Attenuated progression of diet-induced steatohepatitis in glutathione-deficient mice

Abstract

In nonalcoholic fatty liver disease (NAFLD), depletion of hepatic antioxidants may contribute to the progression of steatosis to nonalcoholic steatohepatitis (NASH) by increasing oxidative stress that produces lipid peroxidation, inflammation, and fibrosis. We investigated whether depletion of glutathione (GSH) increases NASH-associated hepatic pathology in mice fed a diet deficient in methionine and choline (MCD diet). Wild-type (wt) mice and genetically GSH-deficient mice lacking the modifier subunit of glutamate cysteine ligase (Gclm null mice), the rate-limiting enzyme for de novo synthesis of GSH, were fed the MCD diet, a methionine/choline-sufficient diet, or standard chow for 21 days. We assessed NASH-associated hepatic pathology, including steatosis, fibrosis, inflammation, and hepatocyte ballooning, and used the NAFLD Scoring System to evaluate the extent of changes. We measured triglyceride levels, determined the level of lipid peroxidation products, and measured by qPCR the expression of mRNAs for several proteins associated with lipid metabolism, oxidative stress, and fibrosis. MCD-fed GSH-deficient Gclm null mice were to a large extent protected from MCD diet-induced excessive fat accumulation, hepatocyte injury, inflammation, and fibrosis. Compared with wt animals, MCD-fed Gclm null mice had much lower levels of F2-isoprostanes, lower expression of acyl-CoA oxidase, carnitine palmitoyltransferase 1a, uncoupling protein-2, stearoyl-coenzyme A desaturase-1, transforming growth factor-β, and plas-minogen activator inhibitor-1 mRNAs, and higher activity of catalase, indicative of low oxidative stress, inhibition of triglyceride synthesis, and lower expression of profibrotic proteins. Global gene analysis of hepatic RNA showed that compared with wt mice, the livers of Gclm null mice have a high capacity to metabolize endogenous and exogenous compounds, have lower levels of lipogenic proteins, and increased antioxidant activity. Thus, metabolic adaptations resulting from severe GSH deficiency seem to protect against the development of steatohepatitis.

Reprint of “In utero exposure to benzo[a]pyrene increases adiposity and causes hepatic steatosis in female mice, and glutathione deficiency is protective”

“…No ballooning, a feature of steatohepatitis, was noted in any of the livers. The nonalcoholic fatty liver disease scores we obtained in the control Gclm −/− and Gclm +/+ mice in the present study are very similar to those reported for control Gclm −/− and Gclm +/+ mice in another recent study (Haque et al, 2010). Table 3 Effects of prenatal BaP exposure and Gclm genotype on hepatic nonalcoholic fatty liver disease (NAFLD) scores (estimated marginal means ± SEM).…”

“…Our previous studies showed that Gclm −/− mice are more sensitive to the gonadal toxicity of gestational exposure to BaP than wild type littermates (Lim et al, 2013;Nakamura et al, 2012) and that female Gclm −/− mice are subfertile (Nakamura et al, 2011). In contrast, Gclm null mice are protected against diet-induced steatohepatitis, showing upregulation of hepatic antioxidant genes and downregulation of triglyceride synthesis and fatty acid ␤-oxidation (Haque et al, 2010;Kendig et al, 2011). Nonalcoholic hepatic steatosis, also called nonalcoholic fatty liver disease, is an independent risk factor for Type 2 diabetes and is prevalent in individuals with metabolic syndrome (Sung and Kim, 2011).…”

A novel murine model for non-alcoholic steatohepatitis developed by combination of a high-fat diet and oxidized low-density lipoprotein

“…28,40 Furthermore, lipid peroxidation, as the main second hit of the two-hit hypothesis, contributes to the pathogenic transition from steatosis to NASH by activating hepatic macrophages to trigger inflammation and inducing hepatocyte injury. 8,41 Therefore, the hepatic injury observed in the HFD-fed mice is considered to be at least partly attributable to elevated oxidative lipids (Figures 1b and c, and 3a). Although 23 weeks of treatment with HFD failed to increase hepatic recruitment of inflammatory cells in the mice, elevated TNF-a gene expression was detected in their livers, showing a trend toward the Th-1 polarized inflammatory response (Figures 2a, and 4a and c-f, and Table 4).…”

“…Hepatic steatosis induced by lipid accumulation in hepatocytes is recognized as the first hit, 7,8 and is also identified as a critical prerequisite for the development of inflammation and an independent risk factor for liver fibrosis. 9,10 The second hits are presumably considered to contribute to the progression from simple steatosis to steatohepatitis.…”

Hepatoprotective effect of tamoxifen on steatosis and non-alcoholic steatohepatitis in mouse models

“…In NASH, the fatty acid overload plays an important role in ROS generation as a result of electron leakage during mitochondrial β-oxidation in energy production (Haque et al, 2010). Levels of CPT-1, a ratelimiting regulator of mitochondrial β-oxidation through its role in mitochondrial fatty acid import, were increased in the steatosis model mice, whereas administration of TAM significantly decreased CPT-1 levels compared to the TAM-unadministered mice.…”