Fig. 5

The direct induction of the pathway by 3 M3 mimics the αMSH mediated effect on proliferation in B16-F10 and Mel 13 melanoma cell lines. a Analysis of cell number performed on B16-F10 and Mel 13, after treatment with 15 μM 3 m3 for 24 and 48 h. Cell number was expressed as a percent variation in comparison with the value of untreated control cells. Data are mean values ± SD of three independent experiments performed in triplicate. *p < 0.01 and **p < 0.001 (vs untreated cells). b Cell-cycle distribution evaluated by flow cytometric analysis on B16-F10 and Mel 13, after treatment for 24 h with 15 μM 3 m3. The bar graph shows the distribution of cells among the different phases of the cell cycle. Data are mean values ± SD of three independent experiments performed in duplicate. *p < 0.01 (vs untreated cells). c Western blot analysis of p27 (4 h), p21 (48 h), cyclin D1 and cyclin E (6 h) protein expression on cell lysate of B16-F10 and Mel 13, treated with 15 μM 3 m3. GAPDH or HSP70 were used as an equal loading control. Results refer to three independent experiments. Representative blots are shown. Densitometric scanning of band intensities was performed to quantify the change of protein expression (control value taken as one fold in each case). ^* p < 0,01 (vs untreated cells)