There is interest in the role of oxidative stress and generation of reactive radical species in the mechanism(s) by which ethanol is toxic. Induction of CYP2E1 is one pathway by which ethanol generates oxidative stress. S-Adenosyl- L-Methionine (SAM) is a regulator of cellular growth, differentiation and function. Impairment of SAM synthesis plays an important role in hepatic injury induced by various agents, including alcohol. CYP2E1 levels were increased in the MAT1A knockout mouse suggesting SAM could regulate or modulate CYP2E1. The goal of this application is to study possible interactions/modulation between CYP2E1 and SAM and to investigate the effects of SAM on CYP2E1- dependent toxicity and generation of reactive oxygen species.
Aim 1 will evaluate the effect of SAM, and the SAM metabolite 5-methylthioadenosine (MTA) on CYP2E 1-dependent toxicity in cultured hepatocytes from pyrazole-treated rats and HepG2 cells overexpressing CYP2E1 (E47 cells).
Aim 2 will study the effect of SAM and MTA on hepatic stellate cell activation by CYP2El-derived diffusible mediators in co-cultures of primary hepatic stellate cells with pyrazole hepatocytes or E47 cells.
Aim 3 will assess the effect of SAM and MTA on CYP2El-dependent activation of antioxidants genes which reflect an adaptive response to CYP2El-dependent oxidative stress. The ability of SAM or MTA to prevent activation of P38 MAP kinasc or other stress kinascs by CYP2E1 will be determined, since such actions may be important in mechanisms by which SAM or MTA prevent CYP2E1 toxicity.
Aim 4 will study in-vivo effects of SAM and MTA on CYP2E1 expression, content and actions. Control rats or rats induced by pyrazole, ethanol, starvation with high levels of CYP2E 1 will be treated with SAM or MTA in-vivo and the effect on basal or induced CYP2E1 protein, activity, mRNA level on up regulation of GSH and antioxidants and on CYP2El-dependent toxicity in several in-vivo models determined. The effect of CYP2E1 induction on expression of the MAT1A and MAT2A genes or enzyme activities responsible for the synthesis of SAM will be determined.
Aim 5 will evaluate the ability of SAM or MTA, in-vitro, to inhibit CYP2El-dependent generation of reactive oxygen species. It is hoped that this study utilizing hepatocyte cell culture models, in-vivo models and mechanistic studies will help to define the effects of SAM on CYP2E 1-dependent toxicity and may prove valuable in understanding the hepatoprotective actions of SAM in many models of liver injury, including alcohol-induced liver injury.

Agency
National Institute of Health (NIH)
Institute
National Institute on Alcohol Abuse and Alcoholism (NIAAA)
Type
Research Project (R01)
Project #
5R01AA014132-03
Application #
6795965
Study Section
Special Emphasis Panel (ZAA1-DD (23))
Program Officer
Gentry, Thomas
Project Start
2002-09-30
Project End
2007-08-31
Budget Start
2004-09-01
Budget End
2005-08-31
Support Year
3
Fiscal Year
2004
Total Cost
$339,000
Indirect Cost
Name
Mount Sinai School of Medicine
Department
Pharmacology
Type
Schools of Medicine
DUNS #
078861598
City
New York
State
NY
Country
United States
Zip Code
10029
Lu, Yongke; Cederbaum, Arthur I (2008) CYP2E1 and oxidative liver injury by alcohol. Free Radic Biol Med 44:723-38
Wang, Xiaodong; Cederbaum, Arthur I (2008) S-adenosyl-L-methionine decreases the elevated hepatotoxicity induced by Fas agonistic antibody plus acute ethanol pretreatment in mice. Arch Biochem Biophys 477:1-11
Zhuge, Jian (2008) A decrease in S-adenosyl-L-methionine potentiates arachidonic acid cytotoxicity in primary rat hepatocytes enriched in CYP2E1. Mol Cell Biochem 314:105-12
Zhuge, Jian; Cederbaum, Arthur I (2007) Depletion of S-adenosyl-l-methionine with cycloleucine potentiates cytochrome P450 2E1 toxicity in primary rat hepatocytes. Arch Biochem Biophys 466:177-85
Dey, Aparajita; Caro, Andres A; Cederbaum, Arthur I (2007) S-adenosyl methionine protects ob/ob mice from CYP2E1-mediated liver injury. Am J Physiol Gastrointest Liver Physiol 293:G91-103
Wang, Xiaodong; Cederbaum, Arthur I (2006) S-adenosyl-L-methionine attenuates hepatotoxicity induced by agonistic Jo2 Fas antibody following CYP2E1 induction in mice. J Pharmacol Exp Ther 317:44-52
Wu, Defeng; Cederbaum, Arthur (2006) Nitric oxide donors prevent while the nitric oxide synthase inhibitor L-NAME increases arachidonic acid plus CYP2E1-dependent toxicity. Toxicol Appl Pharmacol 216:282-92
Wu, Defeng; Cederbaum, Arthur I (2006) Opposite action of S-adenosyl methionine and its metabolites on CYP2E1-mediated toxicity in pyrazole-induced rat hepatocytes and HepG2 E47 cells. Am J Physiol Gastrointest Liver Physiol 290:G674-84
Nieto, Natalia; Cederbaum, Arthur I (2005) S-adenosylmethionine blocks collagen I production by preventing transforming growth factor-beta induction of the COL1A2 promoter. J Biol Chem 280:30963-74
Caro, Andres A; Cederbaum, Arthur I (2005) Inhibition of CYP2E1 catalytic activity in vitro by S-adenosyl-L-methionine. Biochem Pharmacol 69:1081-93

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