Figure 1. Radiation-induced increase in surface PS is dose- and time-dependent.

(A–C) cfPac-1 cells, a human pancreatic cancer cell line, were irradiated at 0, 2, 6, 10 and 16 Gy. 24 hr later Annexin V-FITC staining was performed and the cells were processed by flow cytometry. (A) Gating of cells. FSC: forward light scattering, an indication of cell volume; SSC: side light scattering, an indication of cellular granularity. (B) Gating of live cells. PI (propidium iodide) stains only dead cells. (C) Histogram of cells stained with Annexin V-FITC shows the effects of increasing doses of radiation on Annexin V staining (i.e. surface PS). (D–H) Cells were irradiated at the indicated doses and 24 hr later Annexin V-FITC staining was determined on low surface PS cells (less than 2000 fluorescent units; (D) or moderate or high surface PS cells (greater than 3000 fluorescent units; (E). (F) Cells were irratiated at 5 Gy and analyzed at the indicated times (A2058, cfPac-1 and U87MG). To examine whether tumor cell surface PS was also increased by radiation 2 × 10⁶ cfPac-1 (G) or NCI-H460 (H) cells were injected subcutaneously into nude mice. When the tumors were ~400 mm³ some were treated with 10 Gy of targeted irradiation as described in “Materials and Methods”. The tumors were removed 48 hr. later and the cells were dispersed into single cells. The cells in all panels were then processed and gated as in Figure 1. ^*p < 0.05, ^**p < 0.01. cfPac-1 and PANC-1 are pancreatic cancer cell lines; A2058 is a melanoma cell line; NCI-H460 and H1915 are metastatic lung cancer cell lines; U87MG is a glioblastoma cell line, HPDE is a normal, immortalized pancreatic cell line and HUVEC are primary human umbilical vein endothelial cells.