RESUMO
To study the mechanisms whereby cisplatin suppresses survival of human esophageal squamous cell carcinoma cells.The cytotoxicity of cisplatin in cisplatin-resistant cell line EC109 /CDDP and its parental cell line EC109 was measured by cell viability assay.Western blotting was used to investigate the protein expression of to-tal p53 and phosphorylated p53 at Ser15.Colony formation assay was employed to evaluate the ability of cells to recover from treatments and form colonies.The results indicated that EC109 /CDDP cells were more resistant to cisplatin-induced cytotoxicity than EC109 cells,with the IC50 values of (20.4 ±4.4)μmol /L and (5.7 ±0.1 )μmol /L,respectively.Although cisplatin did not alter the total protein level of p53,it obviously increased the phosphorylation of p53 at Ser15.Cisplatin inhibited survival of both EC109 /CDDP and EC109.Notably,inhibition of p53 by Pifithrin-αsignificantly promoted recovery of cisplatin-treated EC109 and EC109 /CDDP cells to differ-ent degrees.In this respect,p53 protein was found to be activated in response to cisplatin treatment in both EC109 /CDDP and EC109,which may contribute to the cytotoxic effect of cisplatin.
RESUMO
<p><b>OBJECTIVE</b>To study the effect of oncogenic Ras overexpression on autophagic activity in human fibroblast cells in vitro.</p><p><b>METHODS</b>BJ cells were transfected with H-RasV12 or control vector and treated with chloroquine, small interfering RNA (siRNA) for ATG7, or rapamycin. The cellular responses were analyzed by monitoring the parameters and biomarkers for cell growth, senescence and cell death.</p><p><b>RESULTS</b>In BJ cells overexpressing H-RasV12, chloroquine treatment resulted in more prominent cell senescence and a significantly increased cell death rate. Suppression of ATG7 mediated by siRNA also promoted cell senescence. Rapamycin treatment also caused an increased cell death rate but attenuated senescence in surviving cells. In control BJ cells, the cellular response to chloroquine included senescence and cell death, which occurred slowly. Rapamycin treatment and siRNA suppression of ATG7 had no obvious effect on control BJ cells.</p><p><b>CONCLUSION</b>Stable cellular overexpression of oncogenic Ras causes tightly controlled suppression of the autophagic activity of human fibroblast cells, and such changes produce significant effect on cell senescence and survival.</p>
Assuntos
Humanos , Autofagia , Proteína 7 Relacionada à Autofagia , Morte Celular , Células Cultivadas , Senescência Celular , Fibroblastos , Biologia Celular , Genes ras , RNA Interferente Pequeno , Enzimas Ativadoras de Ubiquitina , MetabolismoRESUMO
<p><b>OBJECTIVE</b>To explore the antagonizing effect of celecoxib against the cytotoxicity of carboplatin in human esophageal cancer cells.</p><p><b>METHODS</b>The cell viability of cisplatin-resistant cell line EC109/CDDP and its parental cell line EC109 exposed to carboplatin alone or carboplatin plus celecoxib was determined by MTT assay. The expression of CTR1, caspase-3 activation and PARP cleavage in the exposed cells were examined by Western blotting. Caspase-3 activity and cell apoptosis after the exposure were detected with Caspase-3/7 assay and flow cytometry, respectively. The effect of celecoxib on carboplatin accumulation in the cells was measured using inductively coupled plasma mass spectrometry (ICP-MS).</p><p><b>RESULTS</b>Celecoxib treatment significantly increased the IC50 of carboplatin, suppressed carboplatin-induced caspase-3 and PARP cleavage and caspase-3 activity in EC109 and EC109/CDDP cells. Celecoxib also inhibited carboplatin-induced apoptosis and suppressed intracellular carboplatin accumulation in both cell lines. A combined exposure to celecoxib and carboplatin did not cause significant changes in the protein expression of CTR1.</p><p><b>CONCLUSION</b>Celecoxib antagonizes the cytotoxic effect of carboplatin and inhibits carboplatin-induced apoptosis in human esophageal cancer cells by reducing intracellular carboplatin accumulation.</p>