Cholesterol Induces Oxidative Stress, Mitochondrial Damage and Death in Hepatic Stellate Cells to Mitigate Liver Fibrosis in Mice Model of NASH.

Liver fibrosis and its end-stage disease cirrhosis are major world health problems arising from chronic injury of the liver. In recent years, the hypothesis that hepatic stellate cells' (HSCs') activation and fibrosis can be mitigated by HSC apoptosis and cell death has become of interest. In the current study, we evaluated the effect of cholesterol and bile acids on HSC apoptosis and liver fibrosis. Male C57BL/6J mice (wild type), aged four to five weeks, were fed an AIN-93G based diet (normal diet, ND), ND diet + 1% (w/w) cholesterol (CHOL group), ND diet + 0.5% (w/w) cholic acid (CA group) or ND diet + 1% (w/w) cholesterol + 0.5% (w/w) cholic acid (CHOL + CA group). Female Mdr2(-/-) mice were also treated with ND with and without 1% cholesterol. The effect of cholesterol on liver fibrosis and HSC clearance was evaluated. In addition, we studied the mechanism of cholesterol-induced apoptosis in HSC-T6 and AML-12 hepatocyte cell lines. In animals treated with cholic acids, increased lipid peroxidation and fibrosis were observed after six weeks of treatment. However, addition of cholesterol to the diet of C57BL/6J mice led to HSC-specific apoptosis and resolution of liver fibrosis, verified by double-staining with active caspase and α smooth muscle actin antibodies. In Mdr2 (-/-) mice, a diet supplemented with cholesterol corrected fibrosis and induced active hepatic stellate cells' clearance. HSC-T6 were found to be much more sensitive to cholesterol-induced oxidative stress, mitochondrial damage and apoptosis compared to hepatocytes. These results indicate that cholesterol may be a trigger of HSC lipid peroxidation and death in the liver in a model of non-alcoholic steatohepatitis. A high cholesterol-to-bile acid ratio may determine the trajectory of the liver disease toward mitigation of fibrosis.

Whole body metabolism is improved by hemin added to high fat diet while counteracted by nitrite: a mouse model of processed meat consumption.

Nitrites and nitrates are traditional food additives used as curing agents in the food industry. They inhibit the growth of microorganisms and give a typical pink color to meat. Besides the positive effects of nitrite in foods, if present at high levels in the body, may induce hypoxia and contribute to the production of pro-carcinogenic secondary N-nitrosamines. This study investigated the whole-body metabolic effects of hemin and nitrite added to a high fat diet as red and processed red meat nutritional models. Mice were fed for 11 weeks with five different diets-(1) control diet (ND), (2) high fat diet (HFD) with 60% fat, (3) HFD with hemin (HFD + H, red meat model), (4) HFD with hemin and nitrite (HFD + HN, processed meat model), and (5) HFD with hemin, nitrite, and secondary amine (HFD + HNN, N-nitrosamine generating model)-and several metabolic parameters were determined and respiratory measurements were performed. Mice fed with the HFD + H or HFD + HNN diet had a lower epididymal white adipose tissue (eWAT) : body ratio and lower fasting glucose level than those fed the HFD alone. In addition, our results demonstrated a relief in hepatosteatosis grade among the HFD + H and HFD + HNN diet fed mice. Nitrite added to the HFD impaired the ability to use fat for energy, opposite to the effect of hemin. This study shows that nitrite in addition to pro-carcinogenesis and hypoxia can impact metabolic disease progression when added to meat.

Cholesterol Induces Nrf-2- and HIF-1α-Dependent Hepatocyte Proliferation and Liver Regeneration to Ameliorate Bile Acid Toxicity in Mouse Models of NASH and Fibrosis.

Nonalcoholic steatohepatitis (NASH) is currently one of the most common liver diseases worldwide. The toxic effects of lipids and bile acids contribute to NASH. The regenerative pathway in response to damage to the liver includes activation of the inflammatory process and priming of hepatocytes to proliferate to restore tissue homeostasis. However, the effects of cholesterol on bile acid toxicity, inflammation, and fibrosis remain unknown. We have used two mouse models of bile acid toxicity to induce liver inflammation and fibrosis. A three-week study was conducted using wild-type mice receiving an atherogenic diet (1% (w/w) cholesterol and 0.5% (w/w) cholic acid) and its separate constituents. Mdr2-/- mice were fed a high-cholesterol-enriched diet or standard AIN-93 diet for 6 weeks. We measured serum transaminase levels to assess liver tissue necrosis and fibrosis; iNOS, SAA1, SAA2, and F4/80 levels to determine liver inflammation; PCNA and HGF levels to evaluate proliferative response; and Nrf-2, HIF-1α, and downstream gene expression to establish protective responses. In both studies, high bile acid levels increased serum transaminases and liver fibrosis, whereas cholesterol supplementation attenuated these effects. Cholesterol supplementation activated survival and the robustness of HIF-1α and Nrf-2 gene expression in hepatocytes, induced liver inflammation and hepatocyte proliferation, and inhibited stellate cell hyperplasia and fibrosis. In conclusion, our data show for the first time that cholesterol intake protects against bile acid liver toxicity. The balance between hepatic cholesterol and bile acid levels may be of prognostic value in liver disease progression and trajectory.