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Tumor necrosis factor-alpha-associated lysosomal permeabilization is cathepsin B dependent
Nathan W Werneburg1, M Eugenia Guicciardi, Steven F Bronk
1Division of Gastroenterology and Hepatology, Mayo Medical School, Clinic, and Foundation, Rochester, Minnesota 55905, USA.
American Journal of Physiology. Gastrointestinal and Liver Physiology
|September 12, 2002
Summary
Tumor necrosis factor-alpha (TNF-alpha) triggers lysosomal permeabilization and cell death in hepatocytes. This study reveals sphingosine and Cathepsin B (Cat B) interaction mediates this TNF-alpha-induced lysosomal destabilization.
Area of Science:
- Cell Biology
- Biochemistry
- Hepatology
Background:
- Cathepsin B (Cat B) release from lysosomes contributes to tumor necrosis factor-alpha (TNF-alpha)-induced hepatocyte death.
- Sphingosine, elevated by TNF-alpha in the liver, is implicated in lysosomal permeabilization.
Purpose of the Study:
- To investigate the mechanisms of TNF-alpha-associated lysosomal permeabilization.
- To determine the specific role of sphingosine in this process.
Main Methods:
- Confocal microscopy to visualize Cat B-green fluorescent protein and LysoTracker Red release from lysosomes.
- Fluorescence-based and calcein release assays to assess lysosomal permeabilization in hepatocytes and isolated lysosomes.
- Utilized Cat B(+/+) and Cat B(-/-) hepatocytes and liver lysosomes.
Main Results:
- TNF-alpha/actinomycin D treatment caused release of Cat B and LysoTracker Red from lysosomes, but not Texas Red dextran from endosomes, indicating specific lysosomal targeting.
- Sphingosine induced lysosomal permeabilization in Cat B(+/+) hepatocytes and isolated lysosomes, but not in Cat B(-/-) counterparts.
- C(6) ceramide did not permeabilize lysosomes.
Conclusions:
- Sphingosine plays a critical role in TNF-alpha-induced lysosomal permeabilization.
- A sphingosine-Cathepsin B interaction is implicated in lysosomal destabilization during TNF-alpha cytotoxic signaling in hepatocytes.