Skip to main content

Advertisement

Springer Nature Link
Log in

Alcohol oxidizing enzymes and ethanol-induced cytotoxicity in rat pancreatic acinar AR42J cells

  • Published:
In Vitro Cellular & Developmental Biology - Animal Aims and scope Submit manuscript

Abstract

Alcoholic chronic pancreatitis (ACP) is a serious inflammatory disease causing significant morbidity and mortality. Due to lack of a suitable animal model, the underlying mechanism of ACP is poorly understood. Chronic alcohol abuse inhibits alcohol dehydrogenase (ADH) and facilitates nonoxidative metabolism of ethanol to fatty acid ethyl esters (FAEEs) in the pancreas frequently damaged during chronic ethanol abuse. Earlier, we reported a concentration-dependent formation of FAEEs and cytotoxicity in ethanol-treated rat pancreatic tumor (AR42J) cells, which express high FAEE synthase activity as compared to ADH and cytochrome P450 2E1. Therefore, the present study was undertaken to investigate the role of various ethanol oxidizing enzymes in ethanol-induced pancreatic acinar cell injury. Confluent AR42J cells were pre-treated with inhibitors of ADH class I and II [4-methylpyrazole (MP)] or class I, II, and III [1,10-phenanthroline (PT)], cytochrome P450 2E1 (trans-1,2-dichloroethylene) or catalase (sodium azide) followed by incubation with 800 mg% ethanol at 37°C for 6 h. Ethanol metabolism, cell viability, cytotoxicity (apoptosis and necrosis), cell proliferation status, and formation of FAEEs in AR42J cells were measured. The cell viability and cell proliferation rate were significantly reduced in cells pretreated with 1,10-PT + ethanol followed by those with 4-MP + ethanol. In situ formation of FAEEs was twofold greater in cells incubated with 1,10-PT + ethanol and ∼1.5-fold in those treated with 4-MP + ethanol vs. respective controls. However, cells treated with inhibitors of cytochrome P450 2E1 or catalase in combination of ethanol showed no significant changes either for FAEE formation, cell death or proliferation rate. Therefore, an impaired ADH class I–III catalyzed oxidation of ethanol appears to be a key contributing factor in ethanol-induced pancreatic injury via formation of nonoxidative metabolites of ethanol.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Subscribe and save

Springer+ Basic
$34.99 /Month
  • Get 10 units per month
  • Download Article/Chapter or eBook
  • 1 Unit = 1 Article or 1 Chapter
  • Cancel anytime
Subscribe now

Buy Now

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Figure 1.
Figure 2.
Figure 3.
Figure 4.
Figure 5.
Figure 6.

Similar content being viewed by others

Explore related subjects

Discover the latest articles and news from researchers in related subjects, suggested using machine learning.

References

  • Bhat H. K.; Ansari G. A. S. Cholesterol ester hydrolase-mediated conjugation of haloethanols with fatty acids. Chem. Res. Toxicol. 3: 311–317; 1990.

    Article  PubMed  CAS  Google Scholar 

  • Bhopale K.; Wu H.; Boor P. J.; Ansari G. A. S.; Kaphalia B. S. Metabolic basis of ethanol-induced hepatic and pancreatic toxicity in hepatic alcohol dehydrogenase deficient deer mice. Alcohol 39: 179–188; 2006.

    Article  PubMed  CAS  Google Scholar 

  • Brunner R.; Xie J.; Bank S. Does acute alcoholic pancreatitis exist with preexisting chronic pancreatitis? J. Clin. Gastroenterol. 38: 201–202; 2004.

    Article  PubMed  Google Scholar 

  • Christophe J. Pancreatic tumoral cell line AR42J. An amophicrine model. Am. J. Physiol. 266: G963–G971; 1994.

    PubMed  CAS  Google Scholar 

  • Chrostek L.; Jelski W.; Szmitkowski M.; Puchalski Z. Alcohol dehydrogenase (ADH) isoenzymes and aldehyde dehydrogenase (ALDH) activity in the human pancreas. Dig. Dis. Sci. 48: 1230–1233; 2003.

    Article  PubMed  CAS  Google Scholar 

  • Criddle D. N.; Raraty M. G.; Neoptolemos J. P.; Tepikin A. V.; Petersen O. H.; Sutton R. Ethanol toxicity in pancreatic acinar cells: mediation by nonoxidative fatty acid metabolites. Proc. Natl. Acad. Sci. U. S. A. 101: 10738–10743; 2004.

    Article  PubMed Central  PubMed  CAS  Google Scholar 

  • Criddle D. N.; Sutton R.; Petersen O. H. Role of Ca2+ in pancreatic cell death induced by alcohol metabolites. J. Gastroenterol. Hepatol. Suppl. 3: S14–S17; 2006.

    Article  CAS  Google Scholar 

  • Fernando H.; Wiktorowicz J. E.; Soman K. V.; Kaphalia B. S.; Khan M. F.; Ansari G. A. S. Liver proteomics in progressive alcoholic steatosis. Toxicol. Appl. Pharmacol. 266: 470–480; 2013.

    Google Scholar 

  • Foster J. R.; Idle J. R.; Hardwick J. P. et al. Induction of drug-metabolizing enzymes in human pancreatic cancer and chronic pancreatitis. J. Pathol. 169: 457–463; 1993.

    Article  PubMed  CAS  Google Scholar 

  • Gukovskaya A. S.; Mouria M.; Gukovskaya I.; Reyes C. N.; Kasho V. N.; Faller L. D.; Pandol S. J. Ethanol metabolism and transcription factor activation in pancreatic acinar cells in rats. Gastroenterology 122: 106–118; 2002.

    Article  PubMed  CAS  Google Scholar 

  • Haber P. S.; Apte M. V.; Applegate T. L.; Norton I. D.; Korsten M. A.; Pirola R. C.; Wilson J. S. Metabolism of ethanol by rat pancreatic acinar cells. J. Lab. Clin. Med. 132: 294–302; 1998.

    Article  PubMed  CAS  Google Scholar 

  • Haber P. S.; Wilson J. S.; Apte M. V.; Pirola R. C. Fatty acid ethyl esters increases rat pancreatic lysosomal fragility. J. Lab. Clin. Med. 121: 759–764; 1993.

    PubMed  CAS  Google Scholar 

  • Kaphalia B. S. Pancreatic toxicology. In: Nriagu J. O. (ed) Encyclopedia of Environmental Health, vol. 4. Elsevier, Burlington, pp 315–324; 2011.

    Chapter  Google Scholar 

  • Kaphalia B. S.; Ansari G. A. S. Fatty acid ethyl esters and ethanol-induced pancreatic injury. Cell. Mol. Biol. 47: OL173–OL179; 2001.

    PubMed  CAS  Google Scholar 

  • Kaphalia B. S.; Ansari G. A. S. Purification and characterization of rat pancreatic fatty acid ethyl ester synthase and its structural and functional relationship to pancreatic cholesterol esterase. J. Biochem. Mol. Toxicol. 17: 338–345; 2003.

    Article  PubMed  CAS  Google Scholar 

  • Kaphalia B. S.; Cai P.; Khan M. F.; Okorodudu A. O.; Ansari G. A. S. Fatty acid ethyl esters; markers of alcohol abuse and alcoholism. Alcohol 34: 151–158; 2004.

    Article  PubMed  CAS  Google Scholar 

  • Kaphalia B. S.; Green S. M.; Ansari G. A. S. Fatty acid ethyl and methyl ester synthases, and fatty acid anilide synthase in HepG2 and AR42J cells: interrelationships and inhibition by tri-o-tolyl phosphate. Toxicol. Appl. Pharmacol. 159: 134–141; 1999.

    Google Scholar 

  • Kaphalia B. S.; Khan M. F.; Carroll R. M.; Aronson J.; Ansari G. A. S. Subchronic toxicity of 2-chloroethanol and 2-bromoethanol in rats. Res. Commun. Pharmacol. Toxicol. 1: 173–186; 1996.

    CAS  Google Scholar 

  • Kessova I. G.; DeCarli L. M.; Lieber C. S. Inducibility of cytochrome P4502E1 and P4501A1 in the rat pancreas. Alcohol. Clin. Exp. Res. 22: 501–504; 1998.

    Article  PubMed  CAS  Google Scholar 

  • Lankisch P. G.; Banks P. A. Pancreatitis. Springer, Berlin, Heidelberg, New York, p 377; 1998.

    Book  Google Scholar 

  • Lange L. G.; Sobel B. E. Mitochondrial dysfunction induced by fatty acid ethyl esters, myocardial metabolites of ethanol. J. Clin. Invest. 72: 724–731; 1983.

    Article  PubMed Central  PubMed  CAS  Google Scholar 

  • Laposata E. A.; Lange L. G. Presence of nonoxidative ethanol metabolism in humans organs commonly damaged by ethanol abuse. Science 231: 497–499; 1986.

    Article  PubMed  CAS  Google Scholar 

  • Manautou J. E.; Carlson G. P. Ethanol-induced fatty acid ethyl ester formation in vivo and in vitro in rat lung. Toxicology 70: 303–312; 1991.

    Article  PubMed  CAS  Google Scholar 

  • Mathews J. M.; Etheridge A. S.; Raymer J. H.; Black S. R.; Pulliam Jr. D. W.; Bucher J. R. Selective inhibition of cytochrome P450 2E1 in vivo and in vitro with trans-1,2-dichloroethylene. Chem. Res. Toxicol. 11: 778–785; 1998.

    Article  PubMed  CAS  Google Scholar 

  • Norton I. D.; Apte M. V.; Haber P. S.; McCaughan G. W.; Pirola R. C.; Wilson J. S. Cytochrome P4502E1 is present in rat pancreas and is induced by chronic ethanol administration. Gut 42: 426–430; 1998.

    Article  PubMed Central  PubMed  CAS  Google Scholar 

  • Nuutinen H.; Lindros K. O.; Salaspuro M. Determinants of blood acetaldehyde level during ethanol oxidation in chronic alcoholics. Alcohol. Clin. Exp. Res. 7: 163–168; 1983.

    Article  PubMed  CAS  Google Scholar 

  • Panes J.; Caballeria J.; Guitart R.; Pares A.; Soler X.; Rodamilans M.; Navasa M.; Pares X.; Bosch J.; Rodes J. Determinants of ethanol and acetaldehyde metabolism in chronic alcoholics. Alcohol. Clin. Exp. Res. 17: 48–53; 1993.

    Article  PubMed  CAS  Google Scholar 

  • Panes J.; Soler X.; Pares A.; Caballeria J.; Farres J.; Rodes J.; Pares X. Influence of liver disease on hepatic alcohol and aldehyde dehydrogenases. Gastroenterology 97: 708–714; 1989.

    PubMed  CAS  Google Scholar 

  • Pfutzer R. H.; Tadic S. D.; Li H. S.; Thompson B. S.; Zhang J. Y.; Ford M. E.; Eagon P. K.; Whitcomb D. C. Pancreatic cholesterol esterase, ES-10, and fatty acid ethyl ester synthase III gene expression are increased in the pancreas and liver but not in the brain or heart with long-term ethanol feeding in rats. Pancreas 25: 101–106; 2002.

    Article  PubMed  Google Scholar 

  • Pitchumoni C. S.; Glasser M.; Saran R. M.; Panchacharam P.; Thelmo W. Pancreatic fibrosis in chronic alcoholics and non alcoholics without clinical pancreatitis. Am. J. Gastroenterol. 79: 382–388; 1984.

    PubMed  CAS  Google Scholar 

  • Rognstead R.; Grunnet N. Enzymatic pathways of ethanol metabolism. In: Majchrowicz E.; Noble E. P. (eds) Biochemistry and Pharmacology of Ethanol. Plenum Press, New York, pp 65–86; 1979.

    Chapter  Google Scholar 

  • Singh M.; Simsek H. Ethanol and the pancreas: current status. Gastroenterology 98: 1051–1062; 1990.

    Article  PubMed  CAS  Google Scholar 

  • Werner J.; Laposata M.; Fernandez-del Castillo C.; Saghir M.; Iozzo R. V.; Lewandrowski K. B.; Warshaw A. L. Pancreatic injury in rats induced by fatty acid ethyl ester, a nonoxidative metabolite of alcohol. Gastroenterology 113: 286–294; 1997.

    Article  PubMed  CAS  Google Scholar 

  • Werner J.; Saghir M.; Warshaw A. L.; Lewandrowski K. B.; Laposata M.; Iozzo R. V.; Carter E. A.; Schatz R. J.; Fernandez-Del C. C. Alcoholic pancreatitis in rats: injury from nonoxidative metabolites of ethanol. Am. J. Physiol. Gastroentest. Liver Physiol. 283: G65–G73; 2002.

    CAS  Google Scholar 

  • Wilson J. S.; Korsten M. A.; Apte M. V.; Thomas M. C.; Haber P. S.; Pirola R. C. Both ethanol consumption and protein deficiency increase the fragility of pancreatic lysosomes. J. Lab. Clin. Med. 115: 749–755; 1990.

    PubMed  CAS  Google Scholar 

  • Wu H.; Bhopale K. K.; Ansari G. A. S.; Kaphalia B. S. Ethanol-induced cytotoxicity in rat pancreatic acinar AR42J cells: role of fatty acid ethyl esters. Alcohol Alcohol. 43: 1–8; 2008.

    Article  PubMed  CAS  Google Scholar 

  • Wu H.; Cai P.; Clemens D. L.; Jerrells T. R.; Ansari G. A. S.; Kaphalia B. S. Metabolic basis of ethanol-induced cytotoxicity in recombinant HepG2 cells: role of nonoxidative metabolism. Toxicol. Appl. Pharmacol. 216: 238–247; 2006.

    Article  PubMed  CAS  Google Scholar 

Download references

Acknowledgments

This work was supported by NIH grant AA19812 from the National Institute on Alcohol Abuse and Alcoholism. The authors also acknowledge the support of NIEHS Center grant P30ES06676 for Exposure Assessment and Biomarker Development Core for the analysis of ethanol and acetaldehyde.

Author information

Authors and Affiliations

  1. Department of Pathology, The University of Texas Medical Branch, Galveston, TX, 77555-0609, USA

    Kamlesh K. Bhopale, G. A. S. Ansari & Bhupendra S. Kaphalia

  2. Departments of Pharmacology, The University of Texas Medical Branch, Galveston, TX, 77555, USA

    Miriam Falzon

Authors
  1. Kamlesh K. Bhopale
    View author publications

    You can also search for this author inPubMed Google Scholar

  2. Miriam Falzon
    View author publications

    You can also search for this author inPubMed Google Scholar

  3. G. A. S. Ansari
    View author publications

    You can also search for this author inPubMed Google Scholar

  4. Bhupendra S. Kaphalia
    View author publications

    You can also search for this author inPubMed Google Scholar

Corresponding author

Correspondence to Bhupendra S. Kaphalia.

Additional information

Editor: T. Okamoto

Rights and permissions

Reprints and permissions

About this article

Cite this article

Bhopale, K.K., Falzon, M., Ansari, G.A.S. et al. Alcohol oxidizing enzymes and ethanol-induced cytotoxicity in rat pancreatic acinar AR42J cells. In Vitro Cell.Dev.Biol.-Animal 50, 373–380 (2014). https://doi.org/10.1007/s11626-013-9700-7

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s11626-013-9700-7

Keywords