DDAH/ADMA通路在HDL促进内皮细胞NO生成中的作用

目的:观察高密度脂蛋白(HDL)对氧化低密度脂蛋白(ox-LDL)抑制内皮细胞一氧化氮(NO)生成的保护作用,并探讨其与二甲基精氨酸二甲胺水解酶(DDAH)/非对称性二甲基精氨酸(ADMA)通路的关系.方法:Ox-LDL(1000 g/ml)孵育人脐静脉内皮细胞(HUVECs)24小时或不同浓度的HDL(10、50、100μ g/ml)预处理HUVECs细胞1小时,再与ox-LDL(100μ g/ml)共孵育细胞24小时,收集细胞培养上清液检测NO、ADMA的浓度,收集内皮细胞检测DDAH-Ⅱ的mRNA和蛋白表达以及DDAH的活性.结果:Ox-LDL(100μ g/ml)孵育HUVECs24小时后,细胞培养上清液中NO的浓度显著降低,ADMA水平显著增高,细胞内DDAH-Ⅱ的mRNA和蛋白表达以及DDAH的活性均显著降低.HDL(10、50、100μ g/ml)可以拮抗ox-LDL(100μ g/ml)的上述作用.结论:HDL能显著抑制ox-LDL诱导的内皮细胞NO产生减少,其保护作用与其调节DDAH/ADMA通路有关.     

二甲基精氨酸二甲基氨基水解酶与内皮功能异常

非对称二甲基精氨酸（asymmetric dimethylarginine,ADMA）是一氧化氮合酶（NOS）的内源性竞争性抑制物,竞争性抑制NOS使NO含量减少,引起一系列血管异常效应。二甲基精氨酸Z-甲基氨基水解酶（dimethylarginine dimethylaminohydrolase,DDAH）是主导ADMA代谢的催化酶。DDAH减少或活性降低会使ADMA在体内积累,导致内皮功能异常。本文介绍了DDAH引起内皮功能异常的机制,并对有关影响DDAH活性或表达的药物研究作一概述。     

尾加压素对新生大鼠心肌细胞一氧化氮合成的影响

应用半定量逆转录－多聚酶链反应法,观察尾加压素（urotensin Ⅱ,UⅡ）对培养的新生SD大鼠心肌细胞内皮型一氧化氮合酶（endothelial nitric oxide synthase,eNOS)mRNA表达的影响,并测定UⅡ对心肌细胞内一氧化氮合酶（nitric oxide synthase,NOS)活性和一氧化氮（nitric oxide,NO)释放的影响。结果显示：UⅡ抑制培养的新生大鼠心肌细胞eNOS mRNA表达、抑制NOS的活性及NO释放;0.1μmol/L浓度的UⅡ呈时间依赖性抑制心肌细胞NOS的活性及NO生成。上述实验结果提示UⅡ的心血管作用可能与NO合成系统有关。     

苦荞麦总黄酮对PA诱导脐静脉内皮细胞DDAH_2表达的影响

观察苦荞麦总黄酮对软脂酸(PA)诱导的人脐静脉内皮细胞(EA.hy926)中二甲基精氨酸二甲胺水解酶Ⅱ(DDAH2)表达等指标的影响,探讨苦荞麦总黄酮在胰岛素抵抗信号传导通路中的作用机制。将EA.hy926细胞株常规培养、传代,部分用600μmol/L PA造模后分为对照组、模型组、苦荞麦总黄酮组和二甲双胍组,苦荞麦总黄酮组进一步加入浓度为125 mg/L的苦荞麦总黄酮,二甲双胍组加入浓度为2 mmol/L的二甲双胍。双抗体夹心法测定ADMA含量;硝酸还原酶法测定NO含量;RT-PCR法测定DDAH2mRNA表达;Western-blot测定DDAH2蛋白表达。苦荞麦中、高剂量组与模型组比较,ADMA含量明显减少,NO含量明显增加,均有显著性差异(P0.05);苦荞麦总黄酮组与模型组比较,DDAH2mRNA和蛋白表达量增加,有显著性差异(P0.05)。苦荞麦总黄酮对于PA诱导下EA.hy926细胞中DDAH2、NO的合成具有明显促进作用,对ADMA的合成有明显抑制作用。     

DDAH1参与调控精氨酸代谢介导肿瘤发展的研究进展

代谢重编程是恶性肿瘤的标志之一。调控肿瘤的代谢重编程过程可以用来诊断、监测和治疗癌症。精氨酸在肿瘤生长中发挥重要作用,精氨酸代谢紊乱可能影响肿瘤的进展。双甲基精氨酸水解酶亚基1[N(G),N(G)-dimethylarginine dimethylamino hydrolase 1,DDAH1]可以通过DDAH1/不对称二甲基精氨酸(asymmetric dimethylarginine,ADMA)/一氧化氮(nitric oxide,NO)通路参与调控精氨酸的代谢,进而影响肿瘤的进展。本文主要综述DDAH1代谢通路及其调节机制、DDAH1在肿瘤中的研究进展,以及针对DDAH1靶向分子抑制剂的临床转化研究,旨在系统地展示DDAH1在肿瘤诊断、监测和治疗中的分子病理机制研究进展与临床转化领域的应用前景。     

ADMA-- 一个新的心血管疾病危险因子

非对称性二甲基精氨酸(asymmetric dimethylarginine,ADMA)为内源性一氧化氮合酶(endogenous Nitric-Oxide Synthase,eNOS)竞争性抑制剂,可抑制NOS的活性,减少一氧化氮(nitric oxide,NO)的合成,致使内皮功能障碍。近年来研究认为ADMA是冠心病的独立预测因子,与心功能不全及心血管疾病死亡率明显相关。本文将针对ADMA与心血管疾病的相关性进行综述。     

核因子κB在精氨酸血管升压素诱导大鼠心肌成纤维细胞一氧化氮合成中的作用

本文探讨了精氨酸血管升压素(AVP)刺激下体外培养的大鼠心肌成纤维细胞(CFs)内一氧化氮(NO)含量、一氧化氮合酶(NOS)活性、诱导型一氧化氮合酶基因表达的变化及其与核因子κB(NF-κB)的关系。用胰酶消化法分离培养Sprague Dawley仔鼠的CFs,分别采用硝酸还原酶法、分光光度法、逆转录-聚合酶链式反应(RT-PCR)、免疫荧光-共聚焦显微镜和蛋白质印迹检测AVP干预下CFs的NO含量、NOS活性、iNOS mRNA表达和NF-κB的活化。结果显示,AVP浓度依赖性(0．001—0．1μmol／L)地增加CFs的NO含量,提高NOS活性,增加iNOS mRNA表达;AVP能够活化NF—κB,使其由细胞浆转位于细胞核;NF-κB特异性抑制剂吡咯啉烷二甲基硫脲(PDTC)能够抑制AVP诱导的CFs NO含量增加、NOS活性提高和iNOS mRNA表达增加。上述结果提示,AVP干预下CFs iNOS mRNA表达增加、NOS活性增高、NO合成增多可能通过NF-κB激活途径,NF-κB激活参与心肌纤维化的发生和发展。     

一氧化氮抑制AngⅡ介导的心肌肥大反应的信号机制

本文主要利用培养的新生大鼠心肌细胞,从细胞学及分子生物学角度研究一氧化氮（NO）信号系统在AngⅡ介导的心肌肥大反应中的作用及机制。实验以心肌细胞蛋白合成速率、心房钠尿肽（ANP）的表达作为心肌肥大反应的指标,以硝酸盐及亚硝酸盐含量反映心肌细胞NO水平,以免疫印迹法测定MKP－1蛋白表达,以RT－PCR测定eNOS mRNA水平。结果发现：（1）L－精氨酸（L－Arg)10,100μmol/L分别增加心肌细胞NO水平16％及31％,L－Arg(100μmol/L）还可增加心肌细胞eNOS mRNA表达,其作用可被NOS抑制剂L－NAME所抑制;（2）L－Arg(100μmol/L）可降低AngⅡ(0.1μmol/L）诱导的心肌细胞ANP mRNA表达水平和蛋白合成速率,而在L－Arg处理之前用针对MKP－1的反义寡核苷酸转染心肌细胞,蛋白合成速率明显增加,可取消L－Arg的抑制作用,甚至超过AngⅡ组;（3）L－Arg(100μmol/L）明显增加MKP－1蛋白表达,比对照组增加225％,NOS抑制剂L－NAME及蛋白激酶G（PKG）抑制剂KT－5823皆可抑制L－Arg诱导的MKP－1蛋白表达,分别抑制88％、83％,而AngⅡ能增加L－Arg诱导的MKP－1的表达,较对照组增加365％,增强了L－Arg的作用。以上结果表明,NO抑制AngⅡ介导心肌肥大反应的机制可能是通过激活PKG,促进MKP－1的表达,从而增加MAPK去磷酸化实现的。     

H型高血压合并冠心病患者外周血中EMMPRIN、ADMA、Hcy、MMPs、Treg和Th17细胞水平检测

目的探讨H型高血压合并冠心病患者进行外周血各细胞因子水平检测的意义 方法选取陆军军医大学第一附属医院2015年5月至2018年2月收治的84例高血压合并冠心病患者作为观察组,其中H型高血压合并冠心病患者42例,归为H组,非H型高血压合并冠心病患者42例,归为非H组。随机选取同期我院收治的50例未合并高血压的冠心病患者为冠心病组,50例未合并冠心病的高血压患者为高血压组,45名健康成人为对照组。比较各组外周血EMMPRIN、MMPs、ADMA、Hcy、Treg、Th17细胞水平。采用方差分析和q检验进行统计学分析。 结果H组、非H组、冠心病组、高血压组患者外周血细胞外基质金属蛋白酶诱导因子（EMMPRIN）、基质金属蛋白酶（MMPs）、不对称二甲基精氨酸（ADMA）、Hcy、Th17细胞含量均高于对照组,Treg细胞含量与Treg/Th17的比值低于对照组（P?< 0.05）。H组患者血浆ADMA[（11.01±2.67）μmol/L]、Hcy含量[（17.78±3.32）μmol/L]高于非H组[（7.34±1.34）μmol/L、（6.26±2.17）μmol/L]、冠心病组[（9.11±2.28）μmol/?L、（7.39±3.09）μmol/L]、高血压组[（8.89±1.89）μmol/L、（6.89±2.38）μmol/L],差异有统计学意义（F?= 13.360,8.961,P?< 0.05）。而H组、非H组、冠心病组、高血压组患者外周血单核细胞表面的EMMPRIN表达量、MMPs、Treg细胞、Th17细胞含量与Treg/Th17的比值比较,差异均无统计学意义（P?> 0.05）。 结论H?型高血压合并冠心病患者外周血中EMMPRIN、MMPs、ADMA、Hcy水平升高,Treg/?Th17比值降低,其中ADMA的升高与H型高血压有相关性。     

MAPK信号途径在一氧化氮抑制大鼠心肌肥大中的作用

实验观察了一氧化氮（NO）前体L－精氨酸对肾性高血压大鼠心肌组织eNOS蛋白表达及亚硝酸盐／硝酸盐含量、MKP－1蛋白表达及MAPK活性的影响,以及与心肌肥厚的关系,采用两肾一夹Goldblatt肾性高血压模型,随机分为5组：L－精氨酸高、中、低剂量组,分别于术后第5周给予L－精氨酸50、150及450mg/kg;L－NAME组,腹腔注射L－NAME 10mg/kg,同时给予L－精氨酸150mg/kg;高血压对照组,正常饮水,以及另设的一假手术对照组。用药8周后,用插管法测量大鼠动脉血压、左心室重与体重比值,用胶内原位磷酸化法测MFAPK活性、免疫印迹法检测心肌组织eNOS及MKP－1蛋白表达、酶还原法测定心肌组织亚硝酸盐／硝酸盐－硝酸盐含量。结果表明：（1）L－精氨酸可明显抑制肾动脉狭窄术后的血压升高、左心室重与体重比增加,增加心肌组织eNOS、MKP-1蛋白表达及亚硝酸盐－硝酸盐含量,降低心肌组织MAPK活性,其中以150mg/kg组作用最为明显;（2）NOS抑制剂L－NAME可明显抑制－精氨酸的以上作用,肾性高血压大鼠心肌组织eNOS蛋白表达下降。NO生成减少及MKP－1蛋白表达下降以及MAPK活性增强可能与高血压及心肌厚形成有关,L－精氨酸通过促进心肌组织eNOS蛋白表达、增加NO产生和MKP－1表达、减弱MAPK活性而发挥抗高血压及心肌肥厚的作用。     

Relationship between protective effect of xanthone on endothelial cells and endogenous nitric oxide synthase inhibitors

1,3,5,6-tetrahydroxyxanthone was synthesized. The relationship between protective effect of xanthone on endothelial cells and endogenous nitric oxide synthase inhibitors was investigated. Endothelial cells were treated with ox-LDL (100 microg/mL) for 48 h. Adhesion of monocytes to endothelial cells and release of lactate dehydrogenase (LDH) was determined. Levels of tumor necrosis factor-alpha (TNF-alpha), monocyte chemoattractant protein-1 (MCP-1), nitric oxide (NO) and asymmetric dimethylarginine (ADMA, an endogenous inhibitor of nitric oxide synthase) in conditioned medium and activity of dimethylarginine dimethylaminohydrolase (DDAH) in endothelial cells were measured. Incubation of endothelial cells with ox-LDL (100 microg/mL) for 48 h markedly enhanced the adhesion of monocytes to endothelial cells, increased the release of LDH, the levels of TNF-alpha, MCP-1 and ADMA, and decreased the content of NO and the activity of DDAH. Xanthone (1,3,5,6-tetrahydroxyxanthone) (1, 3 or 10 micromol/L) significantly inhibited the increased adhesion of monocytes to endothelial cells and attenuated the increased levels of LDH, MCP-1 and ADMA induced by ox-LDL. Xanthone (1,3,5,6-tetrahydroxyxanthone) (3 or 10 micromol/L) significantly attenuated the increased level of TNF-alpha and decreased level of NO and activity of DDAH by ox-LDL. The present results suggest that xanthone (1,3,5,6-tetrahydroxyxanthone) preserves endothelial cells and inhibits the increased adhesion of monocytes to endothelial cells induced by ox-LDL, and that the protective effect of xanthone (1,3,5,6-tetrahydroxyxanthone) on endothelial cells is related to reduction of ADMA concentration via increase of DDAH activity.     

Protective effect of HDL on endothelial NO production: the role of DDAH/ADMA pathway

Accumulating studies have demonstrated that the dimethylarginine dimethylaminohydrolase/asymmetric dimethylarginine (DDAH/ADMA) system is a novel pathway for modulating nitric oxide (NO) production. The aim of this study was to investigate whether the protective effect of high density lipoprotein (HDL) on endothelial NO production was related to its effect on DDAH/ADMA pathway. Human umbilical vein endothelial cells (HUVECs) were prior exposed to HDL (10, 50, or 100 μg/ml) for 1 h, and then incubated with oxidized low density lipoprotein (ox-LDL) (100 μg/ml) for 24 h. The cultured medium was collected for measuring the concentration of NO and ADMA. The cells were collected for measuring the mRNA and protein expression of DDAH-II as well as DDAH activity. HUVECs treated with ox-LDL (100 μg/ml) for 24 h significantly decreased the concentration of NO, the mRNA and protein expression of DDAH-II as well as DDAH activity and increased the level of ADMA. Pretreatment with HDL (10, 50, or 100 μg/ml) could counteract these changes induced by ox-LDL (100 μg/ml). HDL significantly increased the attenuated endothelial cell NO production induced by ox-LDL, which was attributed to its effect on DDAH/ADMA pathway.     

Vaspin Increases Nitric Oxide Bioavailability through the Reduction of Asymmetric Dimethylarginine in Vascular Endothelial Cells

Vaspin is an adipocytokine recently identified in the visceral adipose tissue of diabetic rats and having anti-diabetic effects. We have recently shown that vaspin has anti-atherogenic effect through Akt-mediated inhibition of endothelial cell apoptosis. Decreased activity of endothelial nitric oxide synthase (eNOS) plays an important role in the pathogenesis of atherosclerosis. Asymmetric dimethylarginine (ADMA) is a well-known endogenous competitive inhibitor of eNOS and risk factor of cardiovascular diseases. The aim of this study was to examine whether vaspin might protect against atherosclerosis through its beneficial effects on the ADMA-eNOS system. Treatment of vaspin significantly increased NO secretion from endothelial cells and isolated aorta from Sprague-Dawley (SD) rats. Furthermore, treatment of vaspin prevented fatty acid-induced decrease in endothelium-dependent vasorelaxation in isolated aorta of SD rat. For the mechanism of vaspin-induced NO biosynthesis, vaspin activated the STAT3 signaling pathway and stimulated eNOS phosphorylation (Ser 1177), a marker of eNOS activation, through STAT3-dependent mechanism. Furthermore, vaspin treatment increased the expression of dimethylarginine dimethylaminohydrolase (DDAH) II, the responsible enzyme for the degradation of ADMA, leading to a reduction in ADMA levels. Vaspin-induced increase in DDAH II gene expression was through STAT3-mediated stimulation of DDAH II promoter activity. These results suggest that vaspin increases eNOS activity by reducing ADMA level through STAT3-mediated regulation of DDAH II expression. Our findings provide a novel molecular mechanism of antiatherogenic actions of vaspin.     

Dimethylarginine dimethylaminohydrolase-2 overexpression improves impaired nitric oxide synthesis of endothelial cells induced by glycated protein.

Nitric oxide (NO) synthesis is modulated by dimethylarginine dimethylaminohydrolase (DDAH) via metabolizing asymmetric dimethylarginine (ADMA), an endogenous NO synthase (NOS) inhibitor. This study investigated whether glycosylated bovine serum albumin (GBSA) could impair NO synthesis by inhibition of DDAH expression and activity, and whether DDAH2 overexpression could reverse the impaired NO synthesis induced by GBSA in endothelial cells. Overexpression of DDAH2 gene was established by liposome-mediated gene transfection in ECV304 endothelial cell line. Cells were incubated with 1.70 mmol/L GBSA for 48h. And the expressions of DDAH1 and DDAH2, gene activities of DDAH and NOS in cells, as well as concentrations of ADMA and NO in media were assayed. The activity of DDAH and expression of DDAH2 gene but not DDAH1 gene were inhibited in endothelial cells after exposure to GBSA, whereas the concentrations of ADMA were increased concomitantly with the decrease of NOS activity in cells and NO production in media. Overexpression of DDAH2 gene could prevent the inhibition of DDAH activity induced by GBSA (0.55+/-0.02 vs 0.42+/-0.02U/g pro; n=3; P<0.05), decrease ADMA concentration (0.59+/-0.04 vs 1.13+/-0.11 micromol/L; n=3; P<0.01), and increase NOS activity and NO production (53.77+/-3.40 vs 34.59+/-2.57 micromol/L; P<0.05) compared with untransfected cells treated with GBSA. These results suggest that decreased DDAH activity and subsequent elevated endogenous ADMA are implicated in the inhibition of NO synthesis induced by GBSA, and overexpression of DDAH2 gene can prevent these changes in DDAH/ADMA/NOS/NO pathway of endothelial cells exposed to GBSA.     

Involvement of DDAH/ADMA/NOS pathway in nicotine-induced endothelial dysfunction

Asymmetric dimethylarginine (ADMA), an endogenous nitric oxide synthase (NOS) inhibitor, is a key contributor for endothelial dysfunction. Decrease in activity of dimethylarginine dimethylaminohydrolase (DDAH), a major hydrolase of ADMA, causes accumulation of ADMA under cardiovascular abnormalities. The study was to determine whether nicotine-induced endothelial dysfunction is related to modulating DDAH/ADMA/NOS pathway. Four-week oral nicotine treatment (5 mg/kg/day) significantly increased the plasma level of ADMA and decreased aortic DDAH expression as well as impaired endothelial function in Sprague-Dawley rats. Similarly, the medium levels of both ADMA and lactate dehydrogenase were markedly elevated in umbilical vein endothelial cells (HUVECs) treated with nicotine (10 microM) for 48 h. Nicotine-induced endothelial damages were markedly attenuated by L-arginine or overexpression of DDAH-II. Nicotine greatly downregulated both mRNA and protein levels of DDAH-II, and decreased DDAH activity in HUVECs. HUVECs express alpha7 nicotinic acetylcholine receptor (alpha7 nAChR), whose antagonists could block these effects of nicotine mentioned above. Intracellular Ca2+ chelator did not affect nicotine-induced decrease in DDAH-II mRNA level. In conclusion, nicotine modulates DDAH/ADMA/NOS pathway of endothelial cell via activation of alpha7 nAChR, which may be involved in endothelial dysfunction associated to smoking.     

Dimethylarginine dimethylaminohydrolase activity modulates ADMA levels,VEGF expression,and cell phenotype

Asymmetric dimethylarginine (ADMA) is an endogenous inhibitor of nitric oxide synthase and is metabolised by dimethylarginine dimethylaminohydrolase (DDAH). Elevated levels of circulating ADMA correlate with various cardiovascular pathologies less is known about the cellular effects of altered DDAH activity. We modified DDAH activity in cells and measured the changes in ADMA levels, morphological phenotypes on Matrigel, and expression of vascular endothelial growth factor (VEGF). DDAH over-expressing ECV304 cells secreted less ADMA and when grown on Matrigel had enhanced tube formation compared to untransfected cells. VEGF mRNA levels were 2.1-fold higher in both ECV304 and murine endothelial cells (sEnd.1) over-expressing DDAH. In addition the DDAH inhibitor, S-2-amino-4(3-methylguanidino)butanoic acid (4124W 1mM), markedly reduced human umbilical vein endothelial cell tube formation in vitro. We have found that upregulating DDAH activity lowers ADMA levels, increases the levels of VEGF mRNA in endothelial cells, and enhances tube formation in an in vitro model, whilst blockade of DDAH reduces tube formation.     

Mechanisms of homocysteine-induced oxidative stress

Hyperhomocysteinemia decreases vascular reactivity and is associated with cardiovascular morbidity and mortality. However, pathogenic mechanisms that increase oxidative stress by homocysteine (Hcy) are unsubstantiated. The aim of this study was to examine the molecular mechanism by which Hcy triggers oxidative stress and reduces bioavailability of nitric oxide (NO) in cardiac microvascular endothelial cells (MVEC). MVEC were cultured for 0-24 h with 0-100 microM Hcy. Differential expression of protease-activated receptors (PARs), thioredoxin, NADPH oxidase, endothelial NO synthase, inducible NO synthase, neuronal NO synthase, and dimethylarginine-dimethylaminohydrolase (DDAH) were measured by real-time quantitative RT-PCR. Reactive oxygen species were measured by using a fluorescent probe, 2',7'-dichlorofluorescein diacetate. Levels of asymmetric dimethylarginine (ADMA) were measured by ELISA and NO levels by the Griess method in the cultured MVEC. There were no alterations in the basal NO levels with 0-100 microM Hcy and 0-24 h of treatment. However, Hcy significantly induced inducible NO synthase and decreased endothelial NO synthase without altering neuronal NO synthase levels. There was significant accumulation of ADMA, in part because of reduced DDAH expression by Hcy in MVEC. Nitrotyrosine expression was increased significantly by Hcy. The results suggest that Hcy activates PAR-4, which induces production of reactive oxygen species by increasing NADPH oxidase and decreasing thioredoxin expression and reduces NO bioavailability in cultured MVEC by 1) increasing NO2-tyrosine formation and 2) accumulating ADMA by decreasing DDAH expression.     

DDAH1 Deficiency Attenuates Endothelial Cell Cycle Progression and Angiogenesis

Asymmetric dimethylarginine (ADMA) is an endogenous inhibitor of nitric oxide (NO) synthase (NOS). ADMA is eliminated largely by the action of dimethylarginine dimethylaminohydrolase1 (DDAH1). Decreased DDAH activity is found in several pathological conditions and is associated with increased risk of vascular disease. Overexpression of DDAH1 has been shown to augment endothelial proliferation and angiogenesis. To better understand the mechanism by which DDAH1 influences endothelial proliferation, this study examined the effect of DDAH1 deficiency on cell cycle progression and the expression of some cell cycle master regulatory proteins. DDAH1 KO decreased in vivo Matrigel angiogenesis and depressed endothelial repair in a mouse model of carotid artery wire injury. DDAH1 deficiency decreased VEGF expression in HUVEC and increased NF1 expression in both HUVEC and DDAH1 KO mice. The expression of active Ras could overcome the decreased VEGF expression caused by the DDAH1 depletion. The addition of VEGF and knockdown NF1 could both restore proliferation in cells with DDAH1 depletion. Flow cytometry analysis revealed that DDAH1 sRNAi knockdown in HUVEC caused G1 and G2/M arrest that was associated with decreased expression of CDC2, CDC25C, cyclin D1 and cyclin E. MEF cells from DDAH1 KO mice also demonstrated G2/M arrest that was associated with decreased cyclin D1 expression and Akt activity. Our findings indicate that DDAH1 exerts effects on cyclin D1 and cyclin E expression through multiple mechanisms, including VEGF, the NO/cGMP/PKG pathway, the Ras/PI3K/Akt pathway, and NF1 expression. Loss of DDAH1 effects on these pathways results in impaired endothelial cell proliferation and decreased angiogenesis. The findings provide background information that may be useful in the development of therapeutic strategies to manipulate DDAH1 expression in cardiovascular diseases or tumor angiogenesis.     

Protective effect of neferine on endothelial cell nitric oxide production induced by lysophosphatidylcholine: the role of the DDAH-ADMA pathway

Neferine, extracted from the seed embryo of Nelumbo nucifera Gaertn., has multiple cardiovascular pharmacological effects. The dimethylarginine dimethylaminohydrolase (DDAH) - asymmetric dimethylarginine (ADMA) system is a novel pathway for modulating nitric oxide (NO) production. The aim of this study was to investigate whether the protective effect of neferine on endothelial NO production was related to the DDAH-ADMA pathway. Human umbilical vein endothelial cells (HUVECs) were first exposed to neferine (0.1, 1.0, or 10.0?μmol/L) for 1?h, and then incubated with lysophosphatidylcholine (LPC; 10?μg/mL) in the presence of neferine for 24?h. The medium was collected for measuring the levels of NO, maleic dialdehyde (MDA), as well as ADMA. The endothelial cells were collected for measuring DDAH activity and the level of reactive oxygen species (ROS). LPC significantly decreased NO concentration and DDAH activity and increased the levels of ADMA, ROS, and MDA. Neferine could partially counteract the changes induced by LPC. These findings suggested that neferine could modulate the DDAH-ADMA pathway via its antioxidant properties, which was involved in its beneficial effect on endothelial NO production.     

Piceatannol is more effective than resveratrol in restoring endothelial cell dimethylarginine dimethylaminohydrolase expression and activity after high-glucose oxidative stress

Glucose-induced oxidative stress is involved in endothelial dysfunction. Dimethylarginine dimethylaminohydrolase (DDAH) and arginase are regulators of the endothelial NO synthase (eNOS). This study aimed to compare the effect of two polyphenolic antioxidants, resveratrol and piceatannol, on DDAH and arginase pathways in bovine aortic endothelial cells under 25 mM glucose for 24 h. DDAH activity and expression were decreased in these cells as compared to control cells, whereas arginase activity was unchanged. DDAH inhibition led to intracellular accumulation of asymmetric dimethylarginine (ADMA), a natural inhibitor of eNOS. Under these conditions, cell pre-treatment with resveratrol (0.1-10 μM) restored basal DDAH activity and ADMA level with a dose-dependent effect. Piceatannol acted as resveratrol on DDAH pathway but at 10-fold lower concentrations. Resveratrol and piceatannol restored DDAH activity even in the presence of splitomicin, a specific inhibitor of Sirtuin 1. These results suggest potential therapeutic intervention targeting resveratrol or piceatannol administration to improve endothelial dysfunction.