|
Objective: To study geldanamycin (Geldanamycin, GA) and the anti-cancer drug cisplatin (DDP) combined inhibition in vitro of human ovarian cancer cell line SKOV3 analysis geldanamycin inhibition of SKOV3 cell proliferation and induced The possible mechanisms of apoptosis explore geldanamycin treatment for ovarian cancer clinical feasibility. Method: 1 in vitro culture of human ovarian cancer SKOV3, using methyl thiazolyl tetrazolium (tetrzolium-based colorimetric assay, MTT) colorimetric assay geldanamycin affect the cell proliferation. Geldanamycin 48 after Giemsa staining microscope observation SKOV3 cell morphological changes and flow cytometry (flow cytometry, FCM) changes in the cell cycle and apoptosis rate. Immunocytochemistry (immunocytochemistory, ICC) detection geldanamycin for 48 hours after the SKOV3 cells Akt and raf-1 protein expression. 4 Flow cytometry analysis the geldanamycin processing SKOV3 cells 48 Akt, raf-1 protein expression levels. 5 MTT colorimetric assay geldanamycin and cisplatin combination of inhibition of proliferation of SKOV3 cells, Guinness formula to analyze the interaction index (Interaction Index), evaluation of the two drugs in combination, such as interaction index > 0.85 indicates an additive effect, the interaction index <0.85 for antagonism. Result: a different concentration of geldanamycin (20,40,200,400,2000 nmol / L) on the proliferation of SKOV3 cells can produce inhibition geldanamycin inhibits proliferation of SKOV3 cells was time - a dose-dependent relationship, the maximum inhibition rate (74.72 ± 3.16)%. The GA after 48h IC50 496.57nmol / L. 2 is by 20,40,200,400,2000 nmol / L geldanamycin role SKOV3 cells after 48h, with the increase of the concentration of the drug, G 2 / M phase cells gradually increased the The effects were dose - dependent manner. Different concentrations of geldanamycin induced apoptosis and apoptosis rate compared to the control group, the difference was statistically significant (P <0.05). 3 optical microscope observation SKOV3 cells without the the geldanamycin role of the control group fusiform or rhomboid shape and full: while geldanamycin after 48h SKOV3 cell shrinkage, cracking, irregular-shaped some cells at both ends of slender pseudopodia, cytoplasmic vacuoles, an increase in exfoliated cells. Display after Giemsa staining under a light microscope, the control group SKOV3 cells fusiform or diamond-shaped, shape and full uniform nuclear staining visible nucleoli, no apoptotic cells; geldanamycin for 48h after SKOV3 cell rounding, cytoplasmic enrichment, nuclear uneven coloring, nuclear fission phenomenon appears, showing the morphological changes of apoptotic cells. The immunocytochemistry assay show, via 0,20,200,2000 nmol / L geldanamycin for 48 hours, SKOV3 cells Akt, raf-1 protein expression gradually decreased with the increase in drug concentration. The flow cytometry results also showed that Akt, raf-1 protein expression was significantly lower (P <0.05). 5 MTT geldanamycin and cisplatin significantly inhibited the proliferation of SKOV3 cells. 20,40,200,400 nmol / L geldanamycin combined with cisplatin on SKOV3 cell proliferation inhibition with increasing concentrations of gradually enhanced. Guinness formula to analyze interactions Index calculated results show that different concentrations of cisplatin combined with different concentrations of geldanamycin interaction index> 0.85. Conclusion: 1 geldanamycin significantly inhibited the proliferation of human ovarian cancer SKOV3, provides a new means for the clinical treatment of ovarian cancer. Geldanamycin inhibits the proliferation of SKOV3 cells showed the time a dose-dependent manner. Geldanamycin allows SKOV3 cells were arrested in G 2 / M phase and may induce apoptosis. 4 geldanamycin by downregulating Akt, raf-1 protein expression and play to inhibit SKOV3 cell proliferation and induce apoptosis. Geldanamycin and cisplatin can enhance the antiproliferative effects of cisplatin on SKOV3 cells.
|