Effects of asymmetric dimethylarginine on renal arteries in portal hypertension and cirrhosis

AIM To evaluate the effects of asymmetric dimethylarginine(ADMA) in renal arteries from portal hypertensive and cirrhotic rats.METHODS Rat renal arteries from Sham(n = 15), pre-hepatic portal hypertension(PPVL; n = 15) and bile duct ligation and excision-induced cirrhosis(BDL; n = 15) were precontracted with norepinephrine, and additional contractions were induced with ADMA(10-6-10-3 mol/L), an endogenous inhibitor of nitric oxide(NO) synthase. Concentration-response curves to acetylcholine(1 × 10-9^(-3) × 10^(-6) mol/L) were determined in precontractedrenal artery segments with norepinephrine in the absence and in the presence of ADMA. Kidneys were collected to determine the protein expression and activity of dimethylarginine dimethylaminohydrolase(DDAH), an enzyme that catabolizes ADMA. RESULTS In renal arteries precontracted with norepinephrine, ADMA caused endothelium-dependent contractions. The pD 2 values to ADMA were similar in the Sham and PPVL groups(4.20 ± 0.08 and 4.11 ± 0.09, P > 0.05, respectively), but were lower than those of the BDL group(4.79 ± 0.16, P < 0.05). Acetylcholine-induced endothelium-dependent relaxation that did not differ, in terms of p D2 and maximal relaxation, among the 3 groups studied. Treatment with ADMA(3 × 10^(-4) mol/L) inhibited acetylcholine-induced relaxation in the 3 groups, but the inhibition was higher(P < 0.05) in the BDL group compared with that for the Sham and PPVL groups. The m RNA and protein expression of DDAH-1 were similar in kidneys from the three groups. Conversely, DDAH-2 expression was increased(P < 0.05) in PPVL and further enhanced(P < 0.05) in the BDL group. However, renal DDAH activity was significantly decreased in the BDL group. CONCLUSION Cirrhosis increased the inhibitory effect of ADMA on basal- and induced-release of NO in renal arteries, and decreased DDAH activity in the kidney.

Asymmetric dimethylarginine as a mediator of vascular dysfunction in cirrhosis

Cirrhosis is associated with marked abnormalities in the circulatory function that involve a reduction in systemic vascular resistance. An important cause of this vasodilatation is the increased production or activity of nitric oxide(NO) in the splanchnic circulation. During portal hypertension and cirrhosis an increased endothelial NO synthase(e NOS) activity is demonstrated in splanchnic vessels. In contrast, the activity of e NOS in the cirrhotic liver is decreased, which suggests a different regulation of e NOS in the liver and in the splanchnic vessels. Asymmetric dimethylarginine(ADMA) is an endogenous NO inhibitor and higher plasma levels of ADMA are related to increased cardiovascular risk in both the general population and among patients with cirrhosis. It has been demonstrated that the liver is a key player in the metabolism of ADMA. This observation was further supported by investigations in human patients, showing a close correlation between ADMA plasma levels and the degree of hepatic dysfunction. ADMA is degraded to citrulline and dimethylamine by dimethylarginine dimethylaminohydrolases(DDAHs). DDAHs are expressed as type 1 and 2 isoforms and are widely distributed in various organs and tissues, including the liver. In this review, we discuss experimental and clinical data that document the effects of dimethylarginines on vascular function in cirrhosis. Our increasing understanding of the routes of synthesis and metabolism of methylarginines is beginning to provide insights into novel mechanisms of liver disease and allowing us to identify potential therapeutic opportunities.

DDAH2(-449 G/C) G allele is positively associated with leukoaraiosis in northeastern China: a double-blind, intergroup comparison, case-control study

Cerebrovascular endothelial dysfunction is involved in the progression of leukoaraiosis. Asymmetric dimethylarginine is a competitive inhibitor of nitric oxide, which is highly expressed in patients with leukoaraiosis. Dimethylarginine dimethylaminohydrolase(DDAH) is a hydrolytic enzyme that is primarily responsible for eliminating asymmetric dimethylarginine, and it plays a role in the pathogenesis of cardiovascular and cerebrovascular diseases. The DDAH2 subtype is expressed in organs rich in induced nitric oxide synthase, including the heart, the placenta, and the cerebral endothelium during cerebral ischemia, in the stress state, or under neurotoxicity. Overexpression of the DDAH2 gene can inhibit asymmetric dimethylarginine-induced peripheral circulating endothelial cell dysfunction. However, it is unknown whether this polymorphism regulates plasma asymmetric dimethylarginine levels in patients with leukoaraiosis. In this doubleblind study, we recruited 46 patients with leukoaraiosis and 46 healthy, matched controls. Plasma asymmetric dimethylarginine levels were determined using enzyme-linked immunoassays. Genomic DNA was isolated from whole blood samples, and polymerase chain reaction, Sma I restriction enzyme digestion, restriction fragment length polymorphisms, and agarose electrophoresis were used to detect DDAH2(-449 G/C) gene polymorphisms. The results revealed that 95.65% of leukoaraiosis patients had recessive genetic models(GG and CG), while 89.13% of healthy control subjects had dominant genetic models(CC and CG). There was a significant difference in the genotype composition ratio between leukoaraiosis patients and healthy controls(P = 0.0002). The frequency of G alleles in the leukoaraiosis patients(71.74%) was significantly higher than in healthy controls, whereas the frequency of C alleles was lower(χ~2 = 13.9580, P = 0.0002). Furthermore, asymmetric dimethylarginine concentrations in subjects with the GG genotype were significantly higher than in subjects with the CG and CC genotypes(Kruskal–Wallis H = 24.5955, P < 0.0001). In addition, the GG genotype of DDAH2(-449 G/C) was more common in patients with leukoaraiosis. These findings suggest that the G allele of DDAH2(-449 G/C) is a risk factor for leukoaraiosis morbidity and is correlated with high levels of asymmetric dimethylarginine. This study was approved by the Institutional Ethics Committee of the 2~(nd) Affiliated Hospital of Harbin Medical University of China(approval No. KY2016-177) on July 28, 2016.

Dysfunction of endothelial NO system originated from homocysteine-induced aberrant methylation pattern in promoter region of DDAH2 gene

Background Hyperhomocysteinemia （HHcy）-mediated dysfunction of endothelial NO system is an important mechanism for atherosclerotic pathogenesis. Dimethylarginine dimethylaminohydrolase （DDAH） is the key enzyme for degrading asymmetric dimethylarginine （ADMA）, which is an endogenous inhibitor of endothelial nitric oxide （NO） synthase （eNOS）. This study was designed to investigate whether the dysfunction of endothelial NO system originates from HHcy-mediated aberrant methylation modification in promotor region of DDAH2 gene. Methods Human umbilical vein endothelial cells （HUVECs） were cultured to the third generation and treated with homocysteine （Hcy） at different concentrations （0, 10, 30, 100, and 300 μmol/L） for 72 hours. The methylation pattern in promoter region CpG island of DDAH2 gene was analyzed by nested methylation-specific PCR （nMSP）. The mRNA expression of eNOS gene and DDAH2 gene was detected by semi-quantitative RT-PCR. The activity of DDAH2 and eNOS in cells, and the concentrations of ADMA and NO in culture medium were assayed respectively. Results Mild increased concentration of Hcy （10 and 30 μmol/L） induced hypomethylation, while high concentration of Hcy （100 and 300 μmol/L） induced hypermethylation in the promoter CpG island of DDAH2 gene. The mRNA expression of DDAH2 increased in mild enhanced concentration of Hcy, and decreased in high concentration of Hcy correspondingly. The inhibition of DDAH2 activity, the increase of ADMA concentration, the reduction of eNOS activity and the decrease of NO production were all consistently relevant to the alteration of Hcy concentration. Conclusion The increased concentration of Hcy induced aberrant methylation pattern in promotor region of DDAH2 gene and the successive alterations in DDAH/ADMA/NOS/NO pathway, which showed highly relevant and dose-effect relationship. The results suggested that the dysfunction of endothelial NO system induced by HHcy could be partially originated from Hcy-mediated aberrant methylation in DDAH2 gene.

Ep,gallocatechin-3-gallate ameliorates erectile function in aged rats via regulation of PRMT1/ DDAH/ADMA/NOS metabolism pathway

Aging-related ED is predominantly attributed to neurovascular dysfunction mediated by NO suppression and increased oxidative stress in penis. The alterations of protein arginine methyltransferases 1 （PRMT1）/dimethylarginine dimethylaminohydrolase （DDAH）/ asymmetrical dimethylarginine （ADMA）/NO synthase （NOS） pathway regulate NO production in the vascular endothelium. Epigallocatechin-3-gallate （EGCG） is one of the most abundant and antioxidative ingredients isolated from green tea. In the present study, 40 Sprague-Dawley rats were randomly distributed into four groups： one young rat group and three aged rat groups treated with daily gavage feedings of EGCG at doses of O, 10 mg kg-1, and 100 mg kg-1 for 12 weeks, respectively. Erectile function was assessed by electrical stimulation of the cavernous nerves with intracavernous pressure （ICP） measurement. After euthanasia, penile tissue was investigated using Western blot and ELISA to assess the PRMTI/DDAH/ADMA/NOS metabolism pathway. Superoxide dismutase （SOD） and malondialdehyde （MDA） levels were detected by colorimetry. We also evaluated smooth muscle contents. The ratio of maximal ICP and mean systemic arterial pressure （MAP） was markedly higher in EGCG-treated aged rats than in untreated aged rats. We found that DDAH 1 and DDAH2 were expressed in cavernosal tissue, and they were downregulated in corpora of aged rats. The administration of EGCG upregulated the expression and activity of DDAH. In contrast, EGCG treatment downregulated the expression of PRMT1 and ADMA content. Moreover, EGCG-treated rats showed an improvement in smooth muscle expression, the ratio of smooth muscle cell/collagen fibril, SOD activity, and MDA levels when compared with untreated aged rats.