Proton beam irradiation inhibits the migration of melanoma cells

In recent years experimental data have indicated that low-energy proton beam radiation might induce a difference in cellular migration in comparison to photons. We therefore set out to compare the effect of proton beam irradiation and X-rays on the survival and long-term migratory properties of two cell lines: uveal melanoma Mel270 and skin melanoma BLM.Cells treated with either proton beam or X-rays were analyzed for their survival using clonogenic assay and MTT test. Long-term migratory properties were assessed with time-lapse monitoring of individual cell movements, wound test and transpore migration, while the expression of the related proteins was measured with western blot.Exposure to proton beam and X-rays led to similar survival but the quality of the cell colonies was markedly different. More paraclones with a low proliferative activity and fewer highly-proliferative holoclones were found after proton beam irradiation in comparison to X-rays. At 20 or 40 days post-irradiation, migratory capacity was decreased more by proton beam than by X-rays. The beta-1-integrin level was decreased in Mel270 cells after both types of radiation, while vimentin, a marker of EMT, was increased in BLM cells only.We conclude that proton beam irradiation induced long-term inhibition of cellular motility, as well as changes in the level of beta-1 integrin and vimentin. If confirmed, the change in the quality, but not in the number of colonies after proton beam irradiation might favor tumor growth inhibition after fractionated proton therapy

Cellular migration properties of Mel270 cells treated with proton beam radiation or X rays.

<p>Individual cell movements were evaluated at 20 days after irradiation (A, B, C) and at 40 days after irradiation (D, E, F) and three parameters were calculated: ‘Speed’, i.e. average speed of cell movement; ‘Displacement’, i.e. the total linear length of the cell displacement from the starting point (μm) and CME (coefficient of movement efficiency), i.e. the ratio of cell displacement to the cell trajectory length. Mean values presented as percent of control; *p<0.05, **p<0.01, ***p<0.001.</p

Proton beam irradiation inhibits the migration of melanoma cells - Fig 1

<p><b>Clonogenic assay of cell survival of Mel270 (A) and BLM (B) cells, treated with proton beam (■) or X rays (●).</b> Representative images of colonies are presented at <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0186002#pone.0186002.s001" target="_blank">S1 Fig</a>. Cell were seeded immediately after radiation. Mean values with SEM, *p<0.05; **p<0.001. RBE values were determined from a linear-quadratic model and were 1.10 for Mel270, and 1.13 for BLM cells. (C–E) Three types of colonies formed by Mel270 (C) and BLM (D) cells in two weeks after irradiation with 1–5 Gy of proton beam or X-rays, determined as the percentage of the total number of colonies. Mel270 and BLM cells form three types of colonies described as holo-, mero- and paraclones (E). Holoclones are large, packed colonies displaying heterogeneity, which are believed to be derived from cancer initiating cells; meroclones are putatively derived from transit-amplifying cells and paraclones are loosely packed cells, derived from differentiated cells [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0186002#pone.0186002.ref017" target="_blank">17</a>].</p

Cellular migration properties of BLM cells treated with proton beam radiation or X rays.

<p>Individual cell movements were evaluated at 20 days after irradiation (A, B, C) and at 40 days after irradiation (D, E, F) and were evaluated in terms of ‘Speed’, i.e. average speed of cell movement; ‘Displacement’, i.e. the total linear length of the cell displacement from the starting point (μm) and CME (coefficient of movement efficiency), i.e. the ratio of cell displacement to the cell trajectory length. Mean values presented as percent of control; *p<0.05, **p<0.01, ***p<0.001.</p