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Sodium 4-phenylbutyric acid prevents murine acetaminophen hepatotoxicity by minimizing endoplasmic reticulum stress

  • Original Article—Liver, Pancreas, and Biliary Tract
  • Published:
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Abstract

Background

Acetaminophen (APAP) overdose induces severe oxidative stress followed by hepatocyte apoptosis/necrosis. Previous studies have indicated that endoplasmic reticulum (ER) stress is involved in the cell death process. Therefore, we investigated the effect of the chemical chaperone 4-phenyl butyric acid (PBA) on APAP-induced liver injury in mice.

Methods

Eight-week-old male C57Bl6/J mice were given a single intraperitoneal (i.p.) injection of APAP (450 mg/kg body weight), following which some were repeatedly injected with PBA (120 mg/kg body weight, i.p.) every 3 h starting at 0.5 h after the APAP challenge. All mice were then serially euthanized up to 12 h later.

Results

PBA treatment dramatically ameliorated the massive hepatocyte apoptosis/necrosis that was observed 6 h after APAP administration. PBA also significantly prevented the APAP-induced increases in cleaved activating transcription factor 6 and phosphorylation of c-Jun N-terminal protein kinase and significantly blunted the increases in mRNA levels for binding immunoglobulin protein, spliced X-box binding protein-1, and C/EBP homologous protein. Moreover, PBA significantly prevented APAP-induced Bax translocation to the mitochondria, and the expression of heme oxygenase-1 mRNA and 4-hydroxynonenal. By contrast, PBA did not affect hepatic glutathione depletion following APAP administration, reflecting APAP metabolism.

Conclusions

PBA prevents APAP-induced liver injury even when an APAP challenge precedes its administration. The underlying mechanism of action most likely involves the prevention of ER stress-induced apoptosis/necrosis in the hepatocytes during APAP intoxication.

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Abbreviations

ALT:

Alanine aminotransferase

AST:

Aspartate aminotransferase

ANOVA:

Analysis of variance

eIF2α:

Eukaryotic initiation factor 2α

ER:

Endoplasmic reticulum

GAPDH:

Glyceraldehyde 3-phosphate dehydrogenase

GRP78:

Glucose-regulated protein 78

Bip:

Binding immunoglobulin protein

H–E:

Hematoxylin–Eosin

4-HNE:

4-hydroxy-2-nonenal

JNK:

c-Jun N-terminal kinase

HO1:

Heme oxygenase-1

NASH:

Non-alcoholic steatohepatitis

sXBP1:

Spliced X-box binding protein-1

UPR:

Unfolded protein response

NAPQI:

N-acetyl-p-benzoquinone imine

TUNEL:

Terminal deoxynucleotidyl transferase-mediated UTP end labeling

ATF6:

Activating transcription factor 6

IRE1α:

Inositol-requiring kinase 1α

PERK:

Protein kinase RNA-like ER kinase

HSP60:

Heat shock protein 60

CHOP:

C/EBP homologous protein

MPT:

Mitochondrial permeability transition

ROS:

Reactive oxygen species

PBA:

4-phenyl butyric acid

UPR:

Unfolded protein response

References

  1. Brune K, Renner B, Tiegs G. Acetaminophen/paracetamol: a history of errors, failures and false decisions. Eur J Pain. 2015;19:953–65.

    Article  CAS  PubMed  Google Scholar 

  2. Major JM, Zhou EH, Wong HL, et al. Trends in rates of acetaminophen-related adverse events in the United States. Pharmacoepidemiol Drug Saf. 2016;25(5):590–8.

    Article  PubMed  Google Scholar 

  3. Canbay A, Jochum C, Bechmann LP, et al. Acute liver failure in a metropolitan area in Germany: a retrospective study (2002–2008). Z Gastroenterol. 2009;47:807–13.

    Article  CAS  PubMed  Google Scholar 

  4. Krenzelok EP, The FDA. Acetaminophen advisory committee meeting—what is the future of acetaminophen in the United States? The perspective of a committee member. Clin Toxicol (Phila). 2009;47:784–9.

    Article  CAS  Google Scholar 

  5. Xu C, Bailly-Maitre B, Reed JC. Endoplasmic reticulum stress: cell life and death decisions. J Clin Invest. 2005;115:2656–64.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  6. Ferner RE, Dear JW, Bateman DN. Management of paracetamol poisoning. BMJ. 2011;342:d2218.

    Article  PubMed  Google Scholar 

  7. Zyoud SH, Al-Jabi SW, Sweileh WM, et al. Global research productivity of N-acetylcysteine use in paracetamol overdose: a bibliometric analysis (1976–2012). Hum Exp Toxicol. 2015;34:1006–16.

    Article  CAS  PubMed  Google Scholar 

  8. Sudo C, Maekawa K, Segawa K, et al. Trends in drug-induced liver injury based on reports of adverse reactions to PMDA in Japan. Kokuritsu Iyakuhin Shokuhin Eisei Kenkyusho Hokoku. 2012;130:66–70.

    Google Scholar 

  9. Chiew AL, Isbister GK, Duffull SB, et al. Evidence for the changing regimens of acetylcysteine. Br J Clin Pharmacol. 2016;81:471–81.

    Article  CAS  PubMed  Google Scholar 

  10. Doyle KM, Kennedy D, Gorman AM, et al. Unfolded proteins and endoplasmic reticulum stress in neurodegenerative disorders. J Cell Mol Med. 2011;15:2025–39.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  11. Sano R, Reed JC. ER stress-induced cell death mechanisms. Biochim Biophys Acta. 2013;1833:3460–70.

    Article  CAS  PubMed  Google Scholar 

  12. Sasaki M, Yoshimura-Miyakoshi M, Sato Y, et al. A possible involvement of endoplasmic reticulum stress in biliary epithelial autophagy and senescence in primary biliary cirrhosis. J Gastroenterol. 2015;50:984–95.

    Article  PubMed  Google Scholar 

  13. Du RH, Tan J, Yan N, et al. Kir6.2 knockout aggravates lipopolysaccharide-induced mouse liver injury via enhancing NLRP3 inflammasome activation. J Gastroenterol. 2014;49:727–36.

    Article  CAS  PubMed  Google Scholar 

  14. Hamano M, Ezaki H, Kiso S, et al. Lipid overloading during liver regeneration causes delayed hepatocyte DNA replication by increasing ER stress in mice with simple hepatic steatosis. J Gastroenterol. 2014;49:305–16.

    Article  CAS  PubMed  Google Scholar 

  15. Narabayashi K, Ito Y, Eid N, et al. Indomethacin suppresses LAMP-2 expression and induces lipophagy and lipoapoptosis in rat enterocytes via the ER stress pathway. J Gastroenterol. 2015;50:541–54.

    Article  CAS  PubMed  Google Scholar 

  16. Prentice H, Modi JP, Wu JY. Mechanisms of neuronal protection against excitotoxicity, endoplasmic reticulum stress, and mitochondrial dysfunction in stroke and neurodegenerative diseases. Oxid Med Cell Longev. 2015;2015:964518.

    Article  PubMed  PubMed Central  Google Scholar 

  17. Salvado L, Palomer X, Barroso E, et al. Targeting endoplasmic reticulum stress in insulin resistance. Trends Endocrinol Metab. 2015;26:438–48.

    Article  CAS  PubMed  Google Scholar 

  18. Zhang Q, Li Y, Liang T, et al. ER stress and autophagy dysfunction contribute to fatty liver in diabetic mice. Int J Biol Sci. 2015;11:559–68.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  19. Hrincius ER, Liedmann S, Finkelstein D, et al. Acute lung injury results from innate sensing of viruses by an ER stress pathway. Cell Rep. 2015;11:1591–603.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  20. Asano T, Sato A, Isono M, et al. Bortezomib and belinostat inhibit renal cancer growth synergistically by causing ubiquitinated protein accumulation and endoplasmic reticulum stress. Biomed Rep. 2015;3:797–801.

    PubMed  PubMed Central  Google Scholar 

  21. He J, Du L, Bao M, et al. Oroxin A inhibits breast cancer cell growth by inducing robust endoplasmic reticulum stress and senescence. Anticancer Drugs. 2016;27:204–15.

    Article  CAS  PubMed  Google Scholar 

  22. Zhang T, Kho DH, Wang Y, et al. Gp78, an E3 ubiquitin ligase acts as a gatekeeper suppressing nonalcoholic steatohepatitis (NASH) and liver cancer. PLoS One. 2015;10:e0118448.

    Article  PubMed  PubMed Central  Google Scholar 

  23. Uzi D, Barda L, Scaiewicz V, et al. CHOP is a critical regulator of acetaminophen-induced hepatotoxicity. J Hepatol. 2013;59:495–503.

    Article  CAS  PubMed  Google Scholar 

  24. Nagy G, Kardon T, Wunderlich L, et al. Acetaminophen induces ER dependent signaling in mouse liver. Arch Biochem Biophys. 2007;459:273–9.

    Article  CAS  PubMed  Google Scholar 

  25. Song YF, Luo Z, Zhang LH, et al. Endoplasmic reticulum stress and disturbed calcium homeostasis are involved in copper-induced alteration in hepatic lipid metabolism in yellow catfish Pelteobagrus fulvidraco. Chemosphere. 2015;144:2443–53.

    Article  PubMed  Google Scholar 

  26. Kolb PS, Ayaub EA, Zhou W, et al. The therapeutic effects of 4-phenylbutyric acid in maintaining proteostasis. Int J Biochem Cell Biol. 2015;61:45–52.

    Article  CAS  PubMed  Google Scholar 

  27. Vilatoba M, Eckstein C, Bilbao G, et al. Sodium 4-phenylbutyrate protects against liver ischemia reperfusion injury by inhibition of endoplasmic reticulum-stress mediated apoptosis. Surgery. 2005;138:342–51.

    Article  PubMed  Google Scholar 

  28. Lichter-Konecki U, Diaz GA, Merritt JL 2nd, et al. Ammonia control in children with urea cycle disorders (UCDs); phase 2 comparison of sodium phenylbutyrate and glycerol phenylbutyrate. Mol Genet Metab. 2011;103:323–9.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  29. Odievre MH, Brun M, Krishnamoorthy R, et al. Sodium phenyl butyrate downregulates endothelin-1 expression in cultured human endothelial cells: relevance to sickle-cell disease. Am J Hematol. 2007;82:357–62.

    Article  CAS  PubMed  Google Scholar 

  30. Collins AF, Pearson HA, Giardina P, et al. Oral sodium phenylbutyrate therapy in homozygous beta thalassemia: a clinical trial. Blood. 1995;85:43–9.

    CAS  PubMed  Google Scholar 

  31. Morinaga M, Kon K, Saito H, et al. Sodium 4-phenylbutyrate prevents murine dietary steatohepatitis caused by trans-fatty acid plus fructose. J Clin Biochem Nutr. 2015;57:183–91.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  32. Kon K, Ikejima K, Okumura K, et al. Diabetic KK-A(y) mice are highly susceptible to oxidative hepatocellular damage induced by acetaminophen. Am J Physiol Gastrointest Liver Physiol. 2010;299:G329–37.

    Article  CAS  PubMed  Google Scholar 

  33. Jo H, Choe SS, Shin KC, et al. Endoplasmic reticulum stress induces hepatic steatosis via increased expression of the hepatic very low-density lipoprotein receptor. Hepatology. 2013;57:1366–77.

    Article  CAS  PubMed  Google Scholar 

  34. Harding HP, Zhang Y, Ron D. Protein translation and folding are coupled by an endoplasmic-reticulum-resident kinase. Nature. 1999;397:271–4.

    Article  CAS  PubMed  Google Scholar 

  35. Ma Y, Hendershot LM. Delineation of a negative feedback regulatory loop that controls protein translation during endoplasmic reticulum stress. J Biol Chem. 2003;278:34864–73.

    Article  CAS  PubMed  Google Scholar 

  36. Shen J, Chen X, Hendershot L, et al. ER stress regulation of ATF6 localization by dissociation of BiP/GRP78 binding and unmasking of Golgi localization signals. Dev Cell. 2002;3:99–111.

    Article  CAS  PubMed  Google Scholar 

  37. Yoshida H, Okada T, Haze K, et al. ATF6 activated by proteolysis binds in the presence of NF-Y (CBF) directly to the cis-acting element responsible for the mammalian unfolded protein response. Mol Cell Biol. 2000;20:6755–67.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  38. Fu HY, Minamino T, Tsukamoto O, et al. Overexpression of endoplasmic reticulum-resident chaperone attenuates cardiomyocyte death induced by proteasome inhibition. Cardiovasc Res. 2008;79:600–10.

    Article  CAS  PubMed  Google Scholar 

  39. Yoshida H, Matsui T, Yamamoto A, et al. XBP1 mRNA is induced by ATF6 and spliced by IRE1 in response to ER stress to produce a highly active transcription factor. Cell. 2001;107:881–91.

    Article  CAS  PubMed  Google Scholar 

  40. Du K, Xie Y, McGill MR, et al. Pathophysiological significance of c-jun N-terminal kinase in acetaminophen hepatotoxicity. Expert Opin Drug Metab Toxicol. 2015;11:1769–79.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  41. Saito C, Lemasters JJ, Jaeschke H. c-Jun N-terminal kinase modulates oxidant stress and peroxynitrite formation independent of inducible nitric oxide synthase in acetaminophen hepatotoxicity. Toxicol Appl Pharmacol. 2010;246:8–17.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  42. McCullough KD, Martindale JL, Klotz LO, et al. Gadd153 sensitizes cells to endoplasmic reticulum stress by down-regulating Bcl2 and perturbing the cellular redox state. Mol Cell Biol. 2001;21:1249–59.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  43. Puthalakath H, O’Reilly LA, Gunn P, et al. ER stress triggers apoptosis by activating BH3-only protein Bim. Cell. 2007;129:1337–49.

    Article  CAS  PubMed  Google Scholar 

  44. Lee CH, Kuo CY, Wang CJ, et al. A polyphenol extract of Hibiscus sabdariffa L. ameliorates acetaminophen-induced hepatic steatosis by attenuating the mitochondrial dysfunction in vivo and in vitro. Biosci Biotechnol Biochem. 2012;76:646–51.

    Article  CAS  PubMed  Google Scholar 

  45. Bulku E, Stohs SJ, Cicero L, et al. Curcumin exposure modulates multiple pro-apoptotic and anti-apoptotic signaling pathways to antagonize acetaminophen-induced toxicity. Curr Neurovasc Res. 2012;9:58–71.

    Article  CAS  PubMed  Google Scholar 

  46. Xu C, Xu W, Palmer AE, et al. BI-1 regulates endoplasmic reticulum Ca2+ homeostasis downstream of Bcl-2 family proteins. J Biol Chem. 2008;283:11477–84.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  47. Kon K, Kim JS, Uchiyama A, et al. Lysosomal iron mobilization and induction of the mitochondrial permeability transition in acetaminophen-induced toxicity to mouse hepatocytes. Toxicol Sci. 2010;117:101–8.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  48. Uchiyama A, Kim JS, Kon K, et al. Translocation of iron from lysosomes into mitochondria is a key event during oxidative stress-induced hepatocellular injury. Hepatology. 2008;48:1644–54.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  49. Simmen T, Lynes EM, Gesson K, et al. Oxidative protein folding in the endoplasmic reticulum: tight links to the mitochondria-associated membrane (MAM). Biochim Biophys Acta. 2010;1798:1465–73.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  50. Salem IB, Prola A, Boussabbeh M, et al. Activation of ER stress and apoptosis by alpha- and beta-Zearalenol in HCT116 cells, protective role of Quercetin. Neurotoxicology. 2016;53:334–42.

    Article  PubMed  Google Scholar 

  51. Zhong D, Wang H, Liu M, et al. Ganoderma lucidum polysaccharide peptide prevents renal ischemia reperfusion injury via counteracting oxidative stress. Sci Rep. 2015;5:16910.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  52. Egnatchik RA, Leamy AK, Jacobson DA, et al. ER calcium release promotes mitochondrial dysfunction and hepatic cell lipotoxicity in response to palmitate overload. Mol Metab. 2014;3:544–53.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  53. Xie Y, McGill MR, Dorko K, et al. Mechanisms of acetaminophen-induced cell death in primary human hepatocytes. Toxicol Appl Pharmacol. 2014;279:266–74.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  54. Larson AM. Acetaminophen hepatotoxicity. Clin Liver Dis. 2007;11:525–48 (vi).

    Article  PubMed  Google Scholar 

  55. Lauterburg BH, Corcoran GB, Mitchell JR. Mechanism of action of N-acetylcysteine in the protection against the hepatotoxicity of acetaminophen in rats in vivo. J Clin Invest. 1983;71:980–91.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  56. Gilbert J, Baker SD, Bowling MK, et al. A phase I dose escalation and bioavailability study of oral sodium phenylbutyrate in patients with refractory solid tumor malignancies. Clin Cancer Res. 2001;7:2292–300.

    CAS  PubMed  Google Scholar 

  57. Shimizu D, Ishitsuka Y, Miyata K, et al. Protection afforded by pre- or post-treatment with 4-phenylbutyrate against liver injury induced by acetaminophen overdose in mice. Pharmacol Res. 2014;87:26–41.

    Article  CAS  PubMed  Google Scholar 

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Acknowledgments

The authors thank Takako Ikegami and Tomomi Ikeda (Laboratory of Molecular and Biochemical Research, Research Support Center, Juntendo University Graduate School of Medicine, Tokyo, Japan) and Maiko Suzuki and Ryota Kitagawa (Department of Gastroenterology, Juntendo University School of Medicine, Tokyo, Japan) for technical assistance and R. F. Whittier for critical reading of the manuscript and thoughtful discussions.

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Authors and Affiliations

  1. Department of Gastroenterology, Juntendo University School of Medicine, 2-1-1 Hongo, Bunkyo-ku, Tokyo, 113-8421, Japan

    Hiromi Kusama, Kazuyoshi Kon, Kenichi Ikejima, Kumiko Arai, Tomonori Aoyama, Akira Uchiyama, Shunhei Yamashina & Sumio Watanabe

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  1. Hiromi Kusama
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  2. Kazuyoshi Kon
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Correspondence to Kazuyoshi Kon.

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The authors declare that they have no conflict of interest.

Financial support

This work was supported in part by JSPS KAKENHI (24590995 to K.K., 24590996 to K.I., 24390191 to S.W.).

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Kusama, H., Kon, K., Ikejima, K. et al. Sodium 4-phenylbutyric acid prevents murine acetaminophen hepatotoxicity by minimizing endoplasmic reticulum stress. J Gastroenterol 52, 611–622 (2017). https://doi.org/10.1007/s00535-016-1256-3

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