JTE-522-induced apoptosis in human gastric adenocarcinoma [correction of adenocarcinoma] cell line AGS cells by caspase activation accompanying cytochrome C release, membrane translocation of Bax and loss of mitochondrial membrane potential

World J Gastroenterol. 2002 Apr;8(2):217-23. doi: 10.3748/wjg.v8.i2.217.

Abstract

Aim: To investigate the role of the mitochondrial pathway in JTE-522-induced apoptosis and to investigate the relationship between cytochrome C release, caspase activity and loss of mitochondrial membrane potential (Deltapsim).

Methods: Cell culture, cell counting, ELISA assay, TUNEL, flow cytometry, Western blot and fluorometric assay were employed to investigate the effect of JTE-522 on cell proliferation and apoptosis in AGS cells and related molecular mechanism.

Results: JTE-522 inhibited the growth of AGS cells and induced the apoptosis. Caspases 8 and 9 were activated during apoptosis as judged by the appearance of cleavage products from procaspase and the caspase activities to cleave specific fluorogenic substrates. To elucidate whether the activation of caspases 8 and 9 was required for the apoptosis induction, we examined the effect of caspase-specific inhibitors on apoptosis. The results showed that caspase inhibitors significantly inhibited the apoptosis induced by JTE-522. In addition, the membrane translocation of Bax and cytosolic release of cytochrome C accompanying with the decrease of the uptake of Rhodamin 123, were detected at an early stage of apoptosis. Furthermore, Bax translocation, cytochrome C release, and caspase 9 activation were blocked by Z-VAD.fmk and Z-IETD-CHO.

Conclusion: The present data indicate a crucial association between activation of caspases 8, 9, cytochrome C release, membrane translocation of Bax, loss of Deltapsim and JTE-522-induced apoptosis in AGS cells.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adenocarcinoma*
  • Amino Acid Chloromethyl Ketones / pharmacology
  • Anti-Inflammatory Agents, Non-Steroidal / pharmacology
  • Apoptosis / drug effects*
  • Apoptosis / physiology
  • Benzenesulfonates / pharmacology*
  • Caspase Inhibitors
  • Caspases / metabolism*
  • Cyclooxygenase Inhibitors / pharmacology
  • Cysteine Proteinase Inhibitors / pharmacology
  • Cytochrome c Group / metabolism*
  • Enzyme Activation
  • Humans
  • In Situ Nick-End Labeling
  • Membrane Potentials / physiology
  • Mitochondria / drug effects*
  • Mitochondria / metabolism
  • Oxazoles / pharmacology*
  • Proto-Oncogene Proteins / metabolism*
  • Proto-Oncogene Proteins c-bcl-2 / metabolism
  • Stomach Neoplasms*
  • Tumor Cells, Cultured
  • bcl-2-Associated X Protein

Substances

  • 4-(4-cyclohexyl-2-methyloxazol-5-yl)-2-fluorobenzenesulfonamide
  • Amino Acid Chloromethyl Ketones
  • Anti-Inflammatory Agents, Non-Steroidal
  • BAX protein, human
  • Benzenesulfonates
  • Caspase Inhibitors
  • Cyclooxygenase Inhibitors
  • Cysteine Proteinase Inhibitors
  • Cytochrome c Group
  • Oxazoles
  • Proto-Oncogene Proteins
  • Proto-Oncogene Proteins c-bcl-2
  • bcl-2-Associated X Protein
  • benzyloxycarbonylvalyl-alanyl-aspartyl fluoromethyl ketone
  • Caspases