Abstract
We investigated the effects of hepatic ischemia/reperfusion (I/R) injury on asymmetric dimethylarginine (ADMA, a nitric oxide synthase inhibitor), protein methyltransferase (PRMT) and dimethylarginine dimethylaminohydrolase (DDAH) (involved, resp., in ADMA synthesis and degradation), and the cationic transporter (CAT). Male Wistar rats were subjected to 30 or 60 min hepatic ischemia followed by 60 min reperfusion. ADMA levels in serum and bile were determined. Tissue ADMA, DDAH activity, DDAH-1 and CAT-2 protein, DDAH-1 and PRMT-1 mRNA expression, GSH/GSSG, ROS production, and lipid peroxidation were detected. ADMA was found in bile. I/R increased serum and bile ADMA levels while an intracellular decrease was detected after 60 min ischemia. Decreased DDAH activity, mRNA, and protein expression were observed at the end of reperfusion. No significant difference was observed in GSH/GSSG, ROS, lipid peroxidation, and CAT-2; a decrease in PRMT-1 mRNA expression was found after I/R. Liver is responsible for the biliary excretion of ADMA, as documented here for the first time, and I/R injury is associated with an oxidative stress-independent alteration in DDAH activity. These data are a step forward in the understanding of the pathways that regulate serum, tissue, and biliary levels of ADMA in which DDAH enzyme plays a crucial role.
Highlights
Nitric oxide (NO) is abundantly synthesized from the amino acid arginine, by the action of NO-synthase (NOS), a family of enzymes with an endothelial, neuronal, and inducible isoform [1]
To confirm biliary Asymmetric dimethylarginine (ADMA) levels, bile samples were analyzed by the HPLC method equipped with a fluorescence detector and no significant differences when comparing data obtained by the ELISA kit were detected (i.e., 0.31 ± 0.04 versus 0.29 ± 0.02 nmol/mL, resp., in bile obtained after 30 min ischemia followed 60 min reperfusion)
A significant oxidative stress-independent decrease in dimethylarginine dimethylaminohydrolase (DDAH) activity that associates with low mRNA and protein expression occurs at the end of the reperfusion period supporting the hypothesis that the ADMA/DDAH pathway plays a crucial role in acute hepatic I/R injury
Summary
Nitric oxide (NO) is abundantly synthesized from the amino acid arginine, by the action of NO-synthase (NOS), a family of enzymes with an endothelial, neuronal, and inducible isoform [1]. Asymmetric dimethylarginine (ADMA) is an endogenous inhibitor of these enzymes because it competes with L-arginine for each of the three isoforms of NOS: it is considered an important marker of endothelial dysfunction because of its inhibiting role in NO synthesis. An additional pathway was found for ADMA, namely, metabolic degradation by dimethylarginine dimethylaminohydrolase (DDAH), an enzyme that is widely distributed in rats and human subjects, but, in particular, in the liver, kidney, and pancreas [5, 6]. The molecular mechanisms involved in I/R injury are not completely understood and only a few works have reported changes induced by hepatic I/R injury on the ADMA/DDAH pathway which needs to be considered as a point of interest potentially capable of reducing the effects of I/R. Enzymatic activity and protein expression of DDAH and mRNA expression of DDAH-1 and PRMT-1 and CAT-2 protein at the end of reperfusion were examined
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