Study On The Effect And Mechanism Of Physiocn 8-O-?-glucopyranoside On Skin Melanoma Cells

OBJECTIVE: To investigate the effect of natural ingredient physiocn8-O-?-glucopyranoside(PG)on melanoma cells.METHODS: The A375 melanoma cells were pretreated with various doses of physiocn 8-O-?-glucopyranoside.The survival rate of melanoma cells was measured by CCK-8.AnnexinV/PI dying and flow cytometry were used to detect the apoptosis in melanoma cells.The expression of actived caspase-3 and PARP were detected with western-blot.The activities of caspase-9 and caspase-8 were measured by their enzyme substrates.JC-1 fluorescence probe and flow cytometry was used to determine the mitochondria membrane potential.The expression of cytochrome C inside and outside the mitochondria were measured by western-blot.Western-blot and realtime-PCR were proformed to detect the expression and transcription of COX-2.COX-2 was silenced with siRNA and overexpressed with the plasmid,and they were used in the detection of apoptosis in melanoma cells.RESULTS:During the CCK-8 test,physiocn 8-O-?-glucopyranoside was found to inhibit the viability of melanoma cells and the applicable dosed in the following experiment was also determined.Physiocn 8-O-?-glucopyranoside could enhance the apoptosis,increase the expression of active caspase-3 and PARP,impove the activity of caspase-9 but not caspase-8 as well as destroy the mitochondria membrane potential to let cytochrome C flow out to induce apoptosis.The expression and transcription of COX-2 were greatly inhibited in the physiocn 8-O-?-glucopyranoside treated melanoma cells.Silencing COX-2 increased the apoptosis in melanoma cells,and overexpression COX-2 decreased the apoptosis in melanoma which was opposite to the effect of physiocn8-O-?-glucopyranoside.CONCLUSION: Physiocn 8-O-?-glucopyranoside could inhibit the vialbility of melanoma cells as well as increase the apoptosis in cells to yield theanti-melanoma effect.The proapoptotic effect was though mitochondrial pathway but not death receptor pathway.physiocn 8-O-?-glucopyranoside induced apoptosis through suppressing COX-2 in melanoma cells.