Lysosomal instability and cathepsin B release during acetaminophen hepatotoxicity.
Acetaminophen (APAP) overdose is currently the most frequent cause of drug-induced liver failure in the United States. Recently, it was shown that lysosomal iron translocates to mitochondria where it contributes to the collapse of the mitochondrial membrane potential. Therefore, the purpose of this study was to investigate whether cathepsin B, a lysosomal protease, is involved in APAP-induced hepatotoxicity. Cathepsin B activity was measured in subcellular liver fractions of C57Bl/6 mice 3 hr after 300 mg/kg APAP treatment. There was a significant increase in cytoplasmic cathepsin activity, concurrent with a decrease in microsomal activity, indicative of lysosomal cathepsin B release. To investigate the effect of cathepsin B on hepatotoxicity, the cathepsin inhibitor AC-LVK-CHO was given 1 hr prior to 300 mg/kg APAP treatment along with vehicle control. There was no difference between groups in serum alanine aminotransferase (ALT) values, or by histological evaluation of necrosis, although cathepsin B activity was inhibited by 70-80% compared with controls. These findings were confirmed with a different inhibitor (z-FA-fmk) in vivo and in vitro. Hepatocytes were exposed to 5 mM acetaminophen. Lysotracker staining confirmed lysosomal instability and cathepsin B release, but there was no reduction in cell death after treatment with cathepsin B inhibitors. Finally, cathepsin B release was measured in clinical samples from patients with APAP-induced liver injury. Low levels of cathepsin B were released into plasma from overdose patients. APAP overdose causes lysosomal instability and release of cathepsin B into the cytosol but does not contribute to liver injury under these conditions.
Department of Pharmacology, Toxicology & Therapeutics, University of Kansas Medical Center, Kansas City, KS 66160, USA.
Benjamin L
BL
Woolbright
Anup
A
Ramachandran
Mitchell R
MR
McGill
Hui-min
HM
Yan
Mary Lynn
ML
Bajt
Matthew R
MR
Sharpe
John J
JJ
Lemasters
Hartmut
H
Jaeschke
D000082:Acetaminophen / Q000633:toxicity*; D000328:Adult; D000818:Animals; D002401:Cathepsin B / Q000037:antagonists & inhibitors / Q000378:metabolism*; D056486:Chemical and Drug Induced Liver Injury / Q000503:physiopathology*; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D004151:Dipeptides / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D022781:Hepatocytes / Q000187:drug effects* / Q000378:metabolism / Q000473:pathology; D006801:Humans; D007659:Ketones / Q000494:pharmacology; D008099:Liver / Q000187:drug effects / Q000378:metabolism / Q000473:pathology; D008247:Lysosomes / Q000187:drug effects / Q000473:pathology*; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D008875:Middle Aged; D011446:Prospective Studies
D015853:Cysteine Proteinase Inhibitors; D004151:Dipeptides; D007659:Ketones; C057383:MDL 201053; D000082:Acetaminophen; D002401:Cathepsin B