Ubiquitin-proteasome inhibition enhances apoptosis of human pancreatic cancer cells

Autor: Richard A. Perugini, Mark P. Callery, Theodore P. McDade, Frank J. Vittimberga
Rok vydání: 1999
Předmět:
Zdroj: Surgery. 126:371-377
ISSN: 0039-6060
DOI: 10.1016/s0039-6060(99)70179-0
Popis: Background: Tumor necrosis factor (TNF-α)–induced apoptosis is limited by coactivation of nuclear factor kappa B (NF-kb)–dependent antiapoptotic genes. Nuclear translocation of NF-kB requires degradation of ubiquitinated phospho-IkB-a by the 26S proteasome. We examined whether inhibition of the ubiquitin-proteasome pathway enhances TNF-α–induced apoptosis in BxPC-3 human pancreatic cancer cells. Methods: Serum-starved BxPC-3 cells (12 hours) were pretreated or not for 50 minutes with PSI (30 μmol/L), a peptide aldehyde known to inhibit specifically the chymotrypsin-like activity of the 26S proteasome. Cells were subsequently stimulated with recombinant human TNF-α (400 units/mL). Western blots were performed using antibodies to IκB-α and phospho-IκB-α. Level of apoptosis was determined by two methods: enzyme-linked immunosorbent assay detection of interhistone DNA fragments and flow cytometry with propidium iodide staining. Results: TNF-α–induced degradation of IκB-α was inhibited by PSI. Phospho-IκB-α accumulation was observed 20 minutes after TNF-α stimulation. Apoptosis relative to constitutive levels was significantly increased after PSI pretreatment, as measured by DNA fragmentation (P ≤ .05 by Student t test). Percent apoptosis by flow cytometry confirmed marked increases in apoptotic cell fractions from 5.9% (untreated) to 6.8% (TNF-α alone), 16.4% (PSI alone), and 18.9% (PSI and TNF-α). Conclusions: PSI enhances both constitutive and TNF-α–induced apoptosis through inhibition of IκB-α degradation in BxPC-3 human pancreatic cancer cells. (Surgery 1999;371-7.)
Databáze: OpenAIRE
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