八肽缩胆囊素对脂多糖诱导血管内皮细胞诱生型一氧化氮合酶表达变化的抑制作用

目的探讨八肽缩胆囊素（CCK-8）对脂多糖（LPS）诱导血管内皮细胞诱生型一氧化氯合酶（iNOS）表达变化的影响。方法培养人脐静脉内皮细胞株ECV-304细胞。用0．01、0．1和1mg／L LPS处理2～24h，用生理盐水、10mol／LCCK-8和0．1mg／L LPS＋10^-8、10^-7、10^-8mol／L CCK-8处理16h；用比色法检测培养液中一氧化氮（NO）含量、细胞NOS活性，免疫细胞化学及蛋白质免疫印迹法检测iNOS蛋白表达。结果与生理盐水处理的对照组比较，LPS诱导培养液NO含量增多、细胞NOS活性增高、iNOS蛋白表达上调；CCK-8剂量依赣性抑制LPS的上述效应。而单独作用对iNOS蛋白表达、NOS活性和NO含量均无明显影响。结论CCK-8可以明显抑制LPS引起ECV-304细胞iNOS蛋白表达上调。减少NO生成。     

Dietary phosphate restriction ameliorates endothelial dysfunction in adenine-induced kidney disease rats

Hyperphosphatemia causes endothelial dysfunction as well as vascular calcification. Management of serum phosphate level by dietary phosphate restriction or phosphate binders is considered to be beneficial to prevent chronic kidney disease patients from cardiovascular disease, but it has been unclear whether keeping lower serum phosphate level can ameliorate endothelial dysfunction. In this study we investigated whether low-phosphate diet can ameliorate endothelial dysfunction in adenine-induced kidney disease rats, one of useful animal model of chronic kidney disease. Administration of 0.75% adenine-containing diet for 21 days induced renal failure with hyperphosphatemia, and impaired acetylcholine-dependent vasodilation of thoracic aortic ring in rats. Then adenine-induced kidney disease rats were treated with either control diet (1% phosphate) or low-phosphate diet (0.2% phosphate) for 16 days. Low-phosphate diet ameliorated not only hyperphosphatemia but also the impaired vasodilation of aorta. In addition, the activatory phosphorylation of endothelial nitric oxide synthase at serine 1177 and Akt at serine 473 in the aorta were inhibited by in adenine-induced kidney disease rats. The inhibited phosphorylations were improved by the low-phosphate diet treatment. Thus, dietary phosphate restriction can improve aortic endothelial dysfunction in chronic kidney disease with hyperphosphatemia by increase in the activatory phosphorylations of endothelial nitric oxide synthase and Akt.     

川芎嗪对缺氧损伤的人脐静脉内皮细胞分泌一氧化氮的影响

目的观察川芎嗪对缺氧损伤的人脐静脉上皮细胞分泌一氧化氮功能的影响。方法以连二亚硫酸钠作为耗氧剂,建立体外人脐静脉上皮细胞缺氧损伤模型,与不同浓度的川芎嗪共同孵育后测定细胞培养液中的一氧化氮和一氧化氮合酶的含量。结果在以终浓度为1μmol/L的川芎嗪的条件下,细胞培养液中的一氧化氮和一氧化氮合酶的含量显著高于模型组(P<0.05,P<0.01)。结论川芎嗪能提高缺氧损伤造成的内皮细胞一氧化氮合酶的活性,增加分泌一氧化氮分泌。     

Mechanisms of homocysteine-induced atherothrombosis

Elevation of plasma homocysteine level is a risk factor for cardiovascular disease, stroke, and venous thromboembolism. It is still uncertain, however, whether hyperhomocysteinemia is a causative factor or a marker of vascular disease. The strongest evidence that homocysteine plays a causal role in atherothrombosis has been provided by studies using animal models. In the past decade, considerable progress in defining the vascular effects of hyperhomocysteinemia was achieved through the use of genetic and dietary approaches to induce hyperhomocysteinemia in experimental animals. A key vascular phenotype observed in hyperhomocysteinemic animals is endothelial dysfunction, manifested by decreased bioavailability of endothelium-derived nitric oxide. Impairment of endothelial function may be mediated by either accelerated oxidative inactivation of nitric oxide or inhibition of nitric oxide production caused by the endogenous nitric oxide synthase inhibitor, asymmetric dimethylarginine. Hyperhomocysteinemia also increases susceptibility to arterial thrombosis and accelerates the development of atherosclerosis in susceptible models such as the apolipoprotein E-deficient mouse. Mechanisms of atherothrombosis may include homocysteine-induced thiolation or acylation of plasma or endothelial proteins and endoplasmic reticulum stress, which activates signal transduction pathways leading to inflammation and apoptosis.     

Nitric Oxide in Hypertension and Renal Diseases

Nitric oxide (NO) is intimately involved in the regulation of vascular tone, renal haemodynamics and sodium balance. The physiological actions of NO suggest important vascular and renal protective roles for NO. When produced in large amounts, however, NO may also mediate cytotoxic effects. Increasing evidence suggests that endothelial function, notably the NO pathway, may be compromised in hypertension. It is not known, however, whether changes in endothelial function are primary or secondary to the development of hypertension. In renal diseases evidence for both excessive and deficient activity of NO pathway has been found. Increased glomerular production of NO via inducible NO synthase (NOS) with potential cytotoxic consequences has been demonstrated in experimental acute glomerulonephritis. On the other hand, indirect evidence obtained by means of NOS inhibitors point out to an important renoprotective role for NO in renal diseases. NO may counteract disease progression in renal diseases by preventing glomerular microthrombi, maintaining renal perfusion and medullary oxygenation, and via its anti-inflammatory/antiproliferative effects. However, these beneficial effects of NO may be compromised (endothelial and/or tubular dysfunction) in chronic nephropathies resulting in an accelerated course of renal disease. In future, more specific inhibitors and activators of different NOS isoforms are needed to elucidate the role of NO in various renal diseases in detail, and for treatment strategies aimed at modifying the NO pathway.     

Enhanced peroxynitrite formation is associated with vascular aging

Vascular aging is mainly characterized by endothelial dysfunction. We found decreased free nitric oxide (NO) levels in aged rat aortas, in conjunction with a sevenfold higher expression and activity of endothelial NO synthase (eNOS). This is shown to be a consequence of age-associated enhanced superoxide (.O(2)(-)) production with concomitant quenching of NO by the formation of peroxynitrite leading to nitrotyrosilation of mitochondrial manganese superoxide dismutase (MnSOD), a molecular footprint of increased peroxynitrite levels, which also increased with age. Thus, vascular aging appears to be initiated by augmented.O(2)(-) release, trapping of vasorelaxant NO, and subsequent peroxynitrite formation, followed by the nitration and inhibition of MnSOD. Increased eNOS expression and activity is a compensatory, but eventually futile, mechanism to counter regulate the loss of NO. The ultrastructural distribution of 3-nitrotyrosyl suggests that mitochondrial dysfunction plays a major role in the vascular aging process.     

Increased cardiac endothelial nitric oxide synthase expression in patients taking angiotensin-converting enzyme inhibitor therapy

BACKGROUND: The efficacy of angiotensin-converting enzyme (ACE) inhibitors has been demonstrated in large clinical trials, but knowledge of the underlying mechanisms remains incomplete. Therefore, this study investigated the impact of ACE inhibitor therapy on cardiac nitric oxide (NO) synthases in patients with coronary artery disease (CAD) or heart failure. PATIENTS AND METHODS: The mRNA expression was quantified by standard calibrated competitive RT-PCR, protein expression by Western blotting and NOS activity by monitoring the conversion of [3H]arginine to [3H]citrulline during enzymatic formation of NO in tissue homogenates of myocardium of patients with, or without, ACE inhibitor treatment before elective coronary artery bypass grafting or heart transplantation. RESULTS: The mRNA expression (amol microg(-1) RNA) of endothelial NO synthase (eNOS) was higher (22.5 +/- 4.8, n = 23) in the atrial myocardium of patients taking ACE inhibitor treatment, before elective coronary artery bypass grafting, compared with patients not taking this therapy (8.9 +/- 0.7, n = 33, P < 0.0001). The ACE inhibitor therapy increased eNOS protein expression from [(9 +/- 0.7) relative units (RUs) to (12 +/- 0.9) RUs, P < 0.05, respectively] and cardiac NOS activity from 17.6 +/- 1.3 to 23.7 +/- 1.1 pmol mg protein(-1) min(-1) (P < 0.001, respectively). Inducible and neuronal NO synthase expression was not changed by the ACE inhibition. A similar up-regulation of eNOS by ACE inhibition was found in the left ventricles of patients with heart failure. The augmented endothelial NOS expression and activity was not the result of differences in clinical characteristics and concomitant therapy between the patient groups. CONCLUSION: Increased eNOS expression and activity might contribute to the beneficial effects of ACE inhibitor therapy in the treatment of CAD and heart failure.     

Insulin activates vascular endothelial growth factor in vascular smooth muscle cells: influence of nitric oxide and of insulin resistance

BACKGROUND: We aimed to evaluate whether insulin influences vascular endothelial growth factor (VEGF) synthesis and secretion in cultured vascular smooth muscle cells (VSMCs) via nitric oxide (NO) and whether these putative effects are lost in insulin-resistant states. MATERIALS AND METHODS: In VSMC derived from human arterioles and from aortas of insulin-sensitive Zucker fa/+rats and insulin-resistant Zucker fa/fa rats incubated with different concentrations of human regular insulin with or without inhibitors of phosphatidylinositol 4,5-bisphosphate 3-kinase (PI3-K), mitogen-activated protein kinase (MAPK), nitric oxide synthase (NOS) and guanosine 3',5'cyclic monophosphate(cGMP)-dependent protein kinase (PKG), we measured protein expression (Western blot) and secretion (ELISA) of VEGF. RESULTS: We found that in VSMCs from humans and from insulin-sensitive Zucker fa/+rats, insulin increases VEGF protein expression and secretion, with mechanisms blunted by wortmannin and LY294002 (PI3-K inhibitors), PD98059 (MAPK inhibitor), L-NMMA (NOS inhibitor) and Rp-8pCT-cGMPs (PKG inhibitor). Also the NO donor sodium nitroprusside (SNP) and the cGMP analogue 8-Bromo-cGMP increase VEGF protein expression and secretion, with mechanisms inhibited by wortmannin and PD98059. The insulin effects on VEGF are impaired in VSMCs from Zucker fa/fa rats, which also present a reduced insulin ability to increase NO. CONCLUSIONS: In VSMCs from humans and insulin-sensitive Zucker fa/+rats: (i) insulin increases VEGF protein expression and secretion via both PI3-K and MAPK; (ii) the insulin effects on VEGF are mediated by nitric oxide. The insulin action on both nitric oxide and VEGF is impaired in VSMCs from Zucker fa/fa rats, an animal model of metabolic and vascular insulin-resistance.     

Different effects of selective β1‐adrenoceptor antagonists,nebivolol or atenolol in acetaminophen‐induced hepatotoxicity of rats

Acetaminophen (APAP) overdose is a common cause of acute liver failure, and beta‐blockers are commonly used drugs in clinical practice. This study aimed to evaluate the effect of two different beta‐blocker agents as nebivolol and atenolol against APAP‐induced hepatotoxicity. Male Wistar rats were treated with APAP (2 g/kg/day, p.o.) to induce hepatotoxicity. Our results showed that nebivolol and atenolol reduced heart rate and blood pressure. Nebivolol (5 mg/kg/day, p.o.) for 14 days has a hepatoprotective effect shown by significant decrease in hepatic injury parameters (serum AST and ALT) with significant suppression of hepatic malondialdehyde (MDA) and nitric oxide (NO) which were elevated with APAP administration. Also, nebivolol increased reduced glutathione (GSH) which was reduced with APAP administration. Moreover, immunohistochemical examination revealed that nebivolol treatment markedly reduced inducible nitric oxide synthase (iNOS) expression, while expression of endothelial nitric oxide synthase (eNOS) was markedly enhanced, as compared to APAP group. The protective effects of nebivolol were also verified histopathologically. On the other hand, as compared to APAP group, oral administration of atenolol (50 mg/kg) increased hepatic injury parameters but did not change hepatic NO, MDA, and GSH. In conclusion, this study revealed that nebivolol not atenolol is protective against APAP‐induced hepatotoxicity possibly, in part, through its antioxidant activity, inhibition of iNOS expression, and induction of eNOS expression.     

一氧化氮合酶抑制剂对严重烧伤大鼠的作用研究

目的 :研究一氧化氮合酶 (NOS)抑制剂对严重烧伤大鼠体内一氧化氮 (NO)含量及 NOS表达的影响及其与预后的关系。方法 :复制大鼠重症烧伤模型 ,检测应用非选择性 NOS抑制剂 N 硝基 L 精氨酸甲酯(L NAME)和选择性诱生型 NOS(i NOS)抑制剂氨基胍 (AG)后大鼠血液中 NO代谢产物 (NO- 2 /NO- 3 )以及肺和十二指肠组织中神经型 NOS(n NOS) m RNA的表达水平 ,同时统计各组大鼠的存活率。结果 :烧伤后大鼠血液中 NO- 2 /NO- 3 含量明显增高 ,L NAME和 AG都能抑制 NO2 /NO- 3 的升高 ,L NAME作用更为明显 ;烧伤后 n NOS的 m RNA表达在肺和十二指肠中均有不同程度升高 ,AG和 L NAME使 n NOS表达增加 ,AG作用更为明显 ;L NAME组动物存活时间较对照组显著缩短 ,AG组动物存活时间明显延长。结论 :结构型NOS(c NOS)与 i NOS在烧伤休克病理生理过程中的作用明显不同 ,i NOS活性过度增高与烧伤休克发病关系密切