Figure 1

Metformin reduces cell proliferation and viability of Neuroblastoma SKNBE2 cells. A) Cell Index curves reporting SKNBE2 neuroblastoma cell proliferation, untreated (CTR) or treated with metformin (Met), as resulting by Xelligence RTCA DP analysis. Histograms highlight Cell Index values after 24 and 48 hrs. B) Cell counting assay of SKNBE2 cells untreated (CTR) or treated with metformin (Met). Y-axis is referred to cell number x 10⁶. C) [³H]-thymidine incorporation assay of SKNBE2 cells, untreated (CTR) or treated with metformin (Met). Y-axis is referred to cpm x 10³. D) Cell viability analysis of SKNBE2 cells untreated (CTR) or treated with metformin (Met) as determined by ATPlite assay. Y-axis is referred to luminescence x 10⁵. E) Cell death analysis by FACS analysis for sytox blue staining. The percentage of cell death is reported as Met/CTR ratio. F) Western blot analysis of caspase-3 cleavage. Equal loading of proteins was ensured by normalization for pro-caspase 3 and α-tubulin expression. Cleaved Caspase 3 was not detected. G) Western blot analysis of Akt phosphorylation/activation. 1 = Complete Medium (C.M., RPMI 10%), 2 = C.M. + 20 mM Met for 5 min, 3 = C.M. + 20 mM Met for 15 min, 4 = C.M. + 20 mM Met for 30 min, 5 = C.M. + 20 mM Met for 60 min. Equal loading of proteins was ensured by normalization for total Akt and α-tubulin expression. Densitometric analysis of phospho-Akt/total Akt ratio is also reported. (*p < 0.05; **p < 0.01).