Advertisement

Archives of Toxicology

, Volume 88, Issue 2, pp 391–401 | Cite as

Circulating acylcarnitines as biomarkers of mitochondrial dysfunction after acetaminophen overdose in mice and humans

  • Mitchell R. McGill
  • Feng Li
  • Matthew R. Sharpe
  • C. David Williams
  • Steven C. Curry
  • Xiaochao Ma
  • Hartmut JaeschkeEmail author
Molecular Toxicology

Abstract

Acetaminophen (APAP) is a widely used analgesic. However, APAP overdose is hepatotoxic and is the primary cause of acute liver failure in the developed world. The mechanism of APAP-induced liver injury begins with protein binding and involves mitochondrial dysfunction and oxidative stress. Recent efforts to discover blood biomarkers of mitochondrial damage have identified increased plasma glutamate dehydrogenase activity and mitochondrial DNA concentration in APAP overdose patients. However, a problem with these markers is that they are too large to be released from cells without cell death or loss of membrane integrity. Metabolomic studies are more likely to reveal biomarkers that are useful at early time points, before injury begins. Similar to earlier work, our metabolomic studies revealed that acylcarnitines are elevated in serum from mice after treatment with toxic doses of APAP. Importantly, a comparison with furosemide demonstrated that increased serum acylcarnitines are specific for mitochondrial dysfunction. However, when we measured these compounds in plasma from humans with liver injury after APAP overdose, we could not detect any significant differences from control groups. Further experiments with mice showed that N-acetylcysteine, the antidote for APAP overdose in humans, can reduce acylcarnitine levels in serum. Altogether, our data do not support the clinical measurement of acylcarnitines in blood after APAP overdose due to the standard N-acetylcysteine treatment in patients, but strongly suggest that acylcarnitines would be useful mechanistic biomarkers in other forms of liver injury involving mitochondrial dysfunction.

Keywords

Acetaminophen toxicity Biomarkers Mitochondria Acylcarnitines 

Abbreviations

ALT

Alanine aminotransferase

APAP

Acetaminophen

CACT

Carnitine-acylcarnitine translocase

CPT

Carnitine palmitoyltransferase

GDH

Glutamate dehydrogenase

JNK

c-Jun N-terminal kinase

MLK3

Mixed lineage kinase 3

MPT

Mitochondrial membrane permeability transition

mtDNA

Mitochondrial DNA

NAC

N-acetylcysteine

NAPQI

N-acetyl-p-benzoquinone imine

OPLS-DA

Orthogonal projection to latent structures-discriminant analysis

PCA

Principal component analysis

ROS

Reactive oxygen species

Notes

Acknowledgments

This work was supported in part by Grants from McNeil Consumer Health Inc. (to H.J. and S.C.C.), by the University of Kansas Medical Center Liver Center (to H.J.), by the National Institutes of Health Grants R01 DK070195 and R01 AA12916 (to H.J.), and by Grants from the National Center for Research Resources (5P20RR021940-07) and the National Institute of General Medical Sciences (8 P20 GM103549-07) of the National Institutes of Health. Additional support came from an Institutional Development Award (IDeA) from the National Institute of General Medical Sciences of the National Institutes of Health under Grant number P20 GM12345 and, from the “Training Program in Environmental Toxicology” T32 ES007079-26A2 (to M.R.M. and C.D.W.) from the National Institute of Environmental Health Sciences.

Conflict of interest

The authors declare no competing financial interest.

Supplementary material

204_2013_1118_MOESM1_ESM.docx (560 kb)
Supplementary material 1 (DOCX 560 kb)

References

  1. Antoine DJ, Dear JW, Starkey-Lewis P, Platt V, Coyle J, Masson M, Thanacoody RH, Gray AJ, Webb DJ, Moggs JG, Bateman DN, Goldring CE, Park BK (2013) Mechanistic biomarkers provide early and sensitive detection of acetaminophen-induced acute liver injury at first presentation to hospital. Hepatology 58:777–787PubMedCentralPubMedCrossRefGoogle Scholar
  2. Bajt ML, Knight TR, Farhood A, Jaeschke H (2003) Scavenging peroxynitrite with glutathione promotes regeneration and enhances survival during acetaminophen-induced liver injury in mice. J Pharmacol Exp Ther 307:67–73PubMedCrossRefGoogle Scholar
  3. Bajt ML, Knight TR, Lemasters JJ, Jaeschke H (2004) Acetaminophen-induced oxidant stress and cell injury in cultured mouse hepatocytes: protection by N-acetylcysteine. Toxicol Sci 80:343–349PubMedCrossRefGoogle Scholar
  4. Bajt ML, Cover C, Lemasters JJ, Jaeschke H (2006) Nuclear translocation of endonuclease G and apoptosis-inducing factor during acetaminophen-induced liver cell injury. Toxicol Sci 94:217–225PubMedCrossRefGoogle Scholar
  5. Bajt ML, Farhood A, Lemasters JJ, Jaeschke H (2008) Mitochondrial bax translocation accelerates DNA fragmentation and cell necrosis in a murine model of acetaminophen hepatotoxicity. J Pharmacol Exp Ther 324:8–14PubMedCrossRefGoogle Scholar
  6. Bajt ML, Ramachandran A, Yan HM, Lebofsky M, Farhood A, Lemasters JJ, Jaeschke H (2011) Apoptosis-inducing factor modulates mitochondrial oxidant stress in acetaminophen hepatotoxicity. Toxicol Sci 122:598–605PubMedCrossRefGoogle Scholar
  7. Bi H, Li F, Krausz KW, Qu A, Johnson CH, Gonzalez FJ (2013) Targeted metabolomics of serum acylcarnitines evaluates hepatoprotective effect of wuzhi tablet (Schisandra sphenanthera extract) against acute acetaminophen toxicity. Evid Based Complement Alternat Med 2013:985257PubMedCentralPubMedGoogle Scholar
  8. Chen C, Krausz KW, Shah YM, Idle JR, Gonzalez FJ (2009) Serum metabolomics reveals irreversible inhibition of fatty acid beta-oxidation through the suppression of PPARalpha activation as a contributing mechanism of acetaminophen-induced hepatotoxicity. Chem Res Toxicol 22:699–707PubMedCentralPubMedCrossRefGoogle Scholar
  9. Cover C, Mansouri A, Knight TR, Bajt ML, Lemasters JJ, Pessayre D, Jaeschke H (2005) Peroxynitrite-induced mitochondrial and endonuclease-mediated nuclear DNA damage in acetaminophen hepatotoxicity. J Pharmacol Exp Ther 315:879–887PubMedCrossRefGoogle Scholar
  10. Dahlin DC, Miwa GT, Lu AY, Nelson SD (1984) N-acetyl-p-benzoquinone imine: a cytochrome P-450-mediated oxidation product of acetaminophen. Proc Natl Acad Sci USA 81:1327–1331PubMedCrossRefGoogle Scholar
  11. Davern TJ II, James LP, Hinson JA, Polson J, Larson AM, Fontana RJ, Lalani E, Munoz S, Shakil AO, Lee WM, Acute Liver Failure Study Group (2006) Measurement of serum acetaminophen-protein adducts in patients with acute liver failure. Gastroenterology 130:687–694PubMedCrossRefGoogle Scholar
  12. Davis M, Ideo G, Harrison NG, Williams R (1975) Hepatic glutathione depletion and impaired bromosulphthalein clearance early after paracetamol overdose in man and the rat. Clin Sci Mol Med 49:495–502PubMedGoogle Scholar
  13. Gunawan BK, Liu ZX, Han D, Hanawa N, Gaarde WA, Kaplowitz N (2006) c-Jun N-terminal kinase plays a major role in murine acetaminophen hepatotoxicity. Gastroenterology 131:165–178PubMedCrossRefGoogle Scholar
  14. Hanawa N, Shinohara M, Saberi B, Gaarde WA, Han D, Kaplowitz N (2008) Role of JNK translocation to mitochondria leading to inhibition of mitochondria bioenergetics in acetaminophen-induced liver injury. J Biol Chem 283:13565–13577PubMedCrossRefGoogle Scholar
  15. Jaeschke H (1990) Glutathione disulfide formation and oxidant stress during acetaminophen-induced hepatotoxicity in mice in vivo: the protective effect of allopurinol. J Pharmacol Exp Ther 255:935–941PubMedGoogle Scholar
  16. Jaeschke H, McGill MR (2013) Serum glutamate dehydrogenase—biomarker for liver cell death or mitochondrial dysfunction? Toxicol Sci 134:221–222PubMedCrossRefGoogle Scholar
  17. Jaeschke H, McGill MR, Ramachandran A (2012) Oxidant stress, mitochondria, and cell death mechanisms in drug-induced liver injury: lessons learned from acetaminophen hepatotoxicity. Drug Metab Rev 44:88–106PubMedCrossRefGoogle Scholar
  18. Kaufman DW, Kelly JP, Rosenberg L, Anderson TE, Mitchell AA (2002) Recent patterns of medication use in the ambulatory adult population of the United States: the Slone survey. JAMA 287:337–344PubMedCrossRefGoogle Scholar
  19. Knight TR, Ho YS, Farhood A, Jaeschke H (2002) Peroxynitrite is a critical mediator of acetaminophen hepatotoxicity in murine livers: protection by glutathione. J Pharmacol Exp Ther 303:468–475PubMedCrossRefGoogle Scholar
  20. Kon K, Kim JS, Jaeschke H, Lemasters JJ (2004) Mitochondrial permeability transition in acetaminophen-induced necrosis and apoptosis of cultured mouse hepatocytes. Hepatology 40:1170–1179PubMedCrossRefGoogle Scholar
  21. Lauterburg BH, Mitchell JR (1987) Therapeutic doses of acetaminophen stimulate turnover of cysteine and glutathione in man. J Hepatol 4:206–211PubMedCrossRefGoogle Scholar
  22. McGill MR, Jaeschke H (2013) Metabolism and disposition of acetaminophen: recent advances in relation to hepatotoxicity and diagnosis. Pharm Res 30:2174–2187Google Scholar
  23. McGill MR, Yan HM, Ramachandran A, Murray GJ, Rollins DE, Jaeschke H (2011) HepaRG cells: a human model to study mechanisms of acetaminophen hepatotoxicity. Hepatology 53:974–982PubMedCentralPubMedCrossRefGoogle Scholar
  24. McGill MR, Sharpe MR, Williams CD, Taha M, Curry SC, Jaeschke H (2012) The mechanism underlying acetaminophen-induced hepatotoxicity in humans and mice involves mitochondrial damage and nuclear DNA fragmentation. J Clin Invest 122:1574–1583PubMedCentralPubMedCrossRefGoogle Scholar
  25. McGill MR, Lebofsky M, Norris HK, Slawson MH, Bajt ML, Xie Y, Williams CD, Wilkins DG, Rollins DE, Jaeschke H (2013) Plasma and liver acetaminophen-protein adduct levels in mice after acetaminophen treatment: dose-response, mechanisms, and clinical implications. Toxicol Appl Pharmacol 269:240–249PubMedCrossRefGoogle Scholar
  26. Mitchell JR, Jollow DJ, Potter WZ, Gillette JR, Brodie BB (1973) Acetaminophen-induced hepatic necrosis. IV. Protective role of glutathione. J Pharmacol Exp Ther 187:211–217PubMedGoogle Scholar
  27. Nakagawa H, Maeda S, Hikiba Y, Ohmae T, Shibata W, Yanai A, Sakamoto K, Ogura K, Noguchi T, Karin M, Ichijo H, Omata M (2008) Deletion of apoptosis signal-regulating kinase 1 attenuates acetaminophen-induced liver injury by inhibition c-Jun N-terminal kinase activation. Gastroenterology 135:1311–1321PubMedCrossRefGoogle Scholar
  28. Ramachandran A, Lebofsky M, Weinman SA, Jaeschke H (2011) The impact of partial manganese superoxide dismutase (SOD2)-deficiency on mitochondrial oxidant stress, DNA fragmentation and liver injury during acetaminophen hepatotoxicity. Toxicol Appl Pharmacol 251:226–233PubMedCentralPubMedCrossRefGoogle Scholar
  29. Rinaldo P, Matern D, Bennett MJ (2002) Fatty acid oxidation disorders. Annu Rev Physiol 64:477–502PubMedCrossRefGoogle Scholar
  30. Saito C, Lemasters JJ, Jaeschke H (2010a) c-Jun N-terminal kinase modulates oxidant stress and peroxynitrite formation independent of inducible nitric oxide synthase in acetaminophen hepatotoxicity. Toxicol Appl Pharmacol 246:8–17PubMedCentralPubMedCrossRefGoogle Scholar
  31. Saito C, Zwingmann C, Jaeschke H (2010b) Novel mechanisms of protection against acetaminophen hepatotoxicity in mice by glutathione and N-acetylcysteine. Hepatology 51:246–254PubMedCentralPubMedCrossRefGoogle Scholar
  32. Sharma M, Gadang V, Jaeschke A (2012) Critical role for mixed-lineage kinase 3 in acetaminophen-induced hepatotoxicity. Mol Pharmacol 82:1001–1007PubMedCrossRefGoogle Scholar
  33. Tirmenstein MA, Nelson SD (1989) Subcellular binding and effects on calcium homeostasis produced by acetaminophen and a nonhepatotoxic regioisomer, 3′-hydroxyacetanilide, in mouse liver. J Biol Chem 264:9814–9819PubMedGoogle Scholar
  34. Whitehouse LW, Wong LT, Solomonraj G, Paul CJ, Thomas BH (1981) N-acetylcysteine-induced inhibition of gastric emptying: a mechanism affording protection to mice from the hepatotoxicity of concomitantly administered acetaminophen. Toxicology 19:113–125PubMedCrossRefGoogle Scholar
  35. Win S, Than TA, Han D, Petrovic LM, Kaplowitz N (2011) c-Jun N-terminal kinase (JNK)-dependent acute liver injury from acetaminophen or tumor necrosis factor (TNF) requires mitochondrial Sab protein expression in mice. J Biol Chem 286:35071–35078PubMedCrossRefGoogle Scholar
  36. Wong SG, Card JW, Racz WJ (2000) The role of mitochondrial injury in bromobenzene and furosemide induced hepatotoxicity. Toxicol Lett 116:171–181PubMedCrossRefGoogle Scholar

Copyright information

© Springer-Verlag Berlin Heidelberg 2013

Authors and Affiliations

  • Mitchell R. McGill
    • 1
  • Feng Li
    • 1
  • Matthew R. Sharpe
    • 2
  • C. David Williams
    • 1
  • Steven C. Curry
    • 3
    • 4
  • Xiaochao Ma
    • 1
  • Hartmut Jaeschke
    • 1
    Email author
  1. 1.Department of Pharmacology, Toxicology, and TherapeuticsUniversity of Kansas Medical CenterKansas CityUSA
  2. 2.Department of Internal MedicineUniversity of Kansas HospitalKansas CityUSA
  3. 3.Department of Medical ToxicologyBanner Good Samaritan Medical CenterPhoenixUSA
  4. 4.Department of Medicine, Center for Toxicology and Pharmacology Education and ResearchUniversity of Arizona College of MedicinePhoenixUSA

Personalised recommendations