L Enns [1], M Weinfeld [1], KT Bogen [2], A Murtha [1]
[1]Cross Cancer Institute, Edmonton, Alberta, Canada; [2]Lawrence Livermore
National Laboratory, Livermore, California, USA.
Clinical and environmental exposure to ionizing radiation is rarely above 200 cGy. Conventional colony-forming assays are rarely sensitive enough to fully characterize altered cell proliferation/survival at doses <100 cGy. We explored an alternative method in which single cells encapsulated within ~30- to 70-µm-diameter agrarose gel microdrops (GMDs) are exposed and cultured for 4 days at 37ƒC, then analyzed by flow cytometry (FC). Clonogenic proliferation was measured as the fraction of occupied GMDs containing multicellular microcolonies after 4 days in culture. This assay was applied to human A549 lung cells exposed to 0, 5, 10, 20, 50, and 100 cGy of gamma radiation delivered at a relatively low dose rate (0.18 cGy/min). Using the GMD/FC assay, dose-response for gamma-radiation induced arrest of cell proliferation over 4 days was observed to be substantially nonlinear and non-monotonic, with significant hypersensitivity occurring at 10 cGy compared to responses observed at 5, 20, and 50 cGy (p <0.0048, <0.0015, and <0.0059, respectively). The 10-cGy dose was found to be about 10-fold more effective per unit dose than was the 20-cGy dose, and about 3-fold more effective than the 50-cGy dose, for this endpoint in A549 cells exposed at 0.18 cGy/min. The nonlinear/hypersensitivity response revealed by the GMD/FC assay applied to A549 cells underscores the importance of detailed characterizations of low-dose dose-response for radiation effects on cell proliferation, insofar as this endpoint is mechanistically linked to radiation carcinogenesis. [This work was supported by the U.S. Department of Energy (DOE) Low Dose Radiation Research Program and by the National Cancer Institute of Canada; Dr. Bogen's work was performed under DOE auspices by University of California Lawrence Livermore National Laboratory under contract No. W-7405-Eng-48.]