Lipotoxicity induced by saturated fatty acids (SFAs) plays a central role in the pathogenesis of non-alcoholic fatty liver disease (NAFLD); however, the exact mechanism(s) remain to be fully elucidated. SIRT3 is an NAD+-dependent deacetylase primarily located inside mitochondria. In this study, we demonstrated that a SFAs-rich high-fat diet (HFD) was more detrimental to the liver than an isocaloric unsaturated FAs-rich HFD. Unexpectedly, SIRT3 expression/activity were significantly elevated in the livers of mice exposed to the SFAs-rich HFD. Using cultured HepG2 and AML-12 hepatocytes, we demonstrated that unlike monounsaturated FAs, SFAs upregulates SIRT3 expression/activity. SIRT3 overexpression renders both the liver and hepatocytes susceptible to palmitate-induced cell death, which can be alleviated by SIRT3 siRNA transfection. In contrast, SIRT3 suppression protects hepatocytes from palmitate cytotoxicity. Further studies revealed that SIRT3 acts as a negative regulator of autophagy, whereby enhancing the susceptibility of hepatocytes to SFAs-induced cytotoxicity. Mechanistic investigations elucidate that SIRT3 overexpression causes manganese superoxide dismutase (MnSOD) deacetylation/activation, which depleted intracellular superoxide contents, leading to AMP-activated protein kinase (AMPK) inhibition and mTORC1 activation, resulting in autophagy suppression. In contrast, SIRT3 siRNA gene silencing enhances autophagy flux. The similar result was observed in the liver tissue from SIRT3 knockout mice. Conclusion our data identified SIRT3 to be a novel negative regulator of autophagy, whose activation by SFAs contributes to lipotoxicity in hepatocytes and suggest that restraining SIRT3 overactivation can be a potential therapeutic choice for the treatment of NAFLD as well as other metabolic disorders, with lipotoxicity being the principal pathomechanism.

Background & Aim-Abundantly expressed in the metabolically active cells including hepatocytes, N-nicotinamide methyltransferase (NNMT) catalyzes S-adenosylmethionine (SAM)dependent methylation/degradation of nicotinamide, the predominant precursor for intracellular nicotinamide adenine dinucleotide (NAD + ) regeneration via the salvage pathway. The enzyme is emerging to play an important role in regulating metabolism, however, the exact regulatory mechanism(s) underlying NNMT expression remains unclear and its potential implication in alcoholic liver disease (ALD) is completely unknown.Methods-Both traditional Lieber-De Carli and the NIAAA mouse model of ALD were employed. A small scale of chemical screening and ChIP assay were performed. NNMT inhibition was achieved via both genetic (adenoviral shRNA delivery) and pharmacological approach.Results-Chronic alcohol consumption induces liver endoplasmic reticulum (ER) stress and upregulates hepatic NNMT expression. ER stress inducers upregulated NNMT expression in both AML-12 hepatocytes and mice. PERK-ATF4 pathway activation is the main contributor in ER

Background: Salvianolic acid A (Sal A), a natural polyphenol compound extracted from Radix Salvia miltiorrhiza (known as Danshen in China), possesses a variety of potential pharmacological activities. The aim of this study is to determine mechanisms of hepatoprotective effects of Sal A against lipotoxicity both in cultured hepatocytes and in a mouse model of fatty liver disease.Methods: High-fat and high-carbohydrate diet (HFCD)-fed C57BL/6J mice were employed to establish hepatic lipotoxicity in a mouse model. Two doses of Sal A were administered every other day via intraperitoneal injection (20 and 40 mg/kg BW, respectively). After a 10-week intervention, liver injury was detected by immunohistochemical and biochemical analyses. For in vitro studies, we used HepG2, a human hepatoma cell line, and exposed them to palmitic acid to induce lipotoxicity. The protective effects of Sal A on palmitic acid-induced lipotoxicity were examined in Sal A-pretreated HepG2 cells.Results: Sal A treatments attenuated body weight gain, liver injury, and hepatic steatosis in mice exposed to HFCD. Sal A pretreatments ameliorated palmitic acid-induced cell death but did not reverse effects of HFCD- or palmitate-induced activations of JNK, ERK1/2, and PKA. Induction of p38 phosphorylation was significantly reversed by Sal A in HFCD-fed mice but not in palmitate-treated HepG2 cells. However, Sal A rescued hepatic AMP-activated protein kinase (AMPK) suppression and sirtuin 1 (SIRT1) downregulation by both HFCD feeding in mice and exposure to palmitate in HepG2 cells. Sal A dose-dependently up-regulated p-AMPK and SIRT1 protein levels. Importantly, siRNA silencing of either AMPK or SIRT1 gene expression abolished the protective effects of Sal A on lipotoxicity. Moreover, while AMPK silencing blocked Sal A-induced SIRT1, silencing of SIRT1 had no effect on Sal A-triggered AMPK activation, suggesting SIRT1 upregulation by Sal A is mediated by AMPK activation.Conclusion: Our data uncover a novel mechanism for hepatoprotective effects of Sal A against lipotoxicity both in livers from HFCD-fed mice and palmitic acid-treated hepatocytes.

N-acetylcysteine (NAC), a well-accepted antioxidant, has been shown to protect against high fat diet (HFD)-induced obesity-associated non-alcoholic fatty liver disease (NAFLD) in mice. However, the underlying mechanism(s) of the beneficial...

Background Lipotoxicity-induced cell death plays a detrimental role in the pathogenesis of metabolic diseases. Ferulic acid, widespread in plant-based food, is a radical scavenger with multiple bioactivities. However, the benefits of ferulic acid against hepatic lipotoxicity are largely unclear. Here, we investigated the protective effect of ferulic acid against palmitate-induced lipotoxicity and clarified its potential mechanisms in AML-12 hepatocytes. Methods AML-12 mouse hepatocytes were exposed to palmitate to mimic lipotoxicity. Different doses (25, 50, and 100 μM) of ferulic acid were added 2 h before palmitate treatment. Cell viability was detected by measuring lactate dehydrogenase release, nuclear staining, and the expression of cleaved-caspase-3. Intracellular reactive oxygen species content and mitochondrial membrane potential were analysed by fluorescent probes. The potential mechanisms were explored by molecular biological methods, including Western blotting and quantitative real-time PCR, and were further verified by siRNA interference. Results Our data showed that ferulic acid significantly inhibited palmitate-induced cell death, rescued mitochondrial membrane potential, reduced reactive oxygen species accumulation, and decreased inflammatory factor activation, including IL-6 and IL-1beta. Ferulic acid significantly stimulated autophagy in hepatocytes, whereas autophagy suppression blocked the protective effect of ferulic acid against lipotoxicity. Ferulic acid-activated autophagy, which was triggered by SIRT1 upregulation, was mechanistically involved in its anti-lipotoxicity effects. SIRT1 silencing blocked most beneficial changes induced by ferulic acid. Conclusions We demonstrated that the phytochemical ferulic acid, which is found in plant-based food, protected against hepatic lipotoxicity, through the SIRT1/autophagy pathway. Increased intake of ferulic acid-enriched food is a potential strategy to prevent and/or improve metabolic diseases with lipotoxicity as a typical pathological feature.