[Abstract unavailable

Reactive oxygen species (ROS) are highly reactive molecules that are naturally generated in small amounts during the body's metabolic reactions and can react with and damage complex cellular molecules such as lipids, proteins, or DNA. Acute and chronic ethanol treatments increase the production of ROS, lower cellular antioxidant levels, and enhance oxidative stress in many tissues, especially the liver. Ethanol-induced oxidative stress plays a major role in the mechanisms by which ethanol produces liver injury. Many pathways play a key role in how ethanol induces oxidative stress. This review summarizes some of the leading pathways and discusses the evidence for their contribution to alcohol-induced liver injury. Special emphasis is placed on CYP2E1, which is induced by alcohol and is reactive in metabolizing and activating many hepatotoxins, including ethanol, to reactive products, and in generating ROS.

Reactive oxygen species (ROS) are highly reactive molecules that are naturally generated in small amounts during the body's metabolic reactions and can react with and damage complex cellular molecules such as lipids, proteins, or DNA. This review describes pathways involved in ROS formation, why ROS are toxic to cells, and how the liver protects itself against ROS. Acute and chronic ethanol treatment increases the production of ROS, lowers cellular antioxidant levels, and enhances oxidative stress in many tissues, especially the liver. Ethanol-induced oxidative stress plays a major role in the mechanisms by which ethanol produces liver injury. Many pathways play a key role in how ethanol induces oxidative stress. This review summarizes some of the leading pathways and discusses the evidence for their contribution to alcohol-induced liver injury.

A major pathway for chronic ethanol-induced liver injury is ethanol-induced oxidant stress. Several pathways contribute to mechanisms by which ethanol induces oxidant stress. While some studies support a role for cytochrome P450 2E1 (CYP2E1), others do not. Most previous studies were conducted in the intragastric infusion model of ethanol administration. There is a need to develop oral models of significant liver injury and to evaluate the possible role of CYP2E1 in ethanol actions in such models. We evaluated chronic ethanol-induced liver injury, steatosis and oxidant stress in wild type (WT) mice, CYP2E1 knockout (KO) mice and in humanized CYP2E1 knockin (KI) mice, where the human 2E1 was added back to mice deficient in the mouse 2E1. WT mice and the CYP2E1 KO and KI mice (both provided by Dr F. Gonzalez, NCI) were fed a high fat Lieber-DeCarli ethanol liquid diet for 3 weeks; pair-fed controls received dextrose. Ethanol produced fatty liver and oxidant stress in WT mice but liver injury (transaminases, histopathology) was minimal. Ethanol-induced steatosis and oxidant stress was blunted in the KO mice (no liver injury) but restored in the KI mice. Signicant liver injury was produced in the ethanol-fed KI mice with elevated transaminases, necrosis, and increased levels of collagen type 1 and smooth muscle actin. This liver injury in the KI mice was associated with elevated oxidant stress and elevated levels of the human CYP2E1 compared to levels of the mouse 2E1 in WT mice. Activation of JNK and decreased levels of Bcl-2 and Bcl-XL were observed in the ethanol-fed KI mice compared to the other groups. Fatty liver in the WT and the KI mice was associated with lower levels of PPAR alpha and acyl CoA oxidase. No such changes were found in the ethanol-fed KO mice. These results show that CYP2E1 plays a major role in ethanol-induced fatty liver and oxidant stress. It is the absence of CYP2E1 in the KO mice responsible for the blunting of steatosis and oxidant stress since restoring the CYP2E1 restores the fatty liver and oxidant stress. Moreover, it is the human CYP2E1 which restores these effects of ethanol which suggests that results on fatty liver and oxidant stress from rodent models of ethanol intake and mouse CYP2E1 can be extrapolated to human models of ethanol intake and to human CYP2E1.

The effect of ethanol on the viability of a HepG2 cell model which was developed to constitutively express human CYP2E1 was studied in an attempt to establish a linkage between CYP2E1, reactive oxygen intermediates, and ethanol toxicity. Assays of toxicity included leakage of lactate dehydrogenase, trypan blue uptake, morphology, and formazan production. Ethanol was toxic to HepG2 E9 cells, which express CYP2E1, but not to HepG2 MV5 cells, which do not express CYP2E1. The ethanol toxicity was dependent on the concentration of ethanol, starting with 10 m ethanol, and on the time of incubation with ethanol. Phorbol 12-myristate 13-acetate, which increases the expression of CYP2E1 in this model, increased the toxicity by ethanol. Ethanol toxicity was prevented by 4-methylpyrazole and by diallyl sulfide, inhibitors of CYP2E1. The ethanol toxicity was also prevented by radical trapping agents such as N-acetylcysteine and N-t-butyl-alpha-phenylnitrone, antioxidative agents such as catalase, superoxide dismutase, thiourea, and uric acid, and inhibitors of lipid peroxidation, such as vitamin E phosphate, Trolox, and diphenylphenylenediamine. Besides ethanol, other substrates such as Me2SO, CCl4, isoniazid, and N,N-dimethylnitrosamine were cytotoxic to cells expressing CYP2E1 but not to control cells. These results indicate that ethanol was toxic to HepG2 cells which express human CYP2E1 by a pathway sensitive to inhibitors of CYP2E1 and to a variety of antioxidative agents. This model appears to be useful in efforts to establish a CYP2E1-dependent ethanol hepatotoxicity system and to evaluate the role of oxidative stress and reactive radical species in the toxicity by ethanol.