoupling and endothelial nitric oxide synthase-Thr⁴⁹⁵ phosphorylation, restored nitric oxide production, intercellular adhesion molecule 1 expression and monocyte-endothelial interaction.     

Evaluation of the antiperoxidative effects of melatonin in ammonium acetate-treated Wistar rats

The efficacy of melatonin (MLT) against ammonium acetate-induced neurotoxicity was biochemically studied in the experimental rats. The activities of serum transaminases and the levels of thiobarbituric acid reactive substances were significantly increased in ammonium acetate-treated rats. These levels were significantly decreased in MLT and ammonium acetate-treated rats. Further, non-enzymatic (vitamin C and E) and enzymatic (superoxide dismutase and catalase) antioxidants were significantly decreased in ammonium acetate-treated rats and were increased in MLT and ammonium acetate-treated rats. These biochemical alterations during MLT treatment could be due to its ability to: (i) scavenge a variety of radicals and reactive species, (ii) induce antioxidative enzymes which reduce steady state levels of reactive species, (iii) inhibit nitric oxide synthase which generates nitric oxide and (iv) stabilize cell membranes which assists them in reducing oxidative damage and, thus, prevents the oxidative stress in rats.     

17B-estradiol and hypercholesterolemia: involvement of nitric oxide

The mechanisms of the protective effect of 17B-estradiol were investigated in the middle cerebral artery (MCA) and aorta isolated from cholesterol-fed rabbits. Three groups were assigned: control group (standard chow), cholesterol group (standard chow+1% cholesterol) and estradiol group (1% cholesterol+17B-estradiol). The MCA and the aorta were isolated, precontracted respectively with high K(+) solution or with phenylephrine and exposed to cumulative acetylcholine concentrations. In the control group, acetylcholine induced a concentration-dependent relaxation in the aorta and the MCA. Cholesterol diet for eight months reduced significantly the maximal response to acetylcholine by about 50% in the aorta and by about 30% in the MCA. The chronic treatment with 17B-estradiol restored this impaired relaxations to acetylcholine. Incubation of arteries from estradiol group with N(omega)-nitro L-arginine methyl-ester (L-NAME), a potent inhibitor of constitutive nitric oxide synthase, entirely abolished the relaxation to acetylcholine while aminoguanidine, a potent inhibitor of inducible nitric oxide synthase, did not affect this relaxation. These observations suggest that the protective effect of 17B-estradiol against hypercholesterolemia is mediated via a release of endothelial nitric oxide.     

Vitamin C prevents radiation-induced endothelium-dependent vasomotor dysfunction and de-endothelialization by inhibiting oxidative damage in the rat

1. The present study was undertaken to determine whether endothelial function or morphology was altered in aortic rings of rats after irradiation, to investigate the mechanism of radiation effects on the endothelium and to examine the effect of vitamin C treatment against radiation-induced damage of the endothelium. 2. Female Sprague-Dawley rats were randomized into four groups (control, radiation, radiation + vitamin C, radiation + vitamin C + NG-nitro-L-arginine methyl ester (L-NAME); n = 10 for each group and n = 7 for the control group) and were irradiated with 10 Gy of 137Cs as a radiation source. Segments of the thoracic aorta were obtained and isometric tension, levels of 8-hydroxydeoxyguanosine (OH-dG) and immunohistochemical staining were measured. 3. Irradiation significantly impaired the acetylcholine-induced vasodilation of aortic segments, an effect that could be prevented by pretreatment with vitamin C (500 mg/kg per day). This beneficial effect of vitamin C was abolished by the addition of L-NAME (100 microg/kg per day), an inhibitor of nitric oxide (NO) synthesis. Irradiation significantly increased the level of OH-dG in the aorta (1.02 +/- 0.27 vs 2.61 +/- 0.78 OH-dG/105 deoxyguanosine (dG) for control and irradiated tissues, respectively; P < 0.01), an increase that was prevented by vitamin C treatment (1.59 +/- 0.23 OH-dG/105 dG; P < 0.01). Irradiation caused significant de-endothelialization (von Willebrand factor (vWF) staining was 93 +/- 7 vs 100% in irradiated and control tissues, respectively; P < 0.05) and this was prevented by vitamin C treatment (vWF staining 98 +/- 3%; P < 0.05). 4. Radiation caused endothelial damage and impaired NO production through oxidative injury, resulting in a selective impairment of endothelial-dependent vasodilation that could be prevented by vitamin C, partly through anti-oxidant mechanisms.     

Altered active but not passive properties of mesenteric resistance arteries from the vitamin E-deprived rat

1. We tested the hypothesis that lowering antioxidant protection through dietary vitamin E deprivation would alter active and passive mechanical properties in resistance arteries of the rat. Specifically, we hypothesized that vascular tone in isolated mesenteric arteries of the vitamin E-deprived rats would be altered due to impaired endothelial influences of nitric oxide and/or prostaglandins.
2. Lumen diameter and wall thickness were measured in pressurized arteries (≈amp;250 μm diameter) from control (n=9) and vitamin E deprived (n=9) Sprague-Dawley female rats by use of a dimension analysing system.
3. Treatment with a cyclo-oxygenase inhibitor (meclofenamate) did not affect the basal vascular tone in either group. Treatment with a nitric oxide synthase inhibitor (N^G-methyl-L-arginine) caused a significant increase in basal tone only in the vitamin E-deprived rats (% tone: 6.2±1.1 vs 1.2±0.3%; P<0.05). When tone was induced to 25% of the initial diameter with phenylephrine, treatment with the nitric oxide synthase inhibitor resulted in a greater potentiated tone in the vitamin E-deprived rats compared to the controls (26.5±2.7 vs 16.4±3.4%; P<0.05); suggesting a greater nitric oxide affect in the vessels from the vitamin E-deprived rats. Meclofenamate treatment in the induced tone arteries significantly relaxed (−17.4±4.0%; P<0.05) only the arteries from the vitamin E-deprived rats, indicating that a vasoconstrictor was modifying tone. The passive characteristics of distensibility and stress-strain relationship were not different between the two groups of rats.
4. In summary, vitamin E deprivation in the rat enhanced the modulation of vascular tone by both the nitric oxide and cyclo-oxygenase pathways but did not alter passive characteristics of mesenteric arteries.

     

Arsenite increases vasoconstrictor reactivity in rat blood vessels: role of endothelial nitric oxide function

Arsenite has been shown to inhibit endothelium-dependent, nitric oxide-mediated vasodilation in vitro. This study investigated the effects of arsenite on vascular reactivity in vivo. Saline or sodium arsenite (6 mg kg-1) was administered intravenously in Wistar-Kyoto rats for 4 h. As compared to saline, arsenite significantly increased vasoconstrictor responses to phenylephrine in both rat isolated aorta and renal arteries examined in tissue bath. This change was diminished after preincubation of the tissues with the nitric oxide synthase inhibitor NG-nitro-L-arginine methyl ester, which increased phenylephrine-induced vasoconstriction to a similar extent as arsenite. In contrast, acetylcholine-induced vasodilation, mediated by nitric oxide in the aorta and by an endothelium-derived hyperpolarizing factor in renal arteries, was not affected by arsenite. Arsenite induced expression of heat shock proteins Hsp72, Hsp32, and Hsp90, but endothelial nitric oxide synthase expression was not changed. The effects of arsenite on vasoreactivity were unlikely to be mediated by heat shock protein induction, because blockade of heat shock protein induction had little effect on the increased vasoconstriction in vessels from arsenite-treated animals. Our study suggests that in vivo arsenic treatment increases vasoconstrictor reactivity by compromising basal endothelial nitric oxide function, which is not caused by altered endothelial nitric oxide synthase expression.     

Nitro-aspirin is a potential therapy for non alcoholic fatty liver disease

Non-alcoholic fatty liver disease (NAFLD) is the most common cause of chronic liver injury; however its therapeutic strategy has not been established yet. Nitro-aspirin (NO-aspirin) is a new molecule in which aspirin and a NO-donating group are covalently linked. This study investigated the potential protective effect of NO-aspirin on NAFLD. Experimental rats were assigned into 4 groups. Group 1 was fed with normal diet and served as normal control group. Group 2 was fed with 2% cholesterol diet and received vehicle as positive control NAFLD group. Group 3 was fed with 2% cholesterol diet plus NO-aspirin (100 mg/kg/day). Group 4 was fed with 2% cholesterol diet plus aspirin (55 mg/kg/day). Rats were treated for 8 weeks. The results showed that NO-aspirin (but not aspirin) prevented the development of NAFLD as evidenced by significant reduction in liver weight/body weight ratio (liver index) and histopathologic changes. The protective effect of NO-aspirin is accompanied with significant decrease in triglycerides, malondialdehyde (MDA), and nitric oxide (NO) in hepatic tissue. Semi-quantitative immunohistochemical studies showed significant decrease in expression of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) in hepatic tissue. In conclusion, NO-aspirin inhibited multiple pathways involved in the pathogenesis of NAFLD indicating that it might serve as a new therapeutic strategy.     

A flavonoid-rich diet increases nitric oxide production in rat aorta

1. Red wine intake is associated with a low risk of cardiovascular disease. This effect has been partly attributed to the action of polyphenolic compounds, which decrease the oxidation of plasma low density lipoproteins. Moreover, nitric oxide ((*)NO) is a vasodilator and polyphenolic compounds induce endothelium-dependent vasorelaxation in vitro. 2. Here we studied whether a diet rich in dealcoholated red wine (DRW) increases acetylcholine-induced vasorelaxation and whether ingestion of DRW-, quercetin- or catechin-rich diets modifies the (*)NO-cyclic guanosine-3',5'-monophosphate (cyclic GMP) pathway and superoxide anion (O2(.-)) release in aorta in a resting state in rats fed semi-purified diets containing either 35% (v w(-1)) DRW, 0.3% (w w(-1)) quercetin or 0.3% (w w(-1)) catechin for 10 days. 3. (*)NO-mediated vasorelaxation induced by acetylcholine was greater in rats fed the DRW-rich diet than in those that received the control diet. 4. Expression of endothelial (*)NO synthase (eNOS) was similar in the four dietary groups. The aortic rings of rats fed either the DRW-, quercetin-, or catechin-rich diets showed higher NOS activity, (*)NO production and cyclic GMP content than those of rats fed the control diet. No changes were observed in O2(.-) production. 5. In summary, diets rich in either DRW, quercetin or catechin induced endothelium-dependent vasorelaxation in rat aorta in a resting state through the enhancement of (*)NO production, without modifying O2(.-) generation, thus the bioavailability of (*)NO was increased. The increase in the (*)NO-cyclic GMP pathway explains the beneficial effect of flavonoids at vascular level.     

Ameliorative effect of berberine on endothelial dysfunction in diabetic rats induced by high-fat diet and streptozotocin

Berberine can improve insulin resistance, lower blood glucose, and regulate lipid metabolism disorders which cause endothelial dysfunction, leading to vascular complications of type 2 diabetes mellitus. The aim of the present study was to investigate the effects of berberine on endothelial dysfunction of aortas in type 2 diabetes mellitus rats and its mechanism. Wistar rats were randomly divided into four groups: diabetic rats, control rats, diabetic rats treated with berberine (100 mg/kg), and control rats treated with berberine. The serum fasting blood glucose, insulin, total cholesterol, triglyceride and nitric oxide (NO) levels were tested. Acetylcholine-induced endothelium-dependent relaxation and sodium nitroprusside induced endothelium-independent relaxation were measured in aortas for estimating endothelial function. The expression of endothelial nitric oxide synthase (eNOS) mRNA was measured by RT-PCR, and the protein expressions of eNOS and NADPH oxidase (NOX4) were analyzed by western blot. The results showed that berberine significantly decreased fasting blood glucose, and triglyceride levels in diabetic rats. Berberine also improved endothelium-dependent vasorelaxation impaired in aorta. The expressions of eNOS mRNA and protein were significantly increased, while NOX4 protein expression was decreased in aortas from diabetic rats with berberine treatment. Moreover, serum NO levels were elevated after berberine treatment. In conclusion, berberine restores diabetic endothelial dysfunction through enhanced NO bioavailability by up-regulating eNOS expression and down-regulating expression of NADPH oxidase.     

木贼水煎剂对高血脂大鼠主动脉内皮细胞的保护作用

目的 探讨木贼对食饵性高脂血症大鼠主动脉内皮细胞的保护作用.方法 48只SD大鼠随机分为4组,成功复制高脂血症和早期动脉粥样硬化模型,测定血清NO含量及NOS活性;流式细胞仪检测内皮细胞凋亡率及Bcl-2和Bax基因蛋白的表达; 光镜和电镜观察内皮细胞的形态学改变.结果 木贼能纠正由高脂血症所致的NO代谢紊乱;调控Bcl-2和Bax的比值,降低内皮细胞的凋亡率;减轻动脉内皮细胞的损伤程度.结论 木贼可明显改善高脂血症所致的动脉内皮细胞的功能障碍和形态损伤.     

银杏叶提取物对糖尿病大鼠一氧化氮水平的影响

目的 研究银杏叶提取物(EGb)对糖尿病大鼠心肌、睾丸、脑组织一氧化氯水平的影响。方法用链脲佐菌素制备SD大鼠糖尿病模型。测定EGb对糖尿病大鼠心肌、睾丸、脑组织一氧化氯(NO)的含量及一氧化氯合酶(NOS)、诱导型一氧化氯合酶(iNOS)、结构型一氧化氯合酶(cNOS)的活性的影响。结果与正常组相比，糖尿病大鼠心肌、睾丸组织NO含量及NOS，iNOS活性升高，脑组织NO含量及NOS活性升高。与糖尿病组相比，EGb治疗组大鼠心肌、睾丸组织NO含量及NOS、iNOS活性下降。结论 EGb能够对抗糖尿病大鼠过量NO对心肌、睾丸组织的损伤。     

Effect of melatonin on altered expression of vasoregulatory genes during hepatic ischemia/reperfusion

The production of reactive oxygen species during hepatic ischemia/reperfusion (I/R) can help create disturbances in microcirculation. This study examined the effect of melatonin, a pineal secretory product and a potent antioxidant, on the expression of vascular stress genes during hepatic I/R. Rats were subjected to 60 min of hepatic warm ischemia followed by 5 h reperfusion. Melatonin (10 mg/kg) was administered intraperitoneally 15 min before ischemia and immediately before reperfusion. The serum alanine aminotransferase and hepatic malondialdehyde levels increased markedly after I/R. These increases were significantly inhibited by melatonin. The levels of endothelin-1 (ET-1) and its receptor, ET(B) mRNA, were elevated by I/R but attenuated by melatonin. The mRNA levels of endothelial nitric oxide synthase (eNOS), inducible nitric oxide synthase (iNOS), and heme oxygenase-1 were significantly higher after I/ R. Melatonin augmented the increase in the eNOS mRNA level, whereas it reduced the increase in the iNOS mRNA level. The expression of tumor necrosis factor-alpha was increased markedly by I/R. This increase was also attenuated by melatonin. These results suggest that melatonin ameliorates the imbalanced expression of the vascular stress genes during hepatic I/R through its antioxidant property.