Guest editorial
Highlight report: Necrosis-apoptosis conundrum of hepatocytes: mode of hepatocyte death after acetaminophen intoxication
Ahmed Ghallab1
1Forensic Medicine and Toxicology Department, Faculty of Veterinary Medicine, South Valley University, Qena, Egypt
EXCLI J 2018;17:Doc1191
Recently, Huo Du from Hartmut Jaesches's group at University of Kansas published an outstanding study about the mode of hepatocyte killing by acetaminophen and demonstrate that the typical necrotic cell death may switch to secondary apoptosis after specific interventions (Du et al., 2018[7]).
Acetaminophen (APAP) is responsible for more than 70.000 hospitalizations per year and approximately 50 % of acute liver failure cases (Budnitz et al., 2011[5]; Manthripragada et al., 2011[25]). Cell killing by APAP is a consequence of cytochrome P450-mediated formation of the reactive N-acetyl-p-benzoquinone imine (NAPQI) that binds to proteins and glutathione (Mc Gill and Jaeschke, 2013[27], 2015[26]; Nelson, 1989[29]; Xie et al., 2015[38]). It is well accepted that APAP kills hepatocytes predominantly by necrosis rather than by apoptosis (Bajt et al., 2004[3]; Gujral et al., 2002[16]; Jaeschke et al., 2011[21]; McGill et al., 2011[28]). This phenomenon has been observed in vivo and in vitro. The preference for necrosis remains difficult to understand, because some mechanisms considered specific for apoptosis are still induced by APAP, such as mitochondrial translocation of bax and release of cytochrome C, but nevertheless do not lead to apoptotic phenotype (Du et al. 2016[6]; Adams et al., 2001[1]; Knight and Jaeschke, 2002[23]; Bajt et al., 2008[2]).
In their present study, Du and colleagues came much closer to an explanation of this conundrum (Du et al., 2018[7]). They used the mitochondria-targeted superoxide dismutase mimetic Mito-tempo in mice intoxicated with a hepatotoxic dose of 300 mg/kg APAP. As expected, Mito-tempo reduced APAP induced necrosis and led to an overall protection against hepatotoxicity. However, some hepatocytes switched to a clearly apoptotic phenotype as evidenced by morphology, TUNEL positivity and caspase activation, which was not observed after APAP intoxication without Mito-tempo administration (Du et al., 2018[7]). In an elegant series of experiments using RIP3 knockout mice and decreasing RIP3 protein levels by a RIP3-morpholino, the authors demonstrate that the effect of Mito-tempo is due to inhibition of RIP3. In conclusion, the necrosis-apoptosis conundrum of hepatocytes is due to RIP3 kinase, which tips the balance to necrosis, while its inhibition switches cell death to necrosis.
Currently, hepatotoxicity is a major research focus, because drug induced liver injury represents a frequent cause of drug withdrawal from the market (Godoy et al., 2013[11], 2016[12]; Hewitt et al., 2007[20]; Reif et al., 2017[30]).
Numerous studies aim at a better understanding of the molecular and pathophysiological mechanisms of hepatotoxicity (Hassan, 2016[19]; Stöber, 2016[35]; Sezgin et al., 2018[33]; Jansen et al., 2017[22]; Vartak et al., 2016[37]; Ghallab et al., 2016[10]; Bolt, 2017[4]; Schenk et al., 2017[32]; Thiel et al., 2015[36]; Hammad et al., 2014[18]). A frequently applied strategy in toxicology is to construct 'adverse outcome pathways' (Leist et al., 2017[24]; Rodrigues et al., 2018[31]) aiming for possibilities to study hepatoxicity in vitro and in silico (Ghallab, 2017[9]; Hammad, 2013[17]; Grinberg et al., 2014[14], 2018[13]; Gu et al., 2018[15]; Shinde et al., 2015[34]; Frey et al., 2014[8]).
However, this strategy is hampered by the fact that so many aspects of hepatotoxicity in vivo remain elusive, even for a compound as intensively studied as APAP. In conclusion, Huo Du and colleagues are to be congratulated that they unraveled a mystery that confused toxicologists since decades.
References
1.
Adams ML, Pierce RH, Vail ME, White CC, Tonge RP, Kavanagh TJ, et al. Enhanced acetaminophen hepatotoxicity in transgenic mice overexpressing BCL-2. Mol Pharmacol. 2001;60:907-15.2.
Bajt ML, Farhood A, Lemasters JJ, Jaeschke H. Mitochondrial bax translocation accelerates DNA fragmentation and cell necrosis in a murine model of acetaminophen hepatotoxicity. J Pharmacol Exp Ther. 2008;324:8-14.3.
Bajt ML, Knight TR, Lemasters JJ, Jaeschke H. Acetaminophen-induced oxidant stress and cell injury in cultured mouse hepatocytes: protection by N-acetyl cysteine. Toxicol Sci. 2004;80:343-9.4.
Bolt HM. Highlight report: The pseudolobule in liver fibrosis. EXCLI J. 2017;16:1321-2.5.
Budnitz DS, Lovegrove MC, Crosby AE. Emergency department visits for overdoses of acetaminophen-containing products. Am J Prev Med. 2011;40:585-92.6.
Du K, Ramachandran A, Weemhoff JL, Chavan H, Xie Y, Krishnamurthy P, et al. Editor's highlight: Metformin protects against acetaminophen hepatotoxicity by attenuation of mitochondrial oxidant stress and dysfunction. Toxicol Sci. 2016;154:214-26.7.
Du K, Ramachandran A, Weemhoff JL, Woolbright BL, Jaeschke AH, Chao X, et al. Mito-tempo protects against acute liver injury but induces limited secondary apoptosis during the late phase of acetaminophen hepatotoxicity. Arch Toxicol. 2018 Oct 15, epub ahead of print.8.
Frey O, Misun PM, Fluri DA, Hengstler JG, Hierlemann A. Reconfigurable microfluidic hanging drop network for multi-tissue interaction and analysis. Nat Commun. 2014;5:4250.9.
Ghallab A. Highlight report: Metabolomics in hepatotoxicity testing. EXCLI J. 2017;16:1323-5.10.
Ghallab A, Cellière G, Henkel SG, Driesch D, Hoehme S, Hofmann U, et al. Model-guided identification of a therapeutic strategy to reduce hyperammonemia in liver diseases. J Hepatol. 2016;64:860-71.11.
Godoy P, Hewitt NJ, Albrecht U, Andersen ME, Ansari N, Bhattacharya S, et al. Recent advances in 2D and 3D in vitro systems using primary hepatocytes, alternative hepatocyte sources and non-parenchymal liver cells and their use in investigating mechanisms of hepatotoxicity, cell signaling and ADME. Arch Toxicol. 2013;87:1315-530. 12.
Godoy P, Widera A, Schmidt-Heck W, Campos G, Meyer C, Cadenas C, et al. Gene network activity in cultivated primary hepatocytes is highly similar to diseased mammalian liver tissue. Arch Toxicol. 2016;90:2513-29.13.
Grinberg M, Stöber RM, Albrecht W, Edlund K, Schug M, Godoy P, et al. Toxicogenomics directory of rat hepatotoxicants in vivo and in cultivated hepatocytes. Arch Toxicol. 2018;92:3517-33.14.
Grinberg M, Stöber RM, Edlund K, Rempel E, Godoy P, Reif R, et al. Toxicogenomics directory of chemically exposed human hepatocytes. Arch Toxicol. 2014;88:2261-87.15.
Gu X, Albrecht W, Edlund K, Kappenberg F, Rahnenführer J, Leist M, et al. Relevance of the incubation period in cytotoxicity testing with primary human hepatocytes. Arch Toxicol. 2018;92:3505-15.16.
Gujral JS, Knight TR, Farhood A, Bajt ML, Jaeschke H. Mode of cell death after acetaminophen overdose in mice: apoptosis or oncotic necrosis? Toxicol Sci. 2002;67:322-8.17.
Hammad S. Advances in 2D and 3D in vitro systems for hepatotoxicity testing. EXCLI J. 2013;12:993-6. 18.
Hammad S, Hoehme S, Friebel A, von Recklinghausen I, Othman A, Begher-Tibbe B, et al. Protocols for staining of bile canalicular and sinusoidal networks of human, mouse and pig livers, three-dimensional reconstruction and quantification of tissue microarchitecture by image processing and analysis. Arch Toxicol. 2014;88:1161-83.19.
Hassan R. Possibilities and limitations of intravital imaging. EXCLI J. 2016;15:872-4.20.
Hewitt NJ, Lechón MJ, Houston JB, Hallifax D, Brown HS, Maurel P, et al. Primary hepatocytes: current understanding of the regulation of metabolic enzymes and transporter proteins, and pharmaceutical practice for the use of hepatocytes in metabolism, enzyme induction, transporter, clearance, and hepatotoxicity studies. Drug Metab Rev. 2007;39:159-234.21.
Jaeschke H, Williams CD, Farhood A. No evidence for caspase-dependent apoptosis in acetaminophen hepatotoxicity. Hepatology. 2011;53:718-9.22.
Jansen PL, Ghallab A, Vartak N, Reif R, Schaap FG, Hampe J, et al. The ascending pathophysiology of cholestatic liver disease. Hepatology. 2017;65:722-38.23.
Knight TR, Jaeschke H. Acetaminophen-induced inhibition of Fas receptor-mediated liver cell apoptosis: mitochondrial dysfunction versus glutathione depletion. Toxicol Appl Pharmacol. 2002;181:133-41.24.
Leist M, Ghallab A, Graepel R, Marchan R, Hassan R, Bennekou SH, et al. Adverse outcome pathways: opportunities, limitations and open questions. Arch Toxicol. 2017;91:3477-505.25.
Manthripragada AD, Zhou EH, Budnitz DS, Lovegrove MC, Willy ME. Characterization of acetaminophen overdose-related emergency department visits and hospitalizations in the United States. Pharmacoepidemiol Drug Saf. 2011;20:819-26.26.
McGill MR, Jaeschke H. A direct comparison of methods used to measure oxidized glutathione in biological samples: 2-vinylpyridine and N-ethylmaleimide. Toxicol Mech Methods. 2015;25:589-95.27.
McGill MR, Jaeschke H. Metabolism and disposition of acetaminophen: recent advances in relation to hepatotoxicity and diagnosis. Pharm Res. 2013;30:2174-87.28.
McGill MR, Yan HM, Ramachandran A, Murray GJ, Rollins DE, Jaeschke H. HepaRG cells: a human model to study mechanisms of acetaminophen hepatotoxicity. Hepatology. 2011;53:974-82.29.
Nelson SD, Lynch JJ, Sanders D, Montgomery DG, Lucchesi BR. Electrophysiologic actions and antifibrillatory efficacy of subacute left stellectomy in a conscious, post-infarction canine model of ischemic ventricular fibrillation. Int J Cardiol. 1989;22:365-76.30.
Reif R, Ghallab A, Beattie L, Günther G, Kuepfer L, Kaye PM, et al. In vivo imaging of systemic transport and elimination of xenobiotics and endogenous molecules in mice. Arch Toxicol. 2017;91:1335-52.31.
Rodrigues RM, Kollipara L, Chaudhari U, Sachinidis A, Zahedi RP, Sickmann A, et al. Omics-based responses induced by bosentan in human hepatoma HepaRG cell cultures. Arch Toxicol. 2018;92:1939-52.32.
Schenk A, Ghallab A, Hofmann U, Hassan R, Schwarz M, et al. Physiologically-based modelling in mice suggests an aggravated loss of clearance capacity after toxic liver damage. Sci Rep. 2017;7(1):6224.33.
Sezgin S, Hassan R, Zühlke S, Kuepfer L, Hengstler JG, Spiteller M, et al. Spatio-temporal visualization of the distribution of acetaminophen as well as its metabolites and adducts in mouse livers by MALDI MSI. Arch Toxicol. 2018;92:2963-77.34.
Shinde V, Stöber R, Nemade H, Sotiriadou I, Hescheler J, Hengstler J, et al. Transcriptomics of hepatocytes treated with toxicants for investigating molecular mechanisms underlying hepatotoxicity. Methods Mol Biol. 2015;1250:225-40. 35.
Stöber R. Pathophysiology of cholestatic liver disease and its relevance for in vitro tests of hepatotoxicity. EXCLI J. 2016;15:870-1.36.
Thiel C, Schneckener S, Krauss M, Ghallab A, Hofmann U, Kanacher T, et al. A systematic evaluation of the use of physiologically based pharmacokinetic modeling for cross-species extrapolation. J Pharm Sci. 2015;104:191-206.37.
Vartak N, Damle-Vartak A, Richter B, Dirsch O, Dahmen U, Hammad S, et al. Cholestasis-induced adaptive remodeling of interlobular bile ducts. Hepatology. 2016;63:951-64.38.
Xie Y, McGill MR, Du K, Dorko K, Kumer SC, Schmitt TM, et al. Mitochondrial protein adducts formation and mitochondrial dysfunction during N-acetyl-m-aminophenol (AMAP)-induced hepatotoxicity in primary human hepatocytes. Toxicol Appl Pharmacol. 2015;289:213-22.