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pRb suppresses camptothecin‐induced apoptosis in human osteosarcoma Saos‐2 cells by inhibiting c‐Jun N‐terminal kinase
Author(s) -
Lauricella Marianna,
Calvaruso Giuseppe,
Carabillò Maria,
D'Anneo Antonella,
Giuliano Michela,
Emanuele Sonia,
Vento Renza,
Tesoriere Giovanni
Publication year - 2001
Publication title -
febs letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.593
H-Index - 257
eISSN - 1873-3468
pISSN - 0014-5793
DOI - 10.1016/s0014-5793(01)02553-4
Subject(s) - camptothecin , apoptosis , transfection , microbiology and biotechnology , viability assay , acridine orange , dna fragmentation , cell cycle , growth inhibition , chemistry , cytotoxic t cell , cell culture , biology , in vitro , programmed cell death , biochemistry , genetics
This paper studies the cytotoxic effect induced by the topoisomerase I inhibitor camptothecin in human osteosarcoma Saos‐2 cells, which lack p53 and contain a non‐functional form of the product of the retinoblastoma gene, pRb. Cytotoxicity induced by camptothecin was dose‐ and time‐dependent; the treatment with 100 nM camptothecin reduced cell viability by 50% at 32 h and by 75% at 72 h of exposure. The cytotoxic effect was caused by apoptosis, as ascertained by morphological evidence, acridine orange‐ethidium bromide staining and flow cytometric analysis. Apoptosis was accompanied by both the activation of caspase‐3 and the fragmentation of poly(ADP‐ribose) polymerase. Treatment with camptothecin caused a threefold increase in the activity of c‐Jun N‐terminal kinase (JNK) and an eightfold increase in the level of phosphorylated c‐Jun. The introduction of the RB gene into Saos‐2 cells reduced the rate of cell growth. Moreover, stable clones of transfected cells were resistant to camptothecin. Exposure to 100 nM camptothecin for 72 h reduced the viability of transfected cells by only 10%; moreover, very modest effects were observed on the activity of JNK as well as on the level of phosphorylated c‐Jun. The results reported in this paper support the conclusion that the expression of wild‐type pRb in Saos‐2 cells exerts an anti‐apoptotic influence through the control of JNK activity.