Abstract
The metabolism of alcohol involves cytochrome P450 2E1 (CYP2E1)-induced oxidative stress, with the association of phosphatidylinositol-3-kinases (PI3K) and nuclear factor kappa B (NFκB) signalling pathways. CYP2E1 is primarily involved in the microsomal ethanol oxidising system, which generates massive reactive oxygen species (ROS) and ultimately leads to oxidative stress and tissue damage. Lauric acid, a major fatty acid in palm kernel oil, has been shown as a potential antioxidant. Here, we aimed to evaluate the use of lauric acid as a potential antioxidant against ethanol-mediated oxidative stress by investigating its effect on CYP2E1 mRNA expression and the signalling pathway in ethanol-induced HepG2 cells. HepG2 cells were firstly treated with different concentrations of ethanol, and subsequently co-treated with different concentrations of lauric acid for 24 h. Total cellular RNA and total protein were extracted, and qPCR and Western blot was carried out. Ethanol induced the mRNA expression of CYP2E1 significantly, but lauric acid was able to downregulate the induced CYP2E1 expression in a dose-dependent manner. Similarly, Western blot analysis and densitometry analysis showed that the phosphorylated PI3K p85 (Tyr458) protein was significantly elevated in ethanol-treated HepG2 cells, but co-treatment with lauric acid repressed the activation of PI3K. However, there was no significant difference in NFκB pathway, in which the normalised NFκB p105 (Ser933) phosphorylation remained constant in any treatment conditions in this study. This suggests that ethanol induced CYP2E1 expression by activating PI3K p85 (Tyr458) pathway, but not the NFκB p105 (Ser933) pathway in HepG2 cells.
Highlights
The harmful consumption of alcohol has resulted in three million deaths worldwide and 132.6 million disability-adjusted life years in 2016 (World Health Organisation 2018)
Western blot analysis and densitometry analysis showed that the phosphorylated PI3K p85 (Tyr458) protein was significantly elevated in ethanol-treated HepG2 cells, but co-treatment with lauric acid repressed the activation of PI3K
The rise in cytochrome P450 2E1 (CYP2E1) expression is hypothesised to be associated with activation of phosphatidylinositol3-kinases (PI3K) and nuclear factor kappa B (NFκB) pathways as these pathways are involved in alcohol induction (Mandrekar & Szabo 2009; Zeng et al 2012; Wang et al 2015; Zeng et al 2018)
Summary
The harmful consumption of alcohol has resulted in three million deaths worldwide and 132.6 million disability-adjusted life years in 2016 (World Health Organisation 2018). Despite the beneficial roles of moderate alcohol consumption, excessive alcohol intake is detrimental to health and it can lead to various health problems especially alcoholic liver disease (ALD). Alcohol metabolism involves cytochrome P450 2E1 (CYP2E1) enzyme and this enzyme generates acetaldehyde as well as reactive oxygen species (ROS) which lead to ALD (Koop 2006). CYP2E1-mediated alcohol metabolism generates massive amount of reactive oxygen species (ROS) and acetaldehyde which increase the extent of liver injury and lead to the progression of ALD (Koop 2006). Ethanol administration was shown to increase CYP2E1 in wild-type mouse by fivefold but not in CYP2E1 knockout mouse (Bardag-Gorce et al 2000). In CYP2E1 knock-in mice, ethanol administration led to the accumulation of lipid and steatosis as compared to CYP2E1 knockout mice (Wu et al 2012). The rise in CYP2E1 expression is hypothesised to be associated with activation of phosphatidylinositol3-kinases (PI3K) and nuclear factor kappa B (NFκB) pathways as these pathways are involved in alcohol induction (Mandrekar & Szabo 2009; Zeng et al 2012; Wang et al 2015; Zeng et al 2018)
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