title: Isoniazid-induced cell death is precipitated by underlying mitochondrial complex I dysfunction in mouse hepatocytes.
abstract: Isoniazid (INH) is an antituberculosis drug that has been associated with idiosyncratic liver injury in susceptible patients. The underlying mechanisms are still unclear, but there is growing evidence that INH and/or its major metabolite, hydrazine, may interfere with mitochondrial function. However, hepatic mitochondria have a large reserve capacity, and minor disruption of energy homeostasis does not necessarily induce cell death. We explored whether pharmacologic or genetic impairment of mitochondrial complex I may amplify mitochondrial dysfunction and precipitate INH-induced hepatocellular injury. We found that INH (≤ 3000 μM) did not induce cell injury in cultured mouse hepatocytes, although it decreased hepatocellular respiration and ATP levels in a concentration-dependent fashion. However, coexposure of hepatocytes to INH and nontoxic concentrations of the complex I inhibitors rotenone (3 μM) or piericidin A (30 nM) resulted in massive ATP depletion and cell death. Although both rotenone and piericidin A increased MitoSox-reactive fluorescence, Mito-TEMPO or N-acetylcysteine did not attenuate the extent of cytotoxicity. However, preincubation of cells with the acylamidase inhibitor bis-p-nitrophenol phosphate provided protection from hepatocyte injury induced by rotenone/INH (but not rotenone/hydrazine), suggesting that hydrazine was the cell-damaging species. Indeed, we found that hydrazine directly inhibited the activity of solubilized complex II. Hepatocytes isolated from mutant Ndufs4(+/-) mice, although featuring moderately lower protein expression levels of this complex I subunit in liver mitochondria, exhibited unchanged hepatic complex I activity and were therefore not sensitized to INH. These data indicate that underlying inhibition of complex I, which alone is not acutely toxic, can trigger INH-induced hepatocellular injury.
authors.affiliation: University of Connecticut, Department of Pharmaceutical Sciences, Storrs, CT 06269, USA.
authors.forename: Kang Kwang
authors.initials: KK
authors.lastname: Lee
authors.affiliation: University of Connecticut, Department of Pharmaceutical Sciences, Storrs, CT 06269, USA; Daiichi Sankyo, Medicinal Safety Research Laboratories, Japan.
authors.forename: Kazunori
authors.initials: K
authors.lastname: Fujimoto
authors.affiliation: University of Connecticut, Department of Pharmaceutical Sciences, Storrs, CT 06269, USA.
authors.forename: Carmen
authors.initials: C
authors.lastname: Zhang
authors.affiliation: University of Connecticut, Department of Molecular and Cell Biology, Storrs, CT 06029, USA.
authors.forename: Christine T
authors.initials: CT
authors.lastname: Schwall
authors.affiliation: University of Connecticut, Department of Molecular and Cell Biology, Storrs, CT 06029, USA.
authors.forename: Nathan N
authors.initials: NN
authors.lastname: Alder
authors.affiliation: Auburn University, Department of Pathobiology, Auburn, AL 36849, USA.
authors.forename: Carl A
authors.initials: CA
authors.lastname: Pinkert
authors.affiliation: University of Connecticut, Department of Pharmaceutical Sciences, Storrs, CT 06269, USA.
authors.forename: Winfried
authors.initials: W
authors.lastname: Krueger
authors.affiliation: University of Connecticut, Department of Pharmaceutical Sciences, Storrs, CT 06269, USA.
authors.forename: Theodore
authors.initials: T
authors.lastname: Rasmussen
authors.affiliation: University of Connecticut, Department of Pharmaceutical Sciences, Storrs, CT 06269, USA. Electronic address: urs.boelsterli@uconn.edu.
authors.forename: Urs A
authors.initials: UA
authors.lastname: Boelsterli
mesh_terms: D000818:Animals; D000995:Antitubercular Agents / Q000378:metabolism / Q000633:toxicity*; D015153:Blotting, Western; D019069:Cell Respiration / Q000187:drug effects; D056486:Chemical and Drug Induced Liver Injury / Q000378:metabolism*; D042967:Electron Transport Complex I / Q000187:drug effects*; D022781:Hepatocytes / Q000187:drug effects*; D006834:Hydrazines / Q000378:metabolism; D007538:Isoniazid / Q000378:metabolism / Q000633:toxicity*; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D008928:Mitochondria / Q000187:drug effects*
chemical_list: D000995:Antitubercular Agents; D006834:Hydrazines; C029424:hydrazine; D042967:Electron Transport Complex I; D007538:Isoniazid
keywords: ALT; AST; Alanine aminotransferase; Aspartate aminotransferase; BNPP; Bis-p-nitrophenyl phosphate; Complex I; Complex II; DILI; Drug-induced liver injury; ETC; Electron transport chain; Hydrazine; HzN; INH; Isoniazid; Isoniazid (isonicotinic acid hydrazide); LDH; Lactate dehydrogenase; Mitochondria; Ndufs4; OCR; OXPHOS; Oxidative phosphorylation; Oxygen consumption rate; PA; Piericidin A; ROT; Rotenone; TCA; Tricarboxylic acid