内皮源性一氧化氮合酶的活性调节

内皮源性一氧化氮合酶(eNOS)代谢精氨酸产生的一氧化氮(NO)在调节血管的稳态中发挥着重要作用,内皮功能失调所致疾病多与NO的释放和活性受损有关。近年来关于eNOS活性词节的研究很多,机制主要包括乙酰化修饰所决定的亚细胞定位,蛋白-蛋白相互作用以及磷酸化修饰。许多体液因子和机械刺激可通过不同的方式调节eNOS活性。为了更好地认识eNOS的活性调节,本文就该领域最新的研究进展进行归纳总结。     

老年2型糖尿病肾病患者血清C肽与尿白蛋白排泄率的关系

目的研究老年2型糖尿病肾病患者血清C肽水平与尿白蛋白排泄率的关系。方法选取老年2型糖尿病患者44例,分别测定血清C肽水平、糖化血红蛋白(HbA1c)、空腹血糖及尿白蛋白排泄率,并据尿白蛋白排泄率分为正常白蛋白尿组、微量白蛋白尿组、临床白蛋白尿组。结果临床白蛋白尿组血清C肽水平低于其他两组,差异有显著性意义(P<005)。血清C肽水平与尿白蛋白排泄率呈负相关(r=-0629,P<001)。结论血清C肽水平的下降可能参与老年2型糖尿病肾病的发生发展。     

内皮型一氧化氮合酶基因多态性与糖尿病肾病的关系

目的 探讨内皮型一氧化氮合酶(eNOS)基因27bp数目可变的串联重复序列(VNTR)多态性与糖尿病肾病(DN)的关系。方法 应用PCR方法检测了32名健康对照者与84例2型糖尿病(T2DM)患者的eNOS基因a／b基因型，用硝酸盐还原酶法测定上述人群空腹血清一氧化氮代谢物(NOx)水平。并根据24h尿白蛋白排泄率(UAER)将84例T2DM患者分为正常白蛋白尿(UAlb)组(DMl)，微量UAlb组(I)M2)和大量UAlb组(I)M3)。结果 (1)T2DM各组a等位基因频率显著高于对照组，T2DM各组aa ab基因型频率明显高于对照组。DMl组a等位基因频率及aa ab基因型频率高于DM2及DM3组，但差异无显著意义。(2)NC组aa ab基因型空腹血清NOx明显低于bb基因型。(3)血清NOx在DMl组较NC组有明显的升高，但在DM2、DM3组却较NC组显著降低。(4)T2DM患者中，aa ab基因型UAER和血清肌酐(Scr)水平显著高于bb基因型。结论 eNOS基因27bpVNTR多态性与DN发生有关，eNOS基因a等位基因是DN发生和发展的有用的预测标志。     

内皮源性一氧化氮合酶基因多态性的研究进展

内皮源性 NOS(e NOS)产生的 NO可扩张血管、调节血压、改变局部血流、抑制血小板聚集、抗平滑肌细胞的增殖。选择性神经元性 NOS(n NOS)抑制剂因不影响 e NOS活性但有效减弱n NOS活性 ,故在局灶性脑缺血模型中表现出神经保护作用。由于 e NOS表现出的良性循环保护作用 ,诸如 e NOS基因结构和表达及其与心脑肺及周围血管病变的发病的相关性的研究 ,正得到广泛的关注。1　 e NOS基因结构及多态性人类 e NOS是一个复杂的基因位点 ,它含有 2 6个外显子 ,2 5个内含子 ,全长 2 1kb,位于人染色体 7q3 5 - 3 6区域。e NOS常见的突变有…     

氯沙坦对实验性糖尿病大鼠尿白蛋白排泄的影响

目的 观察血管紧张素受体拮抗剂氯沙坦（ｌｏｓａｒｔａｎ）对糖尿病大鼠白蛋白排泄的影响。方法 雄性ＳＤ大鼠分为正常对照组（ＮＣ）、糖尿病对照组（ＤＣ）、糖尿病氯沙坦治疗组（ＤＬ）和糖尿病培哚普利治疗组（ＤＰ）。检测各组第４、８、１２、１６周尿白蛋白排泄变化。结果 ＤＬ组较ＤＣ组尿白蛋白排泄显著下降（Ｐ〈０．０１）,和ＤＰ组比较无显著性差异。结论 血管紧张素受体拮抗剂氯沙坦能有效降低糖尿病大鼠尿白蛋白的排泄。     

内皮源性一氧化氮合酶基因G894T多态性与脑梗死的关联性研究

目的　探讨内皮源性一氧化氮合酶 (e NOS)基因 G894T多态性与脑梗死的关系。方法　取病例组 1 4 8例 ,对照组 1 0 9例 ,各例抽取新鲜静脉血 5 ml,提取 DNA后 ,通过 PCR- RFL P法进行基因分型。结果　病例组和对照组各基因型分布 GG、GT、TT分别是 83.1 % ,1 4 .2 % ,2 .7% ,对照 90 .8% ,9.2 % ,0 .0 0 % (χ2 =3.1 78,P=0 .0 75) ,符合 Hardy- Weinberg平衡 ,两组统计学上无明显差异。但 T等位基因频率在病例组中较对照组明显升高 (P=0 .0 2 7) ,相对危险度 1 .33(95% CI:0 .61 7～ 0 .92 2 )。病例组中 ACE D等位基因频率亦较对照组明显升高 (P=0 .0 1 7)。病例组 T等位基因携带者的血浆胆固醇水平较 GG纯合子患者明显升高 (P=0 .0 2 5) ,而在对照组中并无此差异。结论　 e NOS基因 G894T突变可能是国人脑梗死的遗传学危险因素 ,其致病机理可能与 ACE D缺失型及血浆胆固醇水平有协同作用。     

糖尿病患者尿白蛋白排泄率与骨密度相关性研究

为探讨糖尿病患者尿白蛋白排泄率（AER）与骨密度（BMD）的相关性,测定了106例糖尿病患者和20例正常人（对照组）24小时AER,根据糖尿病患者AER分为Ⅰ组即正常蛋白尿组、Ⅱ组即微量白蛋白尿组、Ⅲ组即大量白蛋白尿组,并分别测定其L2～4椎体、股骨近端BMD。结果显示,除Ⅰ、Ⅱ组男性患者腰椎BMD与对照组无明显差异外,其余各组、各部位BMD与对照组相比均明显下降（P＜0.01）;男女糖尿病AER正常患者分别与同性别对照组相比,其BMD亦降低（P＜0.01）。糖尿病患者中,Ⅲ组BMD明显低于Ⅰ、Ⅱ组（P＜0.05）。提示糖尿病肾病患者BMD比正常人低,大量蛋白尿组BMD较其它组显著下降。     

老年糖尿病患者血清Apelin水平与尿白蛋白排泄率相关性分析

目的探讨老年糖尿病患者血清Apelin水平与尿白蛋白排泄率的相关性,为老年糖尿病肾病的预治提供临床依据。方法59例年龄在65岁以上的糖尿病患者和30例同年龄段健康对照者,采用酶联免疫吸附法测定空腹血清Apelin含量,同时检测空腹血糖及24小时尿白蛋白排泄率（UAER）,分析其相关性。结果59例糖尿病患者血清Apelin值为473±64．34,UAER为87．34±44．32,两者呈在非常显著的正相关（r=0．384,P〈0．01）。而对照组两指标无相关性。同时糖尿病组和对照组的血清Apelin水平与血糖值之间也无明显相关性。结论老年糖尿病患者血清apelin升高与UAER密切相关,提示血清Apelin水平可能是糖尿病肾病发生的因素之一。     

Ⅱ型糖尿病尿转铁蛋白与白蛋白排泄率的比较研究

Ⅱ型糖尿病尿转铁蛋白与白蛋白排泄率的比较研究陈宇钱荣立国外的临床观察表明，在糖尿病早期微量白蛋白尿出现前已有部分病人有尿转铁蛋白的升高，因此推测在诊断糖尿病肾病方面，它比尿白蛋白更敏感［１～３］。本实验通过对Ⅱ型糖尿病病人肌酐清除率、尿白蛋白与尿转铁...     

胰岛素样生长因子1与2型糖尿病尿白蛋白排泄率关系的临床研究

目的 探讨尿和血胰岛素样生长因子1(IGF-1)水平与2型糖尿病(T2DM)尿白蛋白排泄率的关系.方法 根据尿白蛋白排泄率(UAER)将80例T2DM患者分为正常白蛋白尿组(N-UAlb)32例,微量白蛋白尿组(M-UAlb)28例和大量白蛋白尿组(L-UAlb)20例.另设对照(NC)组20名.检测4组尿和血IGF-1水平.结果 (1)与NC组比较,T2DM三组血IGF-1明显降低,三组间血IGF-1水平无明显差异;(2)N-UAlb组尿IGF-1与NC组相比无明显差异;M-UAlb组和L-UAlb组尿IGF-1排泄较N-UAlb组显著增高;(3)尿IGF-1排泄与UAER和尿视黄醇结合蛋白排泄率呈正相关; 结论 2型糖尿病肾病尿IGF-1排泄的增加,提示可能有肾组织局部合成和分泌IGF-1的增加,后者可能是2型糖尿病肾病的发病机制之一.     

内皮型一氧化氮合酶在早期糖尿病大鼠肾组织中表达的定量研究

目的探讨早期糖尿病大鼠肾组织内N0含量变化与一氧化氮合酶（eNOS）表达之间的关系。方法健康雄性SD大鼠60只随机分成糖尿病模型（DM）组和正常对照（NC）组,每组30只。检测两组大鼠的生化指标,检测肾皮质内NO的含量用亚硝酸还原酶法,肾皮质eNOS的含量用Western blot法。结果与同期NC组比较,（1）第1、2、4周DM组大鼠血糖显著升高（P〈0．01）;（2）DM组肾组织内NO含量在第1周时下降（3．52±0．45VS4．23±0．38）,P〈0．01;第2、4周时显著升高（P〈0．01）,第4周时升高最为明显（9．82±0．54VS4．02±0．32）,P〈0．01;（3）Western blot分析显示eNOS含量在第1周时下降,第2～4周时逐渐升高。结论早期糖尿病大鼠肾组织内NO含量逐渐升高,eNOS逐渐增多,提示在糖尿病早期肾组织内N0含量升高主要由eNOS增加引起。     

SIRT1 promotes endothelium-dependent vascular relaxation by activating endothelial nitric oxide synthase

Reduced caloric intake decreases arterial blood pressure in healthy individuals and improves endothelium-dependent vasodilation in obese and overweight individuals. The SIRT1 protein deacetylase mediates many of the effects of calorie restriction (CR) on organismal lifespan and metabolic pathways. However, the role of SIRT1 in regulating endothelium-dependent vasomotor tone is not known. Here we show that SIRT1 promotes endothelium-dependent vasodilation by targeting endothelial nitric oxide synthase (eNOS) for deacetylation. SIRT1 and eNOS colocalize and coprecipitate in endothelial cells, and SIRT1 deacetylates eNOS, stimulating eNOS activity and increasing endothelial nitric oxide (NO). SIRT1-induced increase in endothelial NO is mediated through lysines 496 and 506 in the calmodulin-binding domain of eNOS. Inhibition of SIRT1 in the endothelium of arteries inhibits endothelium-dependent vasodilation and decreases bioavailable NO. Finally, CR of mice leads to deacetylation of eNOS. Our results demonstrate that SIRT1 plays a fundamental role in regulating endothelial NO and endothelium-dependent vascular tone by deacetylating eNOS. Furthermore, our results provide a possible molecular mechanism connecting the effects of CR on the endothelium and vascular tone to SIRT1-mediated deacetylation of eNOS.     

Aminoguanidine reduces regional albumin clearance but not urinary albumin excretion in streptozotocin-diabetic rats

Summary Advanced glycation end-product-formation is thought to play a role in the development of diabetic angiopathy. By altering the structure of different extracellular matrix components advanced glycation end-products might affect vascular and glomerular permeability. In this study we investigated the effect of treatment with an inhibitor of advanced glycation end-product-formation, aminoguanidine, on vascular permeability and the development of albuminuria in streptozotocin-induced diabetic rats. Male Wistar Rp rats were randomized into a control group, a diabetic group, and an aminoguanidine-treated diabetic group. After 8 weeks, 24-h urine collections were taken and rats were implanted with an arterial and a venous catheter. Mean arterial blood pressure was determined by intra-arterial measurement. Regional albumin clearances were assessed in the eye, ileum, lung, skeletal muscle and skin using an isotope technique. Mean arterial pressure in the diabetic group was significantly lower in the control and aminoguanidine-treated groups (p<0.02). Regional albumin clearances were significantly increased in all tissues of diabetic rats compared to control rats (p<0.05). Aminoguanidine treatment of diabetic rats resulted in a significant decrease of regional albumin clearance in all tissues except the lung (p<0.05, lung p=0.07). The development of albuminuria in diabetic rats however, was not affected by aminoguanidine.     

Gene transfer of endothelial nitric oxide synthase reduces angiotensin II-induced endothelial dysfunction

Angiotensin II stimulates vascular NADPH oxidase to produce superoxide, which can react with nitric oxide and impair vasomotor function. We tested the hypothesis that the overexpression of endothelial nitric oxide synthase (eNOS) or superoxide dismutase (SOD) would correct angiotensin II-induced endothelial dysfunction. We examined the effects of the gene transfer of eNOS or 2 isoforms of SOD to the aorta in angiotensin II-treated rabbits on vasomotor function. New Zealand White rabbits were treated for 1 week with angiotensin II (100 ng. kg(-1). min(-1)) or saline by osmotic minipumps. In angiotensin II-treated rabbits, mean blood pressure was 107+/-8 mm Hg; it was 67+/-5 mm Hg in saline-infused rabbits (P<0.05). In aortas from angiotensin II-treated rabbits, lucigenin-enhanced chemiluminescence demonstrated a 2.5-fold increase in superoxide levels, and the oxidative fluorescent probe hydroethidine indicated increased superoxide levels throughout the vascular wall, especially in the endothelium and adventitia. Maximal relaxation to acetylcholine was less in aortas from rabbits treated with angiotensin II (72+/-5% versus 87+/-4% in saline-treated rabbits; P<0.01), but responses to sodium nitroprusside were similar. Segments of the thoracic aorta were incubated in vitro with an adenoviral vector that expressed eNOS, copper zinc SOD (CuZnSOD), extracellular SOD (ECSOD), or beta-galactosidase. beta-Gal treatment with adenovirus containing the gene for eNOS (AdeNOS) but not adenovirus containing the gene for beta-gal (Adbeta-gal) (control virus) restored responses to acetylcholine (82+/-3% after AdeNOS and 67+/-4% after Adbeta-gal). Gene transfer of CuZnSOD or ECSOD did not improve the endothelium-dependent relaxation of the aorta in rabbits that received angiotensin II. Thus, gene transfer of eNOS, but not SOD, effectively restores vasomotor function in angiotensin II-infused rabbits.     

cGMP-mediated negative-feedback regulation of endothelial nitric oxide synthase expression by nitric oxide

Earlier studies have demonstrated that nitric oxide (NO) exerts a fast-acting inhibitory influence on endothelial NO synthase (eNOS) enzymatic activity in isolated vascular tissue preparations. The present study was designed to examine the possible effect of NO on eNOS protein expression in cultured endothelial cells and intact animals. Human coronary endothelial cells were incubated with S-nitroso-N-acetyl-penicillamine (SNAP, an NO donor), oxyhemoglobin (HGB, an NO trapping agent), SNAP plus HGB, or inactive vehicle (control). In other experiments, cells were treated with 3-isobutyl-1-methylxanthine (a phosphodiesterase inhibitor), 1H-[1,2, 4]oxadiazolo-[4,3-2]quinoxalin-1-one (ODQ, a guanylate cyclase inhibitor), SNAP plus ODQ, 8-bromo-cGMP (8-Br-cGMP, a cell-permeable cGMP compound), 8-Br-cGMP plus HGB, or inactive vehicle in order to discern the effect of cGMP. The incubations were conducted for 24 hours, and total nitrate plus nitrite production and eNOS protein abundance (Western analysis) were measured. To determine the effect of NO on eNOS expression in vivo, rats were treated with either the NO donor isosorbide dinitrate or placebo by gastric gavage for 48 hours, and aortic eNOS protein expression was examined. The NO donor SNAP markedly depressed, whereas the NO scavenger HGB significantly raised, eNOS protein expression. The downregulatory action of SNAP was completely abrogated by HGB. Phosphodiesterase inhibitor and 8-Br-cGMP downregulated, whereas the guanylate cyclase inhibitor ODQ upregulated eNOS protein expression. The downregulatory action of SNAP was completely overcome by the guanylate cyclase inhibitor ODQ, and the upregulatory action of the NO scavenger HGB was abrogated by 8-Br-cGMP. Administration of NO donor resulted in a marked downregulation of aortic eNOS protein expression in intact animals, thus confirming the in vitro findings. NO serves as a negative-feedback regulator of eNOS expression via a cGMP-mediated process.     

Reversal of endothelial nitric oxide synthase uncoupling and up-regulation of endothelial nitric oxide synthase expression lowers blood pressure in hypertensive rats.

OBJECTIVES: We sought to examine the hypothesis that a pharmacologic up-regulation of endothelial nitric oxide synthase (eNOS) combined with a reversal of eNOS uncoupling provides a protective effect against cardiovascular disease. BACKGROUND: Many cardiovascular diseases are associated with oxidant stress involving protein kinase C (PKC) and uncoupling of eNOS. METHODS: Messenger ribonucleic acid (mRNA) expression was analyzed with RNase protection assay or quantitative real-time polymerase chain reaction, vascular nitric oxide (NO) with spin trapping, and reactive oxygen species (ROS) with dihydroethidium fluorescence. RESULTS: Aortas of spontaneously hypertensive rats (SHR) showed an elevated production of ROS when compared with aortas of Wistar-Kyoto rats (WKY). The aortic expression of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase subunits (Nox1, Nox2, Nox4, and p22phox) was higher in SHR compared with WKY. In SHR, aortic production of ROS was reduced by the NO synthase inhibitor N(G)-nitro-L-arginine methyl ester (L-NAME), indicating eNOS "uncoupling" in hypertension. Oral treatment with the PKC inhibitor midostaurin reduced aortic Nox1 expression, diminished ROS production, and reversed eNOS uncoupling in SHR. Aortic levels of (6R)-5,6,7,8-tetrahydro-L-biopterin (BH4) were significantly reduced in SHR compared with WKY. Midostaurin normalized BH4 levels in SHR. In both WKY and SHR, midostaurin increased aortic expression of eNOS mRNA and protein, stimulated bioactive NO production, and enhanced relaxation of the aorta to acetylcholine. Midostaurin lowered blood pressure in SHR and, to a lesser extent, in WKY; the compound did not change blood pressure in WKY made hypertensive with L-NAME. CONCLUSIONS: Pharmacologic interventions that combine eNOS up-regulation and reversal of eNOS uncoupling can markedly increase bioactive NO in the vasculature and produce beneficial hemodynamic effects such as a reduction of blood pressure.     

内皮细胞型一氧化氮合酶基因多态性与糖尿病肾病的相关性

糖尿病肾病是糖尿病 (ＤＭ )致死致残的主要原因之一。近年来流行病学调查显示 ,有些ＤＭ患者虽血糖长期控制不良 ,但并不发生ＤＮ ,而有些患者虽代谢控制良好 ,最终仍发生ＤＮ〔1,2〕,提示遗传因素在ＤＮ的发生发展中可能起关键作用。分子水平的研究发现 ,内皮细胞型一氧化氮合酶(ｅＮＯＳ)基因第 7外显子的Ｇｌｕ2 98Ａｓｐ(894Ｇ→Ｔ)基因点突变及第 4内含子一个 2 7ｂｐ的插入 /缺失 (ａ/ｂ)多态与糖尿病微血管并发症及冠心病、心肌梗死的发生有明显相关性〔1 5〕,但该点突变及多态与糖尿病肾病的相关性各家报道不一〔6,7〕,特别是该突…