The absence of an early calcium response to heavy-ion radiation in Mammalian cells.

Intracellular calcium is an important second messenger that regulates many cell functions. Recent studies have shown that calcium ions can also regulate the cellular responses to ionizing radiation. However, previous data are restricted to cells treated with low-LET radiations (X rays, gamma rays and beta particles). In this work, we investigated the calcium levels in cells exposed to heavy-ion radiation of high LET. The experiments were performed at the single ion hit facility of the GSI heavy-ion microprobe. Using a built-in online calcium imaging system, the intracellular calcium concentrations were examined in HeLa cells and human foreskin fibroblast AG1522-D cells before and after irradiation with 4.8 MeV/nucleon carbon or argon ions. Although the experiment was sensitive enough to detect the calcium response to other known stimuli, no response to heavy-ion radiation was found in these two cell types. We also found that heavy-ion radiation has no impact on calcium oscillation induced by hypoxia stress in fibroblast cells.

Targeted irradiation of Mammalian cells using a heavy-ion microprobe.

The existing focusing heavy-ion microprobe at the Gesellschaft für Schwerionenforschung in Darmstadt (Germany) has been modified to enable the targeted irradiation of single, selected cells with a defined number of ions. With this setup, ions in the range from helium to uranium with linear energy transfers (LETs) up to approximately 15,000 keV/microm can be positioned with a precision of a few micrometers in the nuclei of single cells that are growing in culture on a thin polypropylene film. To achieve this accuracy, the microbeam traverses a thin vacuum window with minimal scattering. Electron emission from that window is used for particle detection. The cells are kept in a specially designed dish that is mounted directly behind the vacuum window in a setup allowing the precise movement and the imaging of the sample with microscopic methods. The cells are located by an integrated software program that also controls the rapid deflection and switching of the beam. In this paper, the setup is described in detail together with the first experiments showing its performance. We describe the ability of the microprobe to reliably hit randomly positioned etched nuclear tracks in CR-39 with single ions as well as the ability to visualize the ion hits using immunofluorescence staining for 53BP1 as a marker of DNA damage in the targeted cell nuclei.