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Ethanol-induced cytotoxicity in rat pancreatic acinar AR42J cells: Role of fatty acid ethyl esters

Hai Wu, Kamlesh K. Bhopale, G. A. S. Ansari, Bhupendra S. Kaphalia
DOI: http://dx.doi.org/10.1093/alcalc/agm044 1-8 First published online: 17 October 2007

Abstract

Aims: To understand the mechanism(s) of alcoholic pancreatitis and role of fatty acid ethyl esters (FAEEs, non-oxidative metabolites of ethanol) in ethanol-induced pancreatic injury. Methods: A time- and concentration-dependent synthesis of FAEEs and the cytotoxicity of ethanol and its predominant fatty acid esters were studied in rat pancreatic tumour (AR42J) cells in cultures. Role of FAEEs in ethanol-induced cytotoxicity was investigated by measuring the synthesis of FAEEs, injury markers and apoptosis in cells incubated simultaneously with ethanol and FAEE synthase inhibitor, 3-benzyl-6-chloro-2-pyrone. The cells were pre-incubated with caspase-3 inhibitor (N-acetyl-DEVD-CHO) to measure the effect of caspase-3 inhibition on ethanol-induced apoptosis. Results: The levels of FAEEs synthesized in cell cultures incubated with 800 mg% ethanol for 6 h were ∼10-fold higher (60 nmol/25 × 106 cells) than those in cells incubated with 100 mg% ethanol (5.4 nmol/25 × 106 cells). Ethanol exposure resulted in a concentration-dependent apoptosis (10, 12 and 13% at 200, 400 and 800 mg% ethanol, respectively, vs 5% in controls). A similar concentration-dependent apoptosis was also found in the cells incubated with ethyl oleate (one of the predominant FAEEs reported in alcoholic patients). Inhibition of FAEE synthesis and resultant apoptosis was found in the cells incubated simultaneously with pancreatic FAEE synthase inhibitor and ethanol. Ethanol-induced apoptosis was significantly inhibited in cells pre-incubated with caspase-3 inhibitor. Conclusions: These results support our hypothesis that ethanol-induced cytotoxicity in AR42J cells is mediated by the non-oxidative metabolite(s) of ethanol, and caspase-3 mediated apoptosis could be one of the mechanisms involved in ethanol-induced pancreatic injury.

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