Figure 9.

Combinatorial treatment of metformin and DRE on relatively resistant G361 cells. (a) Cell viability of relatively resistant G361 cells. G361 human melanoma cells were seeded on 96-well plates (~3000 cells/well) and treated at the indicated concentrations of DRE and 4 mM metformin for 72 hours. The WST-1 dye was added to each well after every treatment period and incubated, as described in Section 2. Absorbances were read at 450 nm. Statistical analysis was performed using the GraphPad Prism 5.0; **denotes a P-value <.05 compared to untreated control without metformin; ***denotes a P-value <.0005 compared to untreated control without metformin; ^##denotes a P-value <.05 compared to 5 mg/mL DRE treatment without metformin; VV denotes a P-value <.05 compared to 10 mg/mL treatment without metformin. (b) Induction of apoptosis in metformin-treated G361 cells by DRE. Typical apoptotic morphology was observed in G361 cells treated with DRE starting at 5 mg/mL concentrations for 72 hours. Cells were stained with Hoechst 33342 dye, before images were taken on a fluorescence microscope. Brightly stained, condensed bodies indicate apoptotic nuclei.