Nitric oxide synthase gene transfer for erectile dysfunction in a rat model

OBJECTIVE: To determine whether over-expression of nitric oxide synthase (NOS) in the corpus cavernosum of the penis improves erectile function, as NO is an important transmitter for genitourinary tract function, mediating smooth muscle relaxation and being essential for penile erection. MATERIALS AND METHODS: The inducible form of the enzyme NOS (iNOS) was introduced into the corpus cavernosum of adult Sprague-Dawley rats (250-300 g) by injecting a solution of plasmid, adenovirus or adenovirus-transduced myoblast cells (adeno-myoblasts). Plasmid, adenovirus and adeno-myoblasts encoding the expression of the beta-galactosidase reporter gene were also injected into rats. RESULTS: Throughout the corpora cavernosum there was expression of beta-galactosidase after injecting each of the three solutions. Maximum staining was greatest for adeno-myoblast, then adenovirus and then plasmid. The mean (sd) basal intracavernosal pressure (ICP) of iNOS-treated animals (adenovirus and adeno-myoblast) increased to 55 (23) cmH2O, compared with naive animals with a basal ICP of 5 (6) cmH2O (P = 0.001). Stimulating the cavernosal nerve (15 Hz, 1.5 ms, 10-40 V, 1 min) resulted in a doubling of the ICP (adenovirus and adeno-myoblast) from the basal level of the iNOS-treated animals. Direct in situ measurement of NO showed the release of 1-1.3 micro mol/L in the adeno-myoblast penis. CONCLUSION: Myoblast-mediated gene therapy was more successful for delivering iNOS into the corpus cavernosum than direct adenovirus injection or plasmid transfection. Surprisingly, implanting muscle cells into the penis is not only feasible but also beneficial. Gene therapy for NOS may open new avenues of treatment for erectile dysfunction. Control of NOS expression would be necessary to prevent priapism.     

Effects of the T-786C,G894T,and Intron 4 VNTR (4a/b) polymorphisms of the endothelial nitric oxide synthase gene on the risk of prostate cancer

Several studies have shown that nitric oxide (NO) and nitric oxide synthase (NOS) system plays an important role in carcinogenesis. Endothelial nitric oxide synthase (eNOS) gene polymorphisms significantly affects serum NO concentrations. Studies addressing the relationship between eNOS gene polymorphisms and prostate cancer (CaP) are very scarce. We examined the association between the 3 eNOS gene polymorphisms (T-786C, G894T, and 4a/b) with risk and clinical features of CaP. One hundred seventy patients with CaP (mean age 63.6 ± 12.4 years) and 340 age-matched healthy controls (mean age 64.9 ± 12.9 years) were recruited in this case-control study. Genotyping was performed by polymerase chain reaction restriction fragment length polymorphism (PCR-RLFP) technique. For T-786C polymorphism, we found that CC genotype was associated to CaP risk [odds ratio (OR) = 3.62, 95% confidence interval (CI): 1.89–7.74, P = 0.002), high grade tumor (OR = 2.46, 95% CI:1.78–4.72; P = 0.006), and advanced disease (OR = 4.67, 95% CI: 2.64?8.61; P = 0.002). Neither the CaP risk nor clinical features of CaP were associated with the G894T polymorphism. It was found that, compared with 4a/b bb genotype, the 4a/b “a” variant genotypes were associated with an increased risk of CaP in an allele dose dependent manner (OR = 2.12, 95% CI: 1.68–3.44; P = 0.031 for 4a/b ab genotype, and OR = 4.32, 95% CI: 2.21–6.08; P = 0.001 for 4a/b aa genotype). In addition, genotypes with the “a” allele of the eNOS 4a/b polymorphism predispose the patients to high grade (OR = 4.76, 95% CI: 2.74–8.62; P = 0.001) and advanced CaP (OR = 5.28, 95% CI: 3.64–8.72; P = 0.001). Furthermore, the T-Asp-b and C-Asp-b haplotypes were associated with a significantly decreased risk of CaP (OR = 0.44, 95% CI: 0.33–0.77; P = 0.004, and OR = 0.39, 95% CI: 0.26–0.61; P = 0.001, respectively). We found significant differences in genotype distribution and allelic frequencies between CaP patients and controls for the T-786C, and 4a/b eNOS polymorphisms.     

Association of endothelial nitric oxide synthase gene polymorphisms with end-stage renal disease: a systematic review and meta-analysis

Background and objective: Endothelial nitric oxide synthase (eNOS) is one of the potent regulators of intra renal hemodynamics. Polymorphisms of eNOS gene may be involved in the progression of renal disease, and may be the causative factors that contribute to the deterioration of renal functions. During the past decades, several studies investigated the association of eNOS polymorphisms with the risk of end-stage renal disease (ESRD), but the results remain unclear and the mechanisms are not defined. Our study was designed to examine the role of different eNOS genetic polymorphisms in the progression of ESRD. Materials and methods: Relevant studies were identified through PubMed, Embase, Medline and CNKI (China National Knowledge Infrastructure) database published between January 2000 and November 2013. The association between eNOS polymorphisms and ESRD susceptibility was assessed by pooled odds ratios (ORs) and 95% confidence intervals (95% CI) in fixed or random effects models. Results: Sixteen articles were identified for the analysis of association between eNOS gene polymorphisms and ESRD risk. A total of 2729 patients and 2190 controls for 4b/a, 851 patients and 1171 controls for G894T, and 513 patients and 487 controls for T786C were included in our analysis. Overall, 4a allele of 4b/a polymorphism produced a significant association in the global population (OR?=?1.47, 95% CI?=?1.05–2.06, p?=?0.03) in a random-effect model; T allele of G894T was also significantly associated with ESRD susceptibility in overall populations (OR?=?2.12, 95% CI?=?1.44–3.12, p?=?0.0001). Furthermore, 4a and T carriers were significantly associated with ESRD risk as well. No association was found between T786C polymorphism and ESRD. Conclusion: The evidence accumulated suggested that 4b/a and G894T polymorphisms in the eNOS gene were associated with ESRD susceptibility, indicating that 4a and T allele carriers might become significant genetic molecular markers for the onset of ESRD in overall populations. However, more studies should be performed in the further studies.     

Analysis of endothelial nitric oxide synthase (eNOS) G894T polymorphism and semen parameters in a Chinese Han population

Previous studies have shown that endothelial nitric oxide synthase (eNOS) gene may be involved in abnormal semen parameters. However, the relationship between eNOS G894T polymorphism and semen parameters remains controversial. The purpose of this study was to investigate the association of eNOS G894T polymorphism and semen parameters. The genotype frequency of eNOS G894T was determined in 270 idiopathic asthenozoospermia patients and 248 ethnically matched healthy volunteers using iPLEX genotyping assays on a MassARRAY^® (Sequenom, San Diego, CA, USA) platform. The statistical analysis performed with Fisher's exact test showed no significant difference in frequencies of genotypes between both groups. The logistic regression showed that genotypes GT, TT and allele T were nonassociated with increased risk of asthenozoospermia in the patient group with ≤5% or >5% sperm with normal forms. The dependence on genotypes of semen parameters was further investigated in both patients and control group. There was no significant difference as compared to control group (P > 0.05). Our study indicated that eNOS gene G894T polymorphism may not have an adverse effect on semen parameters in a Chinese Han population.     

Significance of nitric oxide and its modulation mechanisms by endogenous nitric oxide synthase inhibitors and arginase in the micturition disorders and erectile dysfunction

Abstract:   Evidence indicates that nitric oxide (NO) deficiency contributes to micturition disorders, especially in the afferent pathway and erectile dysfunction (ED). Two possible causes of NO deficiency are substrate ( l -arginine) limitation and increased levels of endogenous inhibitors of NO synthase (particularly asymmetric dimethylarginine: ADMA) in plasma and tissues. Elevated tissues of ADMA and N^G-monomethyl-L-arginine (L-NMMA) have been reported to be associated with impaired NO-mediated urethral, trigonal and cavernosal relaxations by pelvic ischemia. Also, plasma ADMA may help to identify underlying cardiovascular disease in men with ED. Decreased l -arginine availability to NO synthase is due to the shunting of l -arginine into other pathways such as arginase. Interaction between NO synthase and arginase has been reported to be involved in NO-mediated urethral and prostatic relaxations. Also, increased arginase activity in cavernosal tissues likely contributes to the ED that accompanies diabetes mellitus and aging. Therefore, arginase inhibition has been reported to enhance the NO-dependent physiological process for erectile function.     

Inhibition of inducible nitric oxide synthase improved erectile dysfunction in rats with type 1 diabetes

Diabetes mellitus (DM), which is closely related to microvascular dysfunction, is a risk factor for erectile dysfunction (ED). Furthermore, the upregulation of inducible nitric oxide synthase (iNOS) is associated with systemic vascular dysfunction in rats with diabetes. The purpose of this study was to investigate the role of iNOS in diabetes mellitus erectile dysfunction (DMED). First, we developed a type 1 DM rat model using streptozotocin and selected those that developed DMED. Then, we injected these rats with the 1400W, an iNOS inhibitor, for 10 weeks and subsequently assessed their ED. Lastly, we performed various molecular studies and histopathological analyses of penile tissues collected from these rats after the experiments. Through the histopathological studies, we also found that the treatment restored the ratios of the smooth muscle to collagen fibres, delayed the development of microvascular injury and alleviated the oxidative stress caused by hyperglycaemia. Based on these results, we confirmed that upregulation of iNOS leads to microvascular dysfunction in patients with ED. Overall, we found that inhibition of iNOS displayed beneficial effects in the treatment of ED, suggesting that its mechanism should be further explored.     

eNOS gene T786C,G894T and 4a4b polymorphisms and male infertility susceptibility: a meta‐analysis

The association between polymorphism of eNOS and male infertility in several studies was controversial. To explore a more precise estimation of the association, a meta‐analysis of eight case–control studies, including 1,968 cases and 1,539 controls, were selected. The meta‐analysis was conducted by calculating the pooled odds ratio (OR) with a 95% confidence interval (95% CI). Overall, the association between T786C and risk of male infertility was obvious (TC vs. TT: OR, 1.20; 95% CI, 1.01–1.42; CC vs. TT: OR, 3.37; 95% CI, 1.65–6.87; TC/CC vs. TT: OR, 1.47; 95% CI, 1.25–1.73; CC vs. TT/TC: OR, 3.18; 95% CI, 1.54–6.56; TC vs. TT: OR, 1.65; 95% CI, 1.27–2.03). However, no overall association was observed between the other two polymorphisms of eNOS (G894T and 4a4b) and male infertility. Stratified analysis showed that significantly strong association between T786C polymorphism and semen quality was present in all three types of male infertility (azoospermia, oligozoospermia and asthenozoospermia). In the subgroup analysis based on ethnicity, both T786C and 4a4b could influence the risk of male infertility in Asian and Caucasian. Further studies of polymorphisms of eNOS with their biological functions are needed to understand the role in the development of male infertility.     

Association of endothelial nitric oxide synthase gene intron 4 polymorphism with end-stage renal disease

BACKGROUND: Nitric oxide (NO) released from endothelial cells is related to the maintenance of physiological vascular tone. The impairment of endothelial NO generation brought about by gene polymorphism is considered one of the deterioration factors in progressive renal disease. In the endothelial nitric oxide synthase (eNOS) intron 4 polymorphism, the presence of the aa genotype has been associated with cardiovascular and renal disease. The aim of this study was to investigate the presence of eNOS gene intron 4 polymorphism in patients with end-stage renal disease (ESRD). METHODS: A total of 114 patients and 94 controls were studied. DNA specimens were extracted from blood and amplified by polymerase chain reaction. The alleles were separated by agarose gel electrophoresis. Genotype distribution and allele frequencies were compared between groups using the chi-squared test. RESULTS: Statistical analysis revealed that the frequency of the eNOS4 genotype aa was significantly different in ESRD patients and in controls (P=0.016, OR=2.07, CI 95%: 1.14-3.74). There was also a statistically significant difference between ESRD patients and controls regarding allele carriers (P=0.004; OR=2.26; CI 95%: 1.29-3.96). When the frequencies of allele carriers in the diabetic nephropathy group and in the control group were compared, a significant difference was found (P=0.034, OR=2.28; CI 95%: 1.04-5.00). CONCLUSION: This study showed a strong correlation between eNOS4a polymorphism and end-stage renal disease.     

Effects of icariin on erectile function and expression of nitric oxide synthase isoforms in castrated rats

Aim： To investigate the effect of icariin on erectile function and the expression of nitric oxide synthase （NOS） isoforms in castrated rats. Methods： Thirty-two adult male Wistar rats were randomly divided into one shamoperated group （A） and three castrated groups （B, C and D）. One week after surgery, rats were treated with normal saline （groups A and B） or oral icariin （1 mg/[kg·day] for group C and 5 mg/[kg·day] for group D） for 4 weeks. One week after treatment, the erectile function of the rats was assessed by measuring intracavernosal pressure （ICP） during electrostimulation of the cavernosal nerve. The serum testosterone （ST） levels, the percent of smooth muscle （PSM） in trabecular tissue, and the expression of mRNA and proteins of neuronal nitric oxide synthase （nNOS）, inducible nitric oxide synthase （iNOS）, endothelial nitric oxide synthase （eNOS） and phosphodiesterase V （PDES） in corpus cavernosum （CC） were also evaluated. Results： ICP, PSM, ST and the expression of nNOS, iNOS, eNOS and PDE5 were significantly decreased in group B compared with those in group A （P 〈 0.01）. However, ICE PSM and the expression of nNOS and iNOS were increased in groups C and D compared with those in group B （P 〈 0.05）. Changes in ST and the expression of eNOS and PDE5 were not significant （P 〉 0.05） in groups C and D compared with those in group B. Conclusion： Oral treatment with icariin （〉 98.6 % purity） for 4 weeks potentially improves erectile function. This effect is correlated with an increase in PSM and the expression of certain NOS in the CC of castrated rats. These results suggest that icariin may have a therapeutic effect on erectile dysfunction.     

内皮型一氧化氮合成酶基因Glu298Asp多态性与糖尿病肾脏病的关系

目的探讨内皮型一氧化氮合成酶（eNOS）基因Glu298Asp多态性与糖尿病肾脏病（DIcD）的关系。方法采用聚合酶链反应（PCR）后限制性内切酶消化技术,对在我院就诊的326例中国汉族2型糖尿病（T2DM）患者和215名健康体检者进行Glu298Asp基因型分析。根据尿白蛋白排泄率（UAER）水平将T2DM患者分为3组：UAER正常组（DM组）102例,微量白蛋白尿组（MAU组）81例,临床肾病组（DKD组）143例。选择同时期本院健康体检者215名为对照组。用多因素Logistic回归分析法判定DKD的独立危险因素。结果DKD组与对照组Glu298Asp基因型频率分布存在显著性差异（GG72．72％、GT26．57％、TT0．70％比GG83．41％、GT16．10％、TT0．49％,P=0．019）,而DM组和MAU组则与对照组无差异（P〉0．05）。Logistic回归分析显示,年龄、平均动脉压、GT基因型是DKD发生的独立危险因素。结论eNOS Glu298Asp基因与中国汉族T2DM患者DKD具有相关性。     

Endothelial nitric oxide synthase gene polymorphisms in patients with nontraumatic femoral head osteonecrosis.

As endothelial nitric oxide synthase (eNOS) has beneficial effects on skeletal, vascular, and thrombotic systems, the association between nontraumatic femoral head osteonecrosis (FHON) and eNOS gene polymorphisms was investigated in Korean patients with FHON. Genomic DNA from 103 patients with nontraumatic FHON (idiopathic in 50, steroid-induced in 29, and alcohol abuse in 24) and 103 control subjects matched for gender and age (3-year range) was analyzed for the 27-bp repeat polymorphism in intron 4 and Glu298Asp polymorphism in exon 7. The frequencies of alleles and genotypes were compared between patients and control subjects. The frequency of 4a allele was significantly higher in total patients than control subjects [6.8% vs. 2.4%, p = 0.0345, odds ratio (OR) 2.931]. In subgroup analysis, the 4a allele significantly increased in patients with idiopathic FHON versus control subjects (9.0% vs. 2.4%, p = 0.0297, OR 3.976). The frequency of the 4a/b genotype in total patients (13.6% vs. 4.9%, p = 0.0302, OR 3.083) as well as patients with idiopathic FHON (18.0% vs. 4.9%, p = 0.0246, OR 4.302) was higher than control subjects. The distribution of Glu298Asp polymorphisms was not significantly different between patients and control subjects. Microstellate polymorphism in intron 4 of eNOS polymorphism was significantly associated with idiopathic FHON in Korean patients. Because 4a allele is associated with lower synthesis of eNOS, these results suggest that carrier state of 4a allele in intron 4 might be a genetic risk factor of FHON and could provide insight into the protective role of nitric oxide in the pathogenesis of FHON.     

Endothelial nitric oxide synthase gene intron 4 a/b VNTR polymorphism in children with APSGN

The role of endothelial nitric oxide synthase gene intron 4 a/b (eNOS4a/b) variable number of tandem repeats (VNTR) polymorphism in various renal diseases was investigated. We investigated whether the eNOS4a/b VNTR polymorphism is associated with susceptibility to acute poststreptococcal glomerulonephritis (APSGN) and its clinical features. Endothelial NOS4a/b VNTR polymorphism is determined by the polymerase chain reaction in 60 children with APSGN, and 66 healthy controls. The genotype distribution of eNOS4 does not differ between the patients and the controls (X²=5.1, p=0.079). However, the frequency of eNOS4a (eNOS4a/a and eNOS4a/b) genotype is higher in the patients than in the controls (X²=4.5, p=0.046). In the APSGN group we performed renal biopsy on eight patients because of nephrotic syndrome accompanies acute nephritic syndrome or glomerular filtration rate (GFR) is lower than 50% of normal, and found that to carry a/a and a/b genotypes were a significant risk factor for this type presentation (OR=17.3, 95% CI:1.95-152.67, p=0.03). Mean serum creatinine values are found statistically significantly higher in a/a and a/b genotypes when compared with b/b genotypes (p=0.022). Children carrying the “aa” and “ab” genotype or “a” allele of eNOS4 have a greater tendency to develop and clinical presentation of APSGN.     

Genetic association study of the GNB3 C825T,the ACE I/D and the eNOS G894T polymorphisms and the risk to develop erectile dysfunction in a German ED population

Erectile dysfunction (ED) is often associated with cardiovascular disorders such as hypertension, coronary heart disease, hypercholesterolaemia and diabetes mellitus. The genotypes in the GNB3 C825T, the ACE I/D and the eNOS G894T polymorphisms have been identified as genetic risk factors for cardiovascular disorders. The association between the genotypes in these polymorphisms and the risk to develop ED was analysed. In 455 German ED patients and 111 age‐matched healthy controls genotyping in the candidate polymorphisms was performed after DNA extraction from whole blood. Association studies between the genotype distribution in the control group in comparison with the ED‐group and age of onset of the disease as well as erectile response to intracorporal prostaglandin injection in dependence of candidate polymorphism genotype were performed using the SPSS‐Software®. Genotype distribution of the GNB3 C825T, the ACE I/D and the eNOS G894T polymorphisms was similar in the ED population and the healthy control group. The age of onset of the disease as well as the erectile response to intracorporal prostaglandin injection was independent of the genotypes in the three candidate polymorphisms. In contrast to the previous studies in this analysis, the risk to develop ED is not influenced by the genotypes in the GNB3 C825T, the ACE I/D and the eNOS G894T polymorphisms.     

Plasma levels of endothelin-1, angiotensin II, nitric oxide and prostaglandin E in the venous and cavernosal blood of patients with erectile dysfunction

OBJECTIVES: To determine the alterations in the plasma levels of endothelin-1, angiotensin II, nitric oxide (NO) and prostaglandin E(2) (PGE(2)) in the venous and cavernosal blood of patients with organic and psychogenic erectile dysfunction (ED). PATIENTS, SUBJECTS AND METHODS: The study included 32 patients complaining of ED; they were subdivided into two equal groups with either organic or psychogenic ED. Fifteen healthy potent age-matched male volunteers were enrolled as a control group. For each patient, venous and cavernosal blood samples were obtained, while venous blood was obtained from the controls. RESULTS: There were significantly greater mean plasma levels of endothelin-1 and angiotensin II, and significantly lower mean plasma levels of NO and PGE(2), in the venous blood of patients with ED than in the controls. Patients with organic ED had significantly higher levels of endothelin-1 and significantly lower levels of NO in both venous and cavernosal blood than had those with psychogenic ED. There were significant positive correlations in both venous and cavernosal blood between endothelin-1 and angiotensin II, and between NO and PGE(2) in all patients with ED and the two subgroups. There were significant negative correlations between venous and cavernosal endothelin-1 and NO, endothelin-1 and PGE(2), angiotensin II and NO, and between angiotensin II and PGE(2). CONCLUSION: The present results suggest that endothelin-1 could be a clinical marker of diffuse endothelial disease manifested by ED. As angiotensin-converting enzyme (ACE) activity controls angiotensin II there might be a rationale for the use of ACE inhibitors to prevent or treat ED. NO and PGE(2) may provide new strategies for the pharmacological treatment of ED.     

Evaluation of tetrahydrobiopterin （BH4） as a potential therapeutic agent to treat erectile dysfunction

Aim： Nitric oxide （NO）-mediated smooth muscle relaxation causes penile erections. The endothelial NO synthase （eNOS） coenzyme tetrahydrobiopterin （BH4） converts eNOS-mediated catalytic activity from oxygen radical to NO production, improving endothelial function and vascular smooth muscle relaxation. Methods： Using quantitative immunohistochemistry, 8-isoprostane and nitrotyrosine concentrations were compared in cavernosal tissue from 17 potent and 7 impotent men, and the effect of single oral doses of BH4 on penile rigidity and tumescence was investigated. The pharmacodynamic effect of single oral doses of BH4 on penile rigidity and tumescence was investigated in a randomized, placebo-controlled, double-blind cross-over fashion in 18 patients with erectile dysfunction （ED） while receiving visual sexual stimulation. Results： 8-isoprostane content in endothelium and smooth muscle was signifi- cantly higher in impotent patient samples; the level of nitrotyrosine was unchanged in ED patients. Relative to placebo, a single dose of 200 mg BH4 led to a mean increase in duration of 〉 60% penile rigidity （33.5 rain [95% confidence interval （CI）： 13.1-49.3] at base and 29.4 rain [95% CI： 8.9-42,2] at tip）. A 500-mg dose increased the relative duration of 〉 60% penile rigidity by 36. I rain （95% CI： 16.3-51.8） at the base and 33.7 rain （95% CI： 11.4-43.9） at the tip. Treatments were well tolerated. Conclusion： BH4 treatment is suggested to switch eNOS catalytic activity from super-oxide to NO formation, leading to a reduced formation of free radical reaction product 8-isoprostane without alteration of nitrotyrosine. The observed results make BH4 a suitable candidate as an ED treatment through reconstitution of altered catalytic activity of the eNOS. （Asian JAndro12006 Mar; 8： 159-167）     

内皮型一氧化氮合酶Glu298Asp基因多态性与老年原发性高血压微量白蛋白尿的关系

目的研究内皮型一氧化氮合酶（eNOS）Glu298Asp基因多态性与老年原发性高血压（EH）微量白蛋白尿（MAU）的关系。方法从到医院就诊的老年EH患者中筛选出202例无显性蛋白尿的患者,行24h MAU测定,并应用基因芯片技术检测eNOS Glu298Asp基因多态性,按照24hMAU定量分为MAU组和非MAU组（NAU组）,比较两组基因型和等位基因分布差异。结果两组等位基因和基因型的分布不同,MAU组等位基因T及含等位基因T的基因型（GT＋TT）分布频率明显升高（χ^2=6.62,P〈0.01;χ^2=7.29,P〈0.01）;T等位基因变异使老年EH患者MAU的相对危险度显著增高（OR=2.361,95%CI=1.256～4.437）。结论 T等位基因是老年EH患者MAU的易感基因,携带T等位基因导致老年EH患者出现MAU的风险显著增高。     

Associations between endothelial nitric oxide synthase polymorphisms and risk of diabetic nephropathy: an updated meta-analysis

Abstract

Background: Genetic polymorphism of endothelial nitric oxide synthase (eNOS) has been implicated in the risk of diabetic nephropathy (DN), but the published findings were inconsistent. We performed a comprehensive meta-analysis to derive a more precise estimation of the association between genetic polymorphisms of eNOS and the risk of DN. Methods: Six online database were researched on the associations between polymorphisms of eNOS (T786C, G894T, 4b/4a) and DN risk. PRISMA statement and Hardy–Weinberg equilibrium assessment were used in this report. Odds ratio and 95% confidence interval were estimated based on the following genetic contrast/models: allelic model, dominant model, recessive model and co-dominant model. The publication bias and sensitivity analysis were also performed to guarantee the statistical power. Results: A total of 49 case–control studies with 11,990/9754/5131 participants for DN/DM/HC group were eligible for meta-analysis (7/25/31 studies for T786C/G984T/4b/a). For the eNOS-T786C, C allele showed a weak association between C allele and DN risk in DN/T2DM group. For eNOS-G894T, there was an association between T allele and DN risk in the global, Asian and African population in DN/T2DM group. For the eNOS-4b/4a, 4a allele was found contributing significantly to increased DN risk in the global population. Conclusion: Our comprehensive meta-analysis suggests that three polymorphisms of eNOS may be the increased risk factors of DN development, especially in Asian population and T2DM group.     

T-786C polymorphism of the endothelial nitric oxide synthase gene is associated with albuminuria in the diabetes heart study

Albuminuria demonstrates significant heritability in multiply affected hypertensive and diabetic families. The role of endothelial nitric oxide synthase (eNOS) gene variants as risk factors for albuminuria was investigated in 590 European American siblings from 230 families in the Diabetes Heart Study. Two polymorphisms in the eNOS gene (T-786C in the promoter region and Glu298Asp in exon 7) were genotyped. Albuminuria was defined as an albumin:creatinine ratio (ACR) >/=17 mg/g in men and >/=25 mg/g in women. Tests of association were based on generalized estimating equations, and tests of linkage disequilibrium were based on the quantitative pedigree disequilibrium test. A total of 83% of participants had type 2 diabetes. The median ACR was 10.7 mg/g (interquartile range, 5.1 to 32.8), and 34% (202 of 590) of participants had an elevated ACR. The eNOS -786C allele but not the Glu298Asp was associated with increased ACR (31% increase in absolute level of ACR for each additional copy of the -786C allele; P < 0.0001) and a higher risk for albuminuria (odds ratio, 1.55 for each additional copy of the -786C allele; P = 0.0005). Adjustment for the nongenetic determinants of ACR had no significant effect on the results; neither did stratification by gender, presence of diabetes, and the Glu298Asp genotype. Results were confirmed by quantitative pedigree disequilibrium test analysis and were consistent with haplotype analysis. The -786C eNOS variant was positively correlated with a higher prevalence and a greater degree of albuminuria in European American families in both diabetic and nondiabetic family members.     

Endothelial nitric oxide synthase gene intron 4 polymorphism in patients with end-stage renal disease.

BACKGROUND: Nitric oxide (NO) synthesized by endothelial cell NO synthase (ecNOS) is a potent regulator of intrarenal haemodynamics. A polymorphism in intron 4 of the ecNOS gene is a candidate gene in cardiovascular and renal diseases. We investigated a potential involvement of this polymorphism in chronic renal failure. METHODS: We performed a case-control study involving 706 patients with end-stage renal disease (ESRD) and 321 healthy controls. All subjects were genotyped for the ecNOS4 polymorphism by the polymerase chain reaction followed by agarose gel electrophoresis. RESULTS: The analysis revealed that the frequencies of the ecNOS4 genotypes were significantly different in ESRD patients, both diabetic and non-diabetic, than in controls. In all dialysis patients for aa, ab and bb genotypes the frequencies were, respectively, 6.5, 35 and 58.5% in the patient group, and 1, 25 and 74% in control subjects. The a allele carriers (aa + ab) were more frequent among ESRD patients than in controls (OR 1.95; 95% CI 1.13-3.4; P = 0.0031). No significant association was found when hypertensive ESRD patients were compared with normotensive patients. The distribution of genotypes was similar in both subgroups (P = 0.21). CONCLUSION: There was a significantly higher frequency of the ecNOS4a allele carriers among ESRD patients, both diabetic and non-diabetic, than in control subjects. This suggests that the ecNOS gene polymorphism may be associated with an increased risk of chronic renal failure.     

Endothelial nitric oxide synthase gene polymorphism in intron 4 affects the progression of renal failure in non-diabetic renal diseases.

BACKGROUND: Nitric oxide is a very potent regulator of intrarenal haemodynamics and is thought to be an important factor in the deterioration of renal function. Our study sought to verify the hypothesis that endothelial nitric oxide synthase (ecNOS) gene polymorphism in intron 4 might have some relevance to progression in chronic renal failure. METHODS: We studied the frequencies of gene polymorphism of ecNOS intron 4 in patients with end-stage renal disease (302 cases) and compared it with that of healthy subjects (248 cases). ecNOS genotypes were determined by the polymerase chain reaction, followed by agarose gel electrophoresis. RESULTS: Two alleles of ecNOS intron 4, labelled a and b could be detected; a has four and b has five tandem 27-bp repeats. The frequencies of ecNOS4b/b, ecNOS4b/a, ecNOS4a/a genotypes were 81.0% (201/248), 19. 0% (47/248), 0.0% (0/248) in the control group, and 74.8% (226/302), 23.5% (71/302), l.7% (5/302) in all the patients, 72.7% (168/231), 25.1% (58/231), 2.2% (5/231) in the group with end-stage renal diseases, excluding diabetic nephropathy (non-DM group), and 81.7% (58/71), 18.3% (13/71), 0.0% (0/71) in diabetic nephropathy (DM group) respectively. The frequency of the ecNOS4a (ecNOSb/a, and ecNOSa/a) in all the patients and in the non-DM group were significantly higher than that in the control group (P=0.021; P=0. 0096 respectively). In contrast, there was no significant difference in the frequencies of ecNOS genotypes between the DM group and the control group (P=0.81). CONCLUSION: Among the frequencies of ecNOS intron 4 gene polymorphism, a allele displayed a significantly higher frequency in cases with end-stage renal failure (ESRF) not caused by diabetic nephropathy. ecNOS gene polymorphism in intron 4 appears, therefore, to affect the progression of renal failure in non- diabetic renal diseases, but the same conclusion could not be drawn in diabetic nephropathy.