Abstract
Purpose: To establish a free fatty acid (FFA)-induced non-alcoholic fatty liver disease (NAFLD) model in HepG2 cells.Methods: HepG2 cells were incubated with 0.1, 1, or 5 mM oleic acid (OA) or palmitic acid (PA) for 24 h. Histological features were examined by oil-red-O staining. Expression levels of metabolic genes (peroxisome proliferator activated receptors α/γ, sterol regulatory element binding proteins 1a/1c, acetyl-CoA carboxylase, acyl-CoA oxidase, and fatty acid synthase), antioxidative genes (catalase and superoxide dismutases 1/2), and cytochrome P450 genes (CYP1A2, CYP2C19, CYP2D6, CYP2E1, CYP3A4, and CYP4A11) were determined by reverse transcription-real time polymerase chain reaction (RT-qPCR).Results: Intracellular lipid storage was observed in cells treated with 1 mM OA or PA while cell shrinkage was present at 5 mM concentrations of both. Expression of all metabolic genes were elevated by 1 mM PA and 5 mM OA and PA. Expression of all antioxidative genes were diminished by 5 mM OA whereas 5 mM PA only reduced superoxide dismutase-2 expression. Expression of CYP1A2, CYP2D6, and CYP3A4 genes were down-regulated by both FFAs, CYP2C19 was induced by PA, while CYP2E1 and CYP4A11 were up-regulated in a concentration-dependent manner.Conclusion: PA was the more potent steatogenic agent in an OA- or PA- induced NAFLD model in HepG2 cells. Increase in intracellular hepatic lipid and expression of metabolic genes, suppression of antioxidative genes, suppression of CYP1A2, CYP2D6, and CYP3A4, and induction of CYP2E1 andCYP4A11 correlated with the multiple-hit pathogenesis model of NAFLD. These findings suggest that PA-induced NAFLD model in HepG2 cells is a suitable in vitro model for studying novel therapeutic approaches to NAFLD treatment.
 Keywords: NAFLD, Multiple-hit pathogenesis, Free fatty acid, Oleic acid, Palmitic acid
Highlights
Non-alcoholic fatty liver disease (NAFLD) is a major chronic liver disease affecting around 30 %of Western and Asian populations [1]
The control HepG2 cells demonstrated regular cell characteristics (Figure 1 A) while those exposed to oleic acid (OA) and palmitic acid (PA) free fatty acids (FFA) showed increasing storage of intracellular lipid in a concentration-dependent manner
The lowest concentration (0.1 mM) of OA (Figure 1 C) or PA (Figure 1 F) caused limited accumulation of lipid droplets with otherwise unchanged histomorphology compared to the controls
Summary
Non-alcoholic fatty liver disease (NAFLD) is a major chronic liver disease affecting around 30 %of Western and Asian populations [1]. Non-alcoholic fatty liver disease (NAFLD) is a major chronic liver disease affecting around 30 %. Fatty liver disease patients have been given lifestyle modification advice comprised of recommendations on diet and encouragement to CYP is a superfamily of mono-oxygenase enzymes that are highly abundant in the liver and play a key role in metabolism of drugs, xenobiotics, and toxic chemicals [6]. CYPs have been implicated in the pathogenesis of fatty liver disease by promoting oxidative stress and inflammation, the metabolic pathways have not been sufficiently described [6]. NAFLD pathogenesis is believed to occur through multiple-hit including insulin resistance, mitochondrial dysfunction, endoplasmic reticulum (ER) stress, oxidantantioxidant imbalance, and inflammation, all of which are targets for NAFLD therapies.
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