Activation of SIRT1 by L-serine increases fatty acid oxidation and reverses insulin resistance in C2C12 myotubes.

Silent information regulator 1 (SIRT1) is a nicotinamide adenine dinucleotide (NAD+)-dependent deacetylase, and the function is linked to cellular metabolism including mitochondrial biogenesis. Hepatic L-serine concentration is decreased significantly in fatty liver disease. We reported that the supplementation of the amino acid ameliorated the alcoholic fatty liver by enhancing L-serine-dependent homocysteine metabolism. In this study, we hypothesized that the metabolic production of NAD+ from L-serine and thus activation of SIRT1 contribute to the action of L-serine. To this end, we evaluated the effects of L-serine on SIRT1 activity and mitochondria biogenesis in C2C12 myotubes. L-Serine increased intracellular NAD+ content and led to the activation of SIRT1 as determined by p53 luciferase assay and western blot analysis of peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC-1α) acetylation. L-Serine treatment increased the expression of the genes associated with mitochondrial biogenesis and enhanced mitochondrial mass and function. In addition, L-serine reversed cellular insulin resistance determined by insulin-induced phosphorylation of Akt and GLUT4 expression and membrane translocation. L-Serine-induced mitochondrial gene expression, fatty acid oxidation, and insulin sensitization were mediated by enhanced SIRT1 activity, which was verified by selective SIRT1 inhibitor (Ex-527) and siRNA directed to SIRT1. L-Serine effect on cellular NAD+ level is dependent on the L-serine metabolism to pyruvate that is subsequently converted to lactate by lactate dehydrogenase. In summary, these data suggest that L-serine increases cellular NAD+ level and thus SIRT1 activity in C2C12 myotubes.

Correction to: Activation of SIRT1 by L-serine increases fatty acid oxidation and reverses insulin resistance in C2C12 myotubes.

AbstractThe original version of this article unfortunately contained a mistake in the article title.

Cinnamamides, Novel Liver X Receptor Antagonists that Inhibit Ligand-Induced Lipogenesis and Fatty Liver.

Liver X receptor (LXR) is a member of the nuclear receptor superfamily, and it regulates various biologic processes, including de novo lipogenesis, cholesterol metabolism, and inflammation. Selective inhibition of LXR may aid the treatment of nonalcoholic fatty liver diseases. In the present study, we evaluated the effects of three cinnamamide derivatives on ligand-induced LXRα activation and explored whether these derivatives could attenuate steatosis in mice. N-(4-trifluoromethylphenyl) 3,4-dimethoxycinnamamide (TFCA) decreased the luciferase activity in LXRE-tk-Luc-transfected cells and also suppressed ligand-induced lipid accumulation and expression of the lipogenic genes in murine hepatocytes. Furthermore, it significantly attenuated hepatic neutral lipid accumulation in a ligand-induced fatty liver mouse system. Modeling study indicated that TFCA inhibited activation of the LXRα ligand-binding domain by hydrogen bonding to Arg305 in the H5 region of that domain. It regulated the transcriptional control exerted by LXRα by influencing coregulator exchange; this process involves dissociation of the thyroid hormone receptor-associated proteins (TRAP)/DRIP coactivator and recruitment of the nuclear receptor corepressor. These results show that TFCA has the potential to attenuate ligand-induced lipogenesis and fatty liver by selectively inhibiting LXRα in the liver.

L-serine supplementation attenuates alcoholic fatty liver by enhancing homocysteine metabolism in mice and rats.

BACKGROUND: Hyperhomocysteinemia plays an important role in the development of hepatic steatosis, and studies indicate that homocysteine-lowering treatment inhibits the development of fatty liver. OBJECTIVE: We evaluated the effects of L-serine on alcoholic fatty liver and homocysteine metabolism. METHODS: In a binge ethanol study, male C57BL/6 mice were divided into 4 groups: control, ethanol + vehicle, and ethanol + 20 or 200 mg/kg L-serine. Mice were gavaged with ethanol (5 g/kg body weight) 3 times every 12 h with or without L-serine which was given twice 30 min before the last 2 ethanol doses. Control mice were fed isocaloric dextran-maltose. In a chronic ethanol study, male Wistar rats were divided into 3 groups: control, ethanol, and ethanol + L-serine. Rats were fed a standard Lieber-DeCarli ethanol diet (36% ethanol-derived calories) for 4 wk with or without dietary L-serine supplementation (1%; wt:vol) for the last 2 wk. In control rats, the ethanol-derived calories were replaced with dextran-maltose. The effects of L-serine were also tested in AML12 cells manipulated to have high homocysteine concentrations by silencing the genes involved in homocysteine metabolism. RESULTS: Binge ethanol treatment increased serum homocysteine and hepatic triglyceride (TG) concentrations by >5-fold vs. controls, which were attenuated in the 200-mg/kg L-serine treatment group by 60.0% and 47.5%, respectively, compared with the ethanol group. In the chronic ethanol study, L-serine also decreased hepatic neutral lipid accumulation by 63.3% compared with the ethanol group. L-serine increased glutathione and S-adenosylmethionine by 94.0% and 30.6%, respectively, compared with the ethanol group. Silencing betaine homocysteine methyltransferase, cystathionine ß-synthase, or methionine increased intracellular homocysteine and TG concentrations by >2-fold, which was reversed by L-serine when L-serine-independent betaine homocysteine methyltransferase was knocked down. CONCLUSION: These results demonstrate that L-serine ameliorates alcoholic fatty liver by accelerating L-serine-dependent homocysteine metabolism.

Expression of CYP3A in chronic ethanol-fed mice is mediated by endogenous pregnane X receptor ligands formed by enhanced cholesterol metabolism.

Pregnane X receptor (PXR) is a nuclear receptor that plays a key regulatory role in xenobiotic metabolism in a ligand-dependent manner. Recently, ethanol was reported to be either an inducer or inhibitor of Cytochrome P450 (CYP) 3A expression. According to our recent microarray data, chronic ethanol upregulates the expression of the genes associated with oxidative phase I drug metabolism, phase II conjugation reaction and phase III xenobiotic transport, most of which are known to be regulated by PXR. In this study, we investigated the effects of chronic ethanol on the expression and activity of CYP3A11 in mice and the role of PXR. Ethanol was administrated to male ICR mice by feeding a standard Lieber-DeCarli diet containing 36 % ethanol for 4 weeks. Ethanol significantly increased hepatic mRNA expression of Pxr and Cyp3a11. Treatment of mice with ethanol increased nuclear translocation of PXR. Consistent with the increase in nuclear PXR, ethanol significantly increased the binding of PXR to the Cyp3a11 promoter. Hepatic cholesterol level and bile acid synthesis are increased by ethanol treatment. The level of some cholesterol metabolites, such as 5ß-cholestane-3α,7α,12α-triol, 7α-hydroxy-4-cholestene-3-one and lithocholic acid, that have been identified as potent PXR agonists are increased in the livers of ethanol-treated mice. In summary, chronic ethanol upregulates the expression of Pxr and Cyp3a11 mRNAs and proteins in mice by PXR activation mediated by enhanced cholesterol metabolism and bile acid synthesis. Our data provide some critical information needed to understand the molecular mechanisms of ethanol-induced CYP3A expression.

Downregulation of PHGDH expression and hepatic serine level contribute to the development of fatty liver disease.

OBJECTIVE: Supplementation with serine attenuates alcoholic fatty liver by regulating homocysteine metabolism and lipogenesis. However, little is known about serine metabolism in fatty liver disease (FLD). We aimed to investigate the changes in serine biosynthetic pathways in humans and animal models of fatty liver and their contribution to the development of FLD. METHODS: High-fat diet (HFD)-induced steatosis and methionine-choline-deficient diet-induced steatohepatitis animal models were employed. Human serum samples were obtained from patients with FLD whose proton density fat fraction was estimated by magnetic resonance imaging. 3-Phosphoglycerate dehydrogenase (Phgdh)-knockout mouse embryonic fibroblasts (MEF) and transgenic mice overexpressing Phgdh (Tg-phgdh) were used to evaluate the role of serine metabolism in the development of FLD. RESULTS: Expression of Phgdh was markedly reduced in the animal models. There were significant negative correlations of the serum serine with the liver fat fraction, serum alanine transaminase, and triglyceride levels among patients with FLD. Increased lipid accumulation and reduced NAD+ and SIRT1 activity were observed in Phgdh-knockout MEF and primary hepatocytes incubated with free fatty acids; these effects were reversed by overexpression of Phgdh. Tg-Phgdh mice showed significantly reduced hepatic triglyceride accumulation compared with wild-type littermates fed a HFD, which was accompanied by increased SIRT1 activity and reduced expression of lipogenic genes and proteins. CONCLUSIONS: Human and experimental data suggest that reduced Phgdh expression and serine levels are closely associated with the development of FLD.

Inhibition of homocysteine-induced endoplasmic reticulum stress and endothelial cell damage by l-serine and glycine.

Hyperhomocysteinemia is an independent risk factor for several cardiovascular diseases. The use of vitamins to modulate homocysteine metabolism substantially lowers the risk by reducing plasma homocysteine levels. In this study, we evaluated the effects of l-serine and related amino acids on homocysteine-induced endoplasmic reticulum (ER) stress and endothelial cell damage using EA.hy926 human endothelial cells. Homocysteine treatment decreased cell viability and increased apoptosis, which were reversed by cotreatment with l-serine. l-Serine inhibited homocysteine-induced ER stress as verified by decreased glucose-regulated protein 78kDa (GRP78) and C/EBP homologous protein (CHOP) expression as well as X-box binding protein 1 (xbp1) mRNA splicing. The effects of l-serine on homocysteine-induced ER stress are not attributed to intracellular homocysteine metabolism, but instead to decreased homocysteine uptake. Glycine exerted effects on homocysteine-induced ER stress, apoptosis, and cell viability that were comparable to those of l-serine. Although glycine did not affect homocysteine uptake or export, coincubation of homocysteine with glycine for 24h reduced the intracellular concentration of homocysteine. Taken together, l-serine and glycine cause homocysteine-induced endothelial cell damage by reducing the level of intracellular homocysteine. l-Serine acts by competitively inhibiting homocysteine uptake in the cells. However, the mechanism(s) by which glycine lowers homocysteine levels are unclear.

LXR-α antagonist meso-dihydroguaiaretic acid attenuates high-fat diet-induced nonalcoholic fatty liver.

Collaborative regulation of liver X receptor (LXR) and sterol regulatory element binding protein (SREBP)-1 are main determinants in hepatic steatosis, as shown in both animal models and human patients. Recent studies indicate that selective intervention of overly functional LXRα in the liver shows promise in treatment of fatty liver disease. In the present study, we evaluated the effects of meso-dihydroguaiaretic acid (MDGA) on LXRα activation and its ability to attenuate fatty liver in mice. MDGA inhibited activation of the LXRα ligand-binding domain by competitively binding to the pocket for agonist T0901317 and decreased the luciferase activity in LXRE-tk-Luc-transfected cells. MDGA significantly attenuated hepatic neutral lipid accumulation in T0901317- and high fat diet (HFD)-induced fatty liver. The effect of MDGA was so potent that treatment with 1mg/kg for 2 weeks completely reversed the lipid accumulation induced by HFD feeding. MDGA reduced the expression of LXRα co-activator protein RIP140 and LXRα target gene products associated with lipogenesis in HFD-fed mice. These results demonstrate that MDGA has the potential to attenuate nonalcoholic steatosis mediated by selective inhibition of LXRα in the liver in mice.

α-Terpineol induces fatty liver in mice mediated by the AMP-activated kinase and sterol response element binding protein pathway.

The use of herbal medicines in disease prevention and treatment is growing rapidly worldwide, without careful consideration of safety issues. α-Terpineol is a monoterpene alcoholic component of Melaleuca alternifolia, Salvia officinalis and Carthamus tinctorius that is used widely as a flavor and essential oil in food. The present study showed that α-terpineol induces fatty liver via the AMP-activated protein kinase (AMPK)-mTOR-sterol regulatory element-binding protein-1 (SREBP-1) pathway. α-Terpineol-treated hepatocytes had significantly increased neutral lipid accumulation. α-Terpineol suppressed AMPK phosphorylation, and increased p70S6 kinase (p70S6K) phosphorylation and SREBP-1 activation. It also increased luciferase activity in cells transfected with LXRE-tk-Luc and SRE-tk-Luc. Inhibition of mTOR signaling by co-treatment with rapamycin or co-transfection with dominant negative p70S6K blocked completely the effects of α-terpineol. α-Terpineol oral administration to mice for 2weeks led to decreased AMPK phosphorylation and increased SREBP-1 activation in the liver, followed by hepatic lipid accumulation. Conversely, rapamycin co-treatment reversed α-terpineol-induced SREBP-1 activation and fatty liver in mice. These data provide evidence that α-terpineol causes fatty liver, an effect mediated by the AMPK/mTOR/SREBP-1 pathway.