妊高征脐动脉内皮细胞损伤与一氧化氮合酶的变化及意义

妊娠高血压综合征(PIH,简称妊高征)是严重危害母婴健康的妊娠并发症,病因至今未明。我们观察了PIH脐动脉内皮细胞损伤程度与ET、NO和一氧化氮合酶(NOS)的变化,旨在探讨血管内皮细胞损伤在PIH发病中的意义。1资料与方法1.1研究对象PIH组选择我院住院病人33例。其诊断及分型标准参考文献犤1犦进行。其中轻度PIH12例;中度PIH11例;重度PIH10例。正常晚妊NLP组17例,均为同期住院健康孕妇。年龄23～34岁,平均25.2±3.1岁。两组均排除肝、肾疾病、糖尿病、慢性高血压,也无其它内科合并症。1.2标本采集和处理两组均在产前取孕妇空腹静…     

妊高征患者血浆一氧化氮变化特点

一氧化氮(NO)化学结构简单、性质活跃,广泛参与机体生理、病理调节。在妊娠期,NO可降低血管对加压物质的反应性、降低外周阻力及血压。目前认为,NO产生减少是妊高征发病的重要原因。本研究通过测定妊高征患者血浆NO的含量,并设正常晚期妊娠妇女为对照组,旨在探讨NO与妊高征之间的关系。1资料与方法1·1分组妊高征组:选自2006年1~6月在本院住院分娩的重度妊高征患者,诊断标准以《妇产科学》第4版为准,共30例。年龄23~34岁,平均孕周38·5周。有慢性高血压、糖尿病、肾病或心血管病等合并征的患者均被除外。对照组:同期本院住院分娩的健康孕妇,共30例,年龄22~32岁,平均孕周40·2周。1·2标本采集在分娩前取肘静脉血3 ml(妊高征患者在使用硫酸镁前采血)。经2 000转/min离心15 min后,取血浆置-30℃低温冰箱待测。1·3检测方法NO的最终代谢产物是亚硝酸盐(NO2-)、硝酸盐(NO3-)。本实验通过检测孕妇血中NO2-/NO3-,间接测量NO的含量。取试管3只,分为测定管、空白管和质控管。每管各加2 g镉粒。镉粒预先以0·1 mol/L盐酸和稀氨缓冲液洗涤,各反应物加完后充分混匀。反应60 mi...     

内皮型一氧化氮合酶DNA多态性与妊高征

妊高征（pregnancy induced hypertension,PIH）是妊娠晚期出现的特发性高血压综合征．其发病率高达7～10％。但一个多世纪以来．有关其病因及发病机制还不甚清楚．这给妊高征的预防和治疗带来了困难。自1992年来,Zeeman等提出血管内皮细胞产生的内皮舒张因子一氧化氮（NO）,在维持正常血管调节及妊高征发病中起一定作用后．NO的研究成为热门课题,截至目前．人们对NO的理化特性、生物活性及作用机理有了较充分认识．由于NO在生物体内极其活跃,难以调控,使人们为之困惑。可以说．现有任何增加或抑制NO合成的治疗措施都不可避免地存在严重副作用。随着后基因时代的到来．人们普遍认识到从基因水平阐明NO代谢异常的发生机理以寻求合理的调控方法是有必要的．于是人们纷纷把目光转向NO代谢异常的遗传学基础的研究。本文就内皮型一氧化氮合酶基因及其DNA多态性与妊高征关系的许多重要发现综述如下。     

ACE基因多态性与原发性高血压患者血清ACE和血浆AngⅡ水平的相关性

目的研究血管紧张素转化酶(ACE)基因插入/缺失(I/D)多态性与原发性高血压(EH)患者血清ACE、血浆血管紧张素Ⅱ(AngⅡ)水平的相关性。方法选择青岛地区EH患者246例和130例正常对照,测定两组血压、血脂、血糖等临床及生化指标,并对高血压进行分级,同时检测血清ACE、血浆AngⅡ水平,采用聚合酶链反应(PCR),检测ACE基因型,比较不同基因型、不同血压分级患者其血清ACE、血浆AngⅡ水平有无差别。结果EH组中,II、ID、DD基因型患者血清ACE水平分别为(36.69±14.05)U/L,(42.98±16.61)U/L,(49.37±17.43)U/L,组间差异有统计学差异(P<0.05);三组基因型患者血浆AngⅡ水平比较,组间差异无统计学差异(P>0.05);随着高血压分级的升高,血清ACE及血浆AngⅡ水平逐渐增加,差异均有统计学意义(P<0.05);EH患者血清ACE水平与ACE基因多态性之间有相关性(r=0.324,P<0.05)。结论 DD基因型EH患者血清ACE水平明显高于ID型和II型患者,血浆AngⅡ水平在不同基因型间无统计学差异。     

尿调素在妊高征中的变化

通过对42例妊高征妇女及103例正常对照组妇女尿中尿调素（UrM）水平的测定，探讨妊高征与UrM之间的关系。结果显示：正常对照组UrM水平随孕周的增长而增加，≥36孕周时增长极为显著（P＜0．01），而妊高征组孕妇UrM水平较同孕周对照组明显下降（P＜0．01），下降程度与妊高征严重程度呈正相关，但产后24～72小时内UrM水平两组间即无差异。说明UrM与妊高征之间存在一定相关性。     

妊高征与尿酸、血脂水平的相关性探讨

目的进一步研究妊高征与高尿酸血症,高血脂的关系及其临床意义。方法检测173例妊娠期各期孕妇及164例正常非孕健康女性的血清尿酸三酰甘油胆固醇水平。结果妊娠孕妇组的血清尿酸水平(367.8±58.7)μmol/L,正常非孕健康组(242.6±43.6)μmol/L三酰甘油水平分别为(1.30±0.25)mmol/L,(1.56±0.30)mmol/L,胆固醇水平(3.95±0.55)mmol/L与(4.53±0.95)mmol/L;妊娠组中高尿酸血症孕妇发生妊高征的比率83.95%(68/81)明显高于尿酸值正常孕妇28.26%(26/92)。妊高征组明显高于对照组,胆固醇水平差异有极显著性t=13.86,P<0.01,三酰甘油水平差异有显著性(t=8.62,P<0.01)。结论妊娠期女性比正常非孕期女性血清尿酸水平升高,妊娠期高尿酸血症发生妊高征的比率明显增加。     

妊高征与血管紧张素转换酶基因多态性的相关性研究

目的探讨妊娠期高血压综合征（pregnancy-induced hypertension,PIH）与血管紧张素转换酶（ACE）基因多态性的关系。方法运用多聚酶链反应技术检测61例PIH患者及胎儿和70例正常妊娠妇女及胎儿血管紧张素转换酶基因多态。结果妊娠妇女两组中DD型、ID型Ⅱ型基因频率分别比较,D型等位基因分别比较差异有统计学意义;胎儿两组中DD型、ID型Ⅱ型基因频率分别比较,D型等位基因分别比较均无统计学差异。结论PIH患者ACE基因多态性与PIH的发病有关;PIH胎儿ACE基因多态性与PIH的发病无关。     

ACE基因多态性与冠心病的相关性研究

目的测定中国汉族人群血管紧张素转换酶基因插入/缺失(I/D)多态性,研究其在中国汉族人群中的分布以及与冠心病发生的关系。方法冠心病患者146例,均经冠状动脉造影确诊;正常对照组113例,选自人群健康查体的随机个体。应用聚合酶链反应-限制性片断长度多态性分析,聚丙稀酰胺凝胶电泳方法检测ACE基因插入/缺失(I/D)多态性。结果①中国汉族人群血管紧张素转换酶基因有Ⅱ、ID、DD三种基因型;②ACE基因I/D多态性与冠心病相关,具有DD基因型比具有Ⅱ基因型的人患冠心病的危险性增加2.06倍。结论①中国汉族人群血管紧张素转换酶基因有Ⅱ、ID、DD三种基因型;②血管紧张素转换酶基因插入/缺失(I/D)多态性与冠心病相关。     

心电图变化与妊高征病情严重程度的相关性分析

目的分析研究在不同严重程度妊娠期高血压综合征患者中心电图的特点及差异。方法选择2010年8月至2013年3月我院妊娠期高血压综合征患者作为临床研究对象,所有患者均经过临床明确诊断,其中轻度42例,中度40例,重度35例。所有患者均给予心电图检查,并比较不同严重程度患者的心电图特点。结果心电图异常以重度妊高征患者的检出率最高,组间对比差异明显,P<0.05,差异有统计学意义;患者心电图异常主要为窦性心动过速、窦性心律不齐等,各心电图异常发生率对比差异明显,P<0.05,差异有统计学意义。结论妊高征患者会有心电图的异常变化,且患者心电图异常情况不同,需要临床医师给予高度重视。     

妊高征患者应用钙剂与内皮素相关性探讨

目的 :探讨妊高征患者应用钙剂预防或治疗时ET水平的变化。以及对妊高征病情的影响。方法 :选择100例妊娠24～26周预测妊高征试验阳性的孕妇。首次测量血清钙平均(2 20±0 3)mmol/L、血清内皮素 (ET)平均 (58 06±9 78)ng/L、平均动脉压(12 5±0 3)kpa。随机分为两组。50例口服葡萄糖酸钙片1～2g/日 ,治疗的病人为观察组 ;50例未口服葡萄糖钙片治疗的病人为对照组。12周后复查血清钙、血清ET、平均动脉压。结果 :观察100例预测妊高征阳性的病人 ,12周以后妊娠达36～38周。观察组和对照组的血清钙 (2 20±0 2)、(1 97±0 3)mmol/L、血清ET(61 76±18 6)、(132±705 64)ng/L、平均动脉压(12 7±0 3)、(13 6±0 3)kpa。两组比较差异有显著性P<0 05。结论 :血清钙与血清ET平均动脉压之间成负相关系。妊高征患者应用钙剂可达到预防和治疗的作用 ,对妊高征患者的病情有显著影响     

内皮型一氧化氮合酶基因G894T多态性与ACEI降压疗效相关性研究

目的探讨原发性高血压患者内皮型一氧化氮合酶基因(eNOS)G894T多态性与血管紧张素转换酶抑制剂(ACEI)降压疗效之间的关系。方法 281例1-2级原发性高血压患者经安慰剂治疗2周后,血压符合入选标准者分别给予咪达普利5～10mg/d或贝那普利10～20mg/d,治疗6周。用酚-氯仿法提取基因组DNA,采用PCR结合限制性内切酶片段长度多态性(RFLP)方法检测eNOS基因型,分析不同基因型组与ACEI降压疗效的关系。结果本研究中eNOS 3种基因型频率分别为GG66.2％、GT27.8％、TT6.0％,等位基因G、T的频率分别为0.80和0.20。ACEI治疗后,eNOS基因GG、GT、TT组收缩压下降值分别为(14.51±11.19)mmHg,(16.11±12.31)mmHg,(12.38±14.30)mmHg;舒张压下降值分别为(8.95±6.56)mmHg,(9.00±6.92)mmHg,(9.37±4.80)mmHg,3组间收缩压与舒张压下降值均无统计学差异。结论 eNOS基因G894T多态性与ACEI降压疗效无关。     

Beneficial effect of the oligomerized polyphenol oligonol on high glucose-induced changes in eNOS phosphorylation and dephosphorylation in endothelial cells

Background and purpose:

Hyperglycaemia is known to reduce nitric oxide (NO) bioavailability by modulating endothelial NO synthase (eNOS) activity, and polyphenols are believed to have cardiovascular benefit. One possible mechanism could be through interaction with eNOS.

Experimental approach:

The effects of the oligomerized polyphenol oligonol on eNOS phosphorylation status and activity were examined in porcine aortic endothelial cells cultured in high glucose concentrations.

Key results:

Exposure to high glucose concentrations strongly inhibited eNOS phosphorylation at Ser-1177 and dephosphorylation at Thr-495 in bradykinin (BK)-stimulated cells. These inhibitory effects of high glucose were significantly prevented by treatment with oligonol. Akt and p38 mitogen-activated protein kinase (MAPK) were activated in BK-stimulated cells. High glucose inhibited Akt activation but enhanced p38 MAPK activation, both of which were reversed by oligonol treatment. The phosphatidylinositol 3-kinase inhibitor wortmannin blocked the reversal by oligonol of phosphorylation at Ser-1177, but not dephosphorylation at Thr-495, in BK-stimulated cells exposed to high glucose. The effect of oligonol on BK dephosphorylation under high glucose was mimicked by protein kinase C (PKC) ε-neutralizing peptides. These data suggest that the effects of oligonol on high glucose-induced attenuation of eNOS Ser-1177 phosphorylation and Thr-495 dephosphorylation may be regulated by Akt activation and PKCε inhibition respectively. Oligonol also prevented high glucose-induced attenuation of BK-stimulated NO production.

Conclusions and implications:

Oligonol prevented the impairment of eNOS activity induced by high glucose through reversing altered eNOS phosphorylation status. This mechanism may underlie the beneficial cardiovascular health effects of this oligomerized polyphenol.     

The effect of puerarin on serum nitric oxide concentration and myocardial eNOS expression in rats with myocardial infarction

Puerarin (1) is a major effective ingredient extracted from the traditional Chinese medicine Ge-gen (Radix Puerariae, RP). Recently, puerarin has been used to treat patients with coronary artery diseases (CAD). However, the mechanisms of puerarin on CAD are still not very clear. In this study, we investigated the role of puerarin on serum nitric oxide (NO) concentration, myocardial endothelial nitric oxide synthase (eNOS) gene expression, the protein expression of eNOS and inducible nitric oxide synthase (iNOS), as well as the level of protein kinase B (Akt/PKB) phosphorylation in rats with myocardial infarction. We found that puerarin (120 mg/kg/day, i.p.) could increase serum nitrite concentration in rat with myocardial ischemia (MI). It also induced the gene expression or activation of eNOS, protein expression of eNOS, and the Akt/PKB phosphorylation. From these results, we suggested that puerarin could increase serum nitric oxide level of rat with myocardial infarction, which should be one of the mechanisms of the therapeutic effect of puerarin on CAD. The increased expression of eNOS and the Akt/PKB pathway may be the underlying mechanism by which puerarin stimulates NO production.     

The effect of puerarin on serum nitric oxide concentration and myocardial eNOS expression in rats with myocardial infarction

Puerarin (1) is a major effective ingredient extracted from the traditional Chinese medicine Ge-gen (Radix Puerariae, RP). Recently, puerarin has been used to treat patients with coronary artery diseases (CAD). However, the mechanisms of puerarin on CAD are still not very clear. In this study, we investigated the role of puerarin on serum nitric oxide (NO) concentration, myocardial endothelial nitric oxide synthase (eNOS) gene expression, the protein expression of eNOS and inducible nitric oxide synthase (iNOS), as well as the level of protein kinase B (Akt/PKB) phosphorylation in rats with myocardial infarction. We found that puerarin (120 mg/kg/day, i.p.) could increase serum nitrite concentration in rat with myocardial ischemia (MI). It also induced the gene expression or activation of eNOS, protein expression of eNOS, and the Akt/PKB phosphorylation. From these results, we suggested that puerarin could increase serum nitric oxide level of rat with myocardial infarction, which should be one of the mechanisms of the therapeutic effect of puerarin on CAD. The increased expression of eNOS and the Akt/PKB pathway may be the underlying mechanism by which puerarin stimulates NO production.     

大鼠对氟西泮抗痫耐受和依赖时海马中一氧化氮合酶的变化

目的通过测定大鼠对氟西泮抗痫耐受性和依赖性时海马中一氧化氮合酶(NOS)蛋白表达及活性的变化,探讨一氧化氮(NO)在此过程中可能的作用。方法建立大鼠对氟西泮抗痫耐受性和依赖性的模型。运用免疫印迹及免疫组化法检测海马中NOS蛋白表达,以及比色法检测NOS活性的变化。结果大鼠对氟西泮抗痫耐受组的海马中NOS蛋白表达明显高于对照组;而对氟西泮抗痫依赖组则与对照组差别无统计学意义。两组NOS活性均显著高于各自的对照组。结论NO可能是介导氟西泮抗痫耐受性和依赖性的因素之一。     

Antioxidant and angiotensin converting enzyme (ACE) inhibitory activities of ethanol extract and pure flavonoids from Adinandra nitida leaves

Context: Adinandra nitida Merr. ex. H.L. Li (Theaceae) is an indigenous plant in south China. Its leaves have been reported to have many curative effects such as reducing blood pressure, as well as antibacterial, antioxidant, and analgesic properties, which could be used in foods and medicines.

Objective: The antioxidant and angiotensin converting enzyme (ACE) inhibitory activities of the main flavonoids and ethanol extract (EE) of A. nitida leaves were investigated for the first time.

Materials and methods: The main flavonoids of A. nitida leaves (camellianin A, camellianin B) were prepared and their contents in EE were determined by HPLC. The antioxidant activities of the samples were measured by DPPH radical scavenging assay and Rancimat test. The ACE inhibitory activities of the samples were carried out by using an assay kit with hippuryl-glycyl-glycine as substrate.

Results: The contents of camellianin A, camellianin B and apigenin in EE were determined as 41.98, 2.67, and 1.73%, respectively. The antioxidant activities of the flavonoids were far lower than that of EE in DPPH radical scavenging and Rancimat assays. However, the ACE-inhibitory activities of camellianin A, camellianin B and apigenin were higher than that of EE.

Discussion and conclusion: The flavonoid content of EE was more than 45%. The high activities of EE in DPPH scavenging and Rancimat assay could be mainly attributed to compounds other than flavonoids. However, the ACE-inhibitory activity of EE could be mainly attributed to the presence of the flavonoids.     

Effects of angiotensin receptor blockers on endothelial nitric oxide release: the role of eNOS variants

AIM

Angiotensin II receptor blockers (ARBs) improve endothelial cell (EC)-dependent vasodilation in patients with hypertension through suppression of angiotensin II type 1 receptors but may have additional and differential effects on endothelial nitric oxide (NO) synthase (eNOS) function. To investigate this question, we tested the effects of various ARBs on NO release in ECs from multiple donors, including those with eNOS genetic variants linked to higher cardiovascular risk.

METHODS

The effects of ARBs (losartan, olmesartan, telmisartan, valsartan), at 1 µm, on NO release were measured with nanosensors in human umbilical vein ECs obtained from 18 donors. NO release was stimulated with calcium ionophore (1 µm) and its maximal concentration was correlated with eNOS variants. The eNOS variants were determined by a single nucleotide polymorphism in the promoter region (T-786C) and in the exon 7 (G894T), linked to changes in NO metabolism.

RESULTS

All of the ARBs caused an increase in NO release as compared with untreated samples (P < 0.01, n = 4–5 in all eNOS variants). However, maximal NO production was differentially influenced by eNOS genotype. Olmesartan increased maximal NO release by 30%, which was significantly greater (P < 0.01, n = 4–5 in all eNOS variants) than increases observed with other ARBs.

CONCLUSIONS

The ARBs differentially enhanced NO release in ECs in a manner influenced by eNOS single nucleotide polymorphisms. These findings provide new insights into the effects of ARBs on EC-dependent vasodilation and eNOS function.     

四逆汤对失血性休克家兔一氧化氮（NO）和一氧化氮合酶（NOS）的影响

目的：探讨中药四逆汤对失血性休克家兔一氧化氮的影响。方法：采用动物分组对照实验，测量不同状态下动物血浆一氧化氮和一氧化氮合酶(NOS)的变化。结果：与休克前比较，休克组的家兔血浆一氧化氮水平各时段均明显提高(P＜0．01)，而治疗组仅在治疗后30min家兔一氧化氮及NOS水平明显提高(P＜0．01)，其他时段与休克前比较，未见显著差异(P＜0．01)，且休克后lh、4h、8h休克组一氧化氮水平显著高于治疗组(P＜0．01)。经药物治疗的失血性休克家兔血浆一氧化氮水平明显降低。     

Relationship of endothelial nitric oxide synthase gene polymorphism with blood pressure, lipid profile and blood glucose level

To study the relationship of the polymorphism of endothelial nitric oxide synthase (eNOS) gene and blood pressure, lipid profiles and blood glucose level. By using PCR-RFLP, the eNOS Glu298Asp gene polymorphism was detected in 184 patients with essential hypertension and 196 matched healthy individuals with normal blood pressure. Taking into account eNOS Glu298Asp polymorphisms, the relationship of blood pressure with triglycerides (TG), total cholesterol (TC), high density lipoprotein (HDL), low density lipoprotein (LDL) and blood glucose level was analyzed. The distribution of eNOS Glu298Asp polymorphism had no significant difference between different blood pressure groups and gender groups, but there was a significant difference between different age groups, diastolic blood pressure groups or BMI groups (P < 0.05). Asp/Asp genotype significantly increased the risk of hypertension in individuals with serum TC above 5.4 mmol/L (P = 0.03, OR = 2.65). eNOS Glu298Asp polymorphism and serum lipid could synergistically modulate the blood pressure. eNOS Asp/Asp genotype could significantly increase the risk of hypertension in individuals with serum TC over 5.4 mmol/L. eNOS Glu298Asp in combination with serum TC could be used to predict the risk of hypertension. __________ Translated from Journal of Third Military Medical University, 2007, 29(5): 448–450 [译自: 第三军医大学学报]     

内外源性EETs对血管内皮一氧化氮合酶的表达及其在Thr-495位磷酸化的作用

目的　研究细胞色素P4 5 0表氧化酶基因转染和直接加入EETs对内皮细胞eNOS表达及其在Thr 4 95位磷酸化的影响。方法　在原代培养的牛主动脉内皮细胞中 ,分别加入生理浓度的 8,9 EET(1 0 0nmol·L-1 )、1 1 ,1 2 EET(1 0 0nmol·L-1 )、1 4 ,1 5 EET(1 0 0nmol·L-1 )孵育 4h ,或直接用重组腺相关病毒介导的花生四烯酸表氧化酶转染牛主动脉内皮细胞2wk ,以产生内源性EETs ,用Westernblot法检测总eNOS蛋白的表达及eNOSThr 4 95磷酸化的水平 ;此外 ,从大鼠尾静脉注射真核表达质粒pCB6、pCB6 2C1 1OR、pCB6 2J2和pCB6 F87V ,2wk后检测大鼠主动脉总eNOS表达及eNOSThr 4 95磷酸化的水平。结果　与空白和溶媒组比较 ,外源性EETs明显促进内皮细胞总eNOS表达 ,增加eNOSThr 4 95的磷酸化 ,而CYP4 5 0抑制剂 (1 7 ODYA)则可明显降低eNOS表达和eNOSThr 4 95的磷酸化水平 ;与相应的对照组比较 ,体内和体外转染不同的表氧化酶基因均能明显上调内皮细胞eNOS的表达 ,增加eNOSThr 4 95的磷酸化水平。结论　EETs和CYP表氧化酶不仅能明显促进血管内皮细胞总eNOS蛋白的表达 ,而且还通过其翻译后修饰来增加其Thr 4 95位蛋白磷酸化水平