ABSTRACT
<p><b>OBJECTIVE</b>To explore Akt-regulated direct p53 mitochondrial translocation in cisplatin-induced apoptosis in ovarian cancer cells and the relationship between this and chemoresistance in ovarian cancer.</p><p><b>METHODS</b>Chemosensitive ovarian cancer cell lines (OV2008 and A2780s) and chemoresistant cells (C13(*) and A2780cp) were treated with cisplatin and whole cell and mitochondrial p53 contents were determined by Western blot. The p53 accumulation in mitochondria was determined in purified mitochondrial fractions in cisplatin-sensitive and -resistant ovarian cancer cells. Akt1/2 siRNA were transfected into C13(*) cells. Cisplatin-induced apoptosis was measured by Hoechst staining and p53 translocation was determined by Western blot.</p><p><b>RESULTS</b>Cisplatin induced mitochondrial p53 accumulation and apoptosis in chemosensitive cells (P < 0.05), but not in resistant cells (P > 0.05). Over-expression of active Akt2 inhibited p53 directly translocate to mitochondria, and downregulation of Akt by Akt1/2 siRNA increased p53 mitochondrial accumulation and sensitize C13(*) cells to cisplatin treatment.</p><p><b>CONCLUSIONS</b>Cisplatin induces direct p53 mitochondrial accumulation in chemosensitive cells, and Akt confers resistance in ovarian cancer cells, in part, by regulating the direct action of p53 in mitochondrial death pathway.</p>
Subject(s)
Female , Humans , Antineoplastic Agents , Metabolism , Apoptosis , Blotting, Western , Cell Line, Tumor , Cisplatin , Metabolism , Pharmacology , Drug Resistance, Neoplasm , Mitochondria , Genetics , Ovarian Neoplasms , Drug Therapy , Proto-Oncogene Proteins c-akt , Genetics , Physiology , RNA, Small Interfering , Metabolism , Tumor Suppressor Protein p53 , Genetics , MetabolismABSTRACT
<p><b>OBJECTIVE</b>To investigate the relationship between Akt and second mitochondria-derived activator of caspases (Smac) in cisplatin (CDDP)-induced apoptosis in human ovarian cancer cells and the role of Akt in the molecular mechanism of chemoresistance in ovarian cancer.</p><p><b>METHODS</b>Chemosensitive (OV2008 and A2780s) and chemoresistant (C13* and A2780cp) ovarian cancer cell lines were treated with CDDP and subcellular Smac contents were determined by Western blot. Smac siRNA and Smac N7 peptide were transfected into OV2008 and C13* cells, respectively. CDDP-induced apoptosis was measured by flow cytometry. A2780s cells stably transfected with Akt2 (A2780s-AAkt2) and C13* cells transfected with Aktl/2 siRNA were treated with CDDP, and Smac content and apoptosis in the cells were determined to detect the changes of their chemoresistance to CDDP.</p><p><b>RESULTS</b>CDDP induced mitochondrial Smac release and apoptosis in chemosensitive cells (P < 0.05), but not resistant cells (P > 0.05). Downregulation of Smac by Smac siRNA confer resistance in OV2008 cells and Smac N7 peptide sensitized C13* cells to CDDP treatment. Overexpression of Akt2 inhibited mitochondrial Smac release and downregulation of Akt by siRNA sensitized C13* cells to CDDP treatment.</p><p><b>CONCLUSION</b>Smac is required in CDDP-induced apoptosis in ovarian cancer cells and overexpression of Akt inhibits mitochondrial Smac release. Akt is closely related to the chemoresistance of ovarian cancer.</p>
Subject(s)
Female , Humans , Antineoplastic Agents , Pharmacology , Apoptosis , Cell Line, Tumor , Cisplatin , Pharmacology , Cytoplasm , Metabolism , Drug Resistance, Neoplasm , Intracellular Signaling Peptides and Proteins , Genetics , Metabolism , Mitochondria , Metabolism , Mitochondrial Proteins , Genetics , Metabolism , Ovarian Neoplasms , Metabolism , Pathology , Proto-Oncogene Proteins c-akt , Genetics , Metabolism , RNA Interference , RNA, Small InterferingABSTRACT
<p><b>OBJECTIVE</b>To study the mechanism of multidrug resistance and its reversal by mdr-1 ribozyme in human ovarian cancer.</p><p><b>METHODS</b>The expression of mdr-1 and p-glycoprotein (p-gp) was studied by confocal laser microscope (Confocal), RT-PCR and Western blot analysis in adriamycin-resistant human ovarian cancer cell line (A2780/ADM) and adriamycin-sensitive one (A2780). The mdr-1 ribozyme was transfected into the A2780/ADM by Lipofectamine 2000 to overcome the multidrug resistance in ovarian cancer.</p><p><b>RESULTS</b>The expression of mdr-1 mRNA and p-gp in A2780/ADM was significantly higher than that in A2780. The expression of mdr-1 mRNA and p-gp in A2780/ADM was lowered after being transfected by mdr-1 ribozyme.</p><p><b>CONCLUSION</b>Multidrug resistance of A2780/ADM, possibly being caused by overexpression of mdr-1 gene, can be partially reversed by mdr-1 ribozyme.</p>