Tumor necrosis factor-α-associated lysosomal permeabilization is cathepsin B dependent

Nathan W. Werneburg, M. Eugenia Guicciardi, Steven F. Bronk, Gregory J. Gores

Research output: Contribution to journalArticle

146 Scopus citations

Abstract

Cathepsin B (Cat B) is released from lysososomes during tumor necrosis factor-α (TNF-α) cytotoxic signaling in hepatocytes and contributes to cell death. Sphingosine has recently been implicated in lysosomal permeabilization and is increased in the liver by TNF-α. Thus the aims of this study were to examine the mechanisms involved in TNF-α-associated lysosomal permeabilization, especially the role of sphingosine. Confocal microscopy demonstrated Cat B-green fluorescent protein and LysoTracker Red were both released from lysosomes after treatment of McNtcp.24 cells with TNF-α/actinomycin D, a finding compatible with lysosomal destabilization. In contrast, endosomes labeled with Texas Red dextran remained intact, suggesting lysosomes were specifically targeted for permeabilization. LysoTracker Red was released from lysosomes in hepatocytes treated with TNF-α or sphingosine in Cat B(+/+) but not Cat B(-/-) hepatocytes, as assessed by a fluorescence-based assay. With the use of a calcein release assay in isolated lysosomes, sphingosine permeabilized liver lysosomes isolated from Cat B(+/+) but not Cat B(-/-) liver. C6 ceramide did not permeabilize lysosomes. In conclusion, these data implicate a sphingosine-Cat B interaction inducing lysosomal destabilization during TNF-α cytotoxic signaling.

Original languageEnglish (US)
Pages (from-to)G947-G956
JournalAmerican Journal of Physiology - Gastrointestinal and Liver Physiology
Volume283
Issue number4 46-4
DOIs
StatePublished - Oct 2002

Keywords

  • Calcein release assay
  • Cathepsin B-green fluorescence protein
  • LysoTracker Red
  • Sphingosine

ASJC Scopus subject areas

  • Physiology
  • Hepatology
  • Gastroenterology
  • Physiology (medical)

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