Intracerebral Administration of S-Adenosylhomocysteine or S-Adenosylmethionine Attenuates the Increases in the Cortical Extracellular Levels of Dimethylarginines Without Affecting cGMP Level in Rats with Acute Liver Failure.

Alterations in brain nitric oxide (NO)/cGMP synthesis contribute to the pathogenesis of hepatic encephalopathy (HE). An increased asymmetrically dimethylated derivative of L-arginine (ADMA), an endogenous inhibitor of NO synthases, was observed in plasma of HE patients and animal models. It is not clear whether changes in brain ADMA reflect its increased local synthesis therefore affecting NO/cGMP pathway, or are a consequence of its increased peripheral blood content. We measured extracellular concentration of ADMA and symmetrically dimethylated isoform (SDMA) in the prefrontal cortex of control and thioacetamide (TAA)-induced HE rats. A contribution of locally synthesized dimethylarginines (DMAs) in their extracellular level in the brain was studied after direct infusion of the inhibitor of DMAs synthesizing enzymes (PRMTs), S-adenosylhomocysteine (AdoHcy, 2 mM), or the methyl donor, S-adenosylmethionine (AdoMet, 2 mM), via a microdialysis probe. Next, we analyzed whether locally synthesized ADMA attains physiological significance by determination of extracellular cGMP. The expression of PRMT-1 was also examined. Concentration of ADMA and SDMA, detected by positive mode electrospray LC-DMS-MS/MS, was greatly enhanced in TAA rats and was decreased (by 30 %) after AdoHcy and AdoMet infusion. TAA-induced increase (by 40 %) in cGMP was unaffected after AdoHcy administration. The expression of PRMT-1 in TAA rat brain was unaltered. The results suggest that (i) the TAA-induced increase in extracellular DMAs may result from their effective synthesis in the brain, and (ii) the excess of extracellular ADMA does not translate into changes in the extracellular cGMP concentration and implicate a minor role in brain NO/cGMP pathway control.

S-Adenosylhomocysteine enhances DNA damage through increased ß-amyloid formation and inhibition of the DNA-repair enzyme OGG1b in microglial BV-2 cells.

S-Adenosylhomocysteine (SAH) is a risk factor for neurodegenerative diseases such as Alzheimer's disease, for which ß-Amyliod (Aß) formation is a major risk factor. We recently showed that SAH increases Aß formation in mouse microglial BV2 cells. Here, we show that incubation of BV2 cells with SAH (0-500nM) for 6-24h sequentially increased Aß formation, ROS and DNA damage measured as 8-oxo-deoxyguanosine (8-oxo-dG) levels. Pre-incubation of BV2 cells with 20µM ß-secretase inhibitor IV for 30min followed by incubation with SAH (500nM) markedly decreased Aß formation and 8-oxo-dG levels. Treatment with SAH for 24h concentration-dependently inhibited DNA methyltransferase (DNMT1) activity and inhibited DNMT1 binding to Sp1 site of 8-oxoG-DNA glycosylases I (OGG1) promoter and OGG1 protein and mRNA expression at 24h; the latter effect was attributed to hypomethylation of the OGG1 gene promoter, because pre-incubation of cells with betaine (1.0mM for 30 min) markedly prevented the inhibition of OGG1 protein expression induced by SAH. Overall, we demonstrate that SAH increases DNA damage in BV-2 cells possible by increased Aß formation leading to increased formation of ROS. Furthermore, the DNA damage is enhanced by SAH through inhibition of DNMT1 activity and hypomethylation of OGG1 gene promoter.

Homocysteine, a risk factor for atherosclerosis, biphasically changes the endothelial production of kynurenic acid.

Increased serum level of homocysteine is an independent risk factor for vascular disease. The effect of DL-homocysteine on the endothelial production of kynurenic acid, an antagonist of alpha7-nicotinic and N-methyl-D-aspartate (NMDA) glutamate receptors, has been evaluated in vitro and in vivo. In rat aortic rings, DL-homocysteine at 40-100 microM enhanced, whereas at >or=400 microM decreased the synthesis of kynurenic acid. S-adenosylhomocysteine mimicked the biphasic action of DL-homocysteine. On the contrary, thiol-containing compounds, L-cysteine and L-methionine, were only inhibiting kynurenic acid production. L-kynurenine uptake blockers, L-phenylalanine and L-leucine, reversed the stimulatory effect of S-adenosylhomocysteine. L-glycine, co-agonist of NMDA receptor, and cis-4-phosphonomethyl-2-piperidine carboxylic acid (CGS 19755), an antagonist of NMDA receptor, have not influenced kynurenic acid formation. In vivo, DL-homocysteine (1.3 mmol, i.p.) increased the level of kynurenic acid in rat serum from 23.7+/-7.1 to 60.7+/-14.2 (15 min, P<0.01) and 55.7+/-13.6 (60 min, P<0.01) pmol/ml, respectively; the endothelial content of kynurenic acid was also increased (51.6+/-5.8 vs. 73.2+/-9.4 fmol/microg of protein; 15 min; P<0.01). DL-homocysteine seems to modulate the production of kynurenic acid both directly and indirectly, possibly following the conversion to S-adenosylhomocysteine. The obtained data suggest a potential contribution of altered formation of kynurenic acid to the endothelial changes induced by hyperhomocysteinemia.

Striatal dopamine depletion, tremors, and hypokinesia following the intracranial injection of S-adenosylmethionine: a possible role of hypermethylation in parkinsonism.

The major symptoms of Parkinson disease (PD) are tremors, hypokinesia, rigidity, and abnormal posture, caused by the degeneration of dopamine (DA) neurons in the substantia nigra (SN) and deficiency of DA in the neostriatal DA terminals. Norepinephrine (NE) and serotonin (5-HT) levels in the neostriatum and tyrosine hydroxylase and melanin pigments in the substantia nigra are also decreased, and brain cholinergic activity is increased. The cause of PD is unknown, but PD is an age-related disorder, suggesting that changes that occur during the aging process may help to precipitate PD. Methylation increases in aging animals. Increased methylation can deplete DA, NE, and 5-HT; increase acetylcholine; and cause hypokinesia and tremors. These effects are similar to changes seen in PD, and interestingly also, they are similar to some of the changes that are associated with the aging process. It is suggested, therefore, that increased methylation may be an inducing factor in parkinsonism. Accordingly, the effects of an increase in methylation in the brain of rats were studied. S-adenosylmethionine (AdoMet), the limiting factor in the methylation process, was injected into the lateral ventricle of rats. Specific behavioral changes that resemble changes seen in PD were investigated. The results showed that AdoMet caused tremors, rigidity, hypokinesia, and depleted DA. The hypokinetic effects of a single dose of AdoMet lasted for about 90 min. AdoMet has a dose-dependent hypokinetic effect. A dose of 9.4 nmol reduced movement time (MT) by 68.9% and increased rest time (RT) by 20.7%, and a dose of 400 nmol reduced MT by 92.4% and increased RT by 27.6%. The normethyl analog of AdoMet, S-adenosylhomocysteine, did not cause hypokinesia or tremors, but it blocked the AdoMet-induced motor effects. L-dopa, the precursor of DA, also blocked the AdoMet-induced motor effects. These data suggest that the methyl group of AdoMet as well as DA depletion are involved in the AdoMet-induced motor effects. A dose of 0.65 mumol of AdoMet depleted DA in the ipsilateral caudate nucleus (CN) or neostriatum by 50.1%, and DA in the contralateral CN was reduced by 9.3%. Double the dose of AdoMet did not increase the depletion of DA on the ipsilateral CN, but DA in the contralateral CN was decreased by 26.3%. Taken together, the results suggest that increased methylation may contribute to the symptoms of PD.