Figure 4: Synergetic effect of gemcitabine and irradiation in vitro and in vivo.

(a) For in vitro analysis, a proliferation assay was performed. After seeding 300 cells per well of a 96-well plate for more than 2 h, the conditioned medium was changed to medium containing gemcitabine. After 12 h, the plate was irradiated and the medium replaced with gemcitabine-free medium and incubated for 5 days. All data were normalised to the 0 Gy/gemcitabine-free condition. The synergistic effect was analysed using a generalised linear model, which revealed that not only gemcitabine and irradiation alone but also their combination significantly impaired proliferation (p < 0.01, p < 0.01, and p = 0.034, respectively). Data are derived from three independent experiments. (b) Representative images for in vivo analysis. B16BL6 cells were inoculated into the left thigh and treated with irradiation and gemcitabine. Mice treated by gemcitabine before irradiation were defined as the synchronised group, while mice irradiated before gemcitabine treatment were defined as the unsynchronised group. Xenografts grew rapidly and mice with too large xenografts were euthanised. (c) Compared with the unsynchronised group, the growth of xenografts in the synchronised group was prevented significantly. (n = 11 for the synchronised group, n = 10 for the unsynchronised group) bar: SEM, *p < 0.05.