Folate deficiency disturbs hepatic methionine metabolism and promotes liver injury in the ethanol-fed micropig

Abstract

Alcoholic liver disease is associated with abnormal hepatic methionine metabolism and folate deficiency. Because folate is integral to the methionine cycle, its deficiency could promote alcoholic liver disease by enhancing ethanol-induced perturbations of hepatic methionine metabolism and DNA damage. We grouped 24 juvenile micropigs to receive folate-sufficient (FS) or folate-depleted (FD) diets or the same diets containing 40% of energy as ethanol (FSE and FDE) for 14 wk, and the significance of differences among the groups was determined by ANOVA. Plasma homocysteine levels were increased in all experimental groups from 6 wk onward and were greatest in FDE. Ethanol feeding reduced liver methionine synthase activity, S-adenosylmethionine (SAM), and glutathione, and elevated plasma malondialdehyde (MDA) and alanine transaminase. Folate deficiency decreased liver folate levels and increased global DNA hypomethylation. Ethanol feeding and folate deficiency acted together to decrease the liver SAM͞S-adenosylhomocysteine (SAH) ratio and to increase liver SAH, DNA strand breaks, urinary 8-oxo-2 -deoxyguanosine [oxo(8)dG]͞mg of creatinine, plasma homocysteine, and aspartate transaminase by more than 8-fold. Liver SAM correlated positively with glutathione, which correlated negatively with plasma MDA and urinary oxo(8)dG. Liver SAM͞SAH correlated negatively with DNA strand breaks, which correlated with urinary oxo(8)dG. Livers from ethanol-fed animals showed increased centrilobular CYP2E1 and protein adducts with acetaldehyde and MDA. Steatohepatitis occurred in five of six pigs in FDE but not in the other groups. In summary, folate deficiency enhances perturbations in hepatic methionine metabolism and DNA damage while promoting alcoholic liver injury. F olate deficiency is among the most common nutritional abnormalities in chronic alcoholic patients, especially in those who have developed alcoholic liver injury (1-5). In addition to poor diet, folate deficiency in chronic alcoholism can be ascribed to decreased intestinal absorption and hepatic uptake, increased renal excretion, and increased oxidative cleavage of the folate molecule (6-12). Folate in its 5-methyltetrahydrofolate (5-MTHF) form is integral to methionine metabolism. Folate deficiency perturbs hepatic methionine metabolism (13,14), which is associated with DNA nucleotide imbalance and increased hepatocellular apoptosis in experimental animals fed folate-deficient (FD) diets or exposed to chronic ethanol (15,16).Hepatic methionine metabolism is regulated by the availability of dietary and endogenous folate that appears in the circulation as 5-MTHF and is the substrate with cofactor vitamin B 12 for the methionine synthase (MS) reaction that generates methionine from homocysteine (Hcy) (see supporting information, which is published on the PNAS web site, www.pnas.org). In the alternate salvage pathway for methionine synthesis, choline is the precursor of betaine, which is the substrate for betaine homocysteine methyltransferase (BHMT). The methionine adeno...

Alcohol-induced S-adenosylhomocysteine accumulation in the liver sensitizes to TNF hepatotoxicity: Possible involvement of mitochondrial S-adenosylmethionine transport

“…Abnormal hepatic methionine metabolism is an acquired metabolic abnormality in ALD. Previous studies, including ours, reported that chronic alcohol in experimental animals not only caused SAM deficiency, but also elevation of SAH [3,12,13]. These observations are consistent with the study by Lieber et al showing that alcohol consumption decreased hepatic phosphatidylethanolamine N-methyltransferase activity in a baboon model of alcohol-induced hepatic fibrosis [14].…”

“…Decreased hepatic SAM and increased homocysteine levels in ALD have been extensively reported. In addition to its effects on SAM and homocysteine levels, chronic alcohol consumption results in a significant increase in hepatic SAH levels [3,12,13]. Importantly, we showed that accumulation of intracellular SAH sensitized hepatocytes to TNF induced cytotoxicity in vitro, which we postulate is a clinically relevant metabolic alteration in the pathogenesis of ALD [12].…”

Hepatic transmethylation reactions in micropigs with alcoholic liver disease

“…As previously described, 15 24 juvenile Yucatan micropigs were obtained from Sinclair Farms (Columbia, MO) and were pair-fed diets that provided 90 kcal/kg body weight with 15% of kcal as vitamin-free casein, 30% of kcal as corn oil, and 55% of kcal as carbohydrate (control) or diets in which carbohydrate was reduced to 15% and ethanol was substituted to provide 40% of kcal. Folic acid was absent or was added to diets at 14.5 g/kg body Arrows indicate the directions of change in methionine metabolites in response to the diets as previously described.…”

“…[3][4][5][6][7]16 We developed a micropig model for alcoholic liver disease based on 14 weeks of feeding diets that either contained ethanol or were deficient in folate, or both factors in combination. 15 Compared with the control group fed a folate-sufficient diet without ethanol, each folate-deficient diet reduced liver folate by one half, and the combined diet resulted in a fourfold elevation in plasma homocysteine. Ethanol feeding alone increased liver homocysteine levels by one half and reduced liver SAM by one third and the GSH/ oxidized glutathione (GSSG) ratio by one half.…”

“…3 Recently, we explored the hypothesis that the combination of folate deficiency with chronic ethanol feeding would accelerate the development of alcoholic liver injury by enhancing abnormal methionine metabolism. 15 This hypothesis is based on the integral role of folate in methionine metabolism ( Fig. 1) and prior evidence for the effects of ethanol on promoting folate deficiency and altering methionine metabolism.…”