Mechanistic Studies On The Radioresistance Of G0 Cells To Space Heavy Ion Radiation

Health risks of astronauts exposed to high energy heavy ions in outer space have been discussed a lot mainly because its accurate assessment is complex and multifactorial. Apparently, appropriate experimental models are critical to the accurate risk assessment. Most of the cells in human body are in G0(quiescent) phase and may remain in this stage for days or even years before resuming cell division and enter G1 phase in response to exogenous stimuli such as injury. In this study, we established a pair of cell models, G0 cells obtained by contact inhibition and G1 cells that exhibit similar radiosensitivity to exponentially growing cells but contain the same amount of DNA. We compared their biological effects induced by heavy ion beams expecting to obtain the more accurate and realistic vision of risk assessment. These models may help to accelerate translational research in radiobiological studies.We tried to reveal the mechanisms underlying the cellular response of G0 cells by investigating the regulatory funcitons of P38 and RAC2 by using the cell cycle assay, survival assay, immunofluorescence, cytokinesis-block micronucleus cytome assay, Western blot and GST pull down assay. Main results of the study are as follows:1. G0 cells are more resistant to ionizing radiation than G1 cells.2. The expression of RAC2, a catalytic subunit of NADPH oxidase, was relatively low in G0 cells but high in G1 cells. Also, relative to G1 cells, the total antioxidant capacity(TAC) markedly increased upon exposure to X-rays whereas the intracellular yield of ROS only slightly increased in G0 cells. We found that knockdown of RAC2 resulted in significant reduction in the yield of ROS in G1 cells. The induction of DNA single- and double-strand breaks in G1 cells by X-rays was also inhibited by the knockdown of RAC2. These findings suggest that low level of RAC2 in G0 cells exposed to X-rays results in low level of oxidative stress and DNA damage.3. P38 MAP kinase(P38MAPK) is activated in response to ionizing radiation. P38 MAPK was expressed at low levels in G0 cells while high levels in G1 cells after X-rays exposure. P38 MAPK was phosphorylated more in G0 cells than G1 cells. The intracellularlocalization of P38 MAPK together with RAC2 leads to their nuclear translocation in G0 cells after X-rays exposure, which resulted in the inactivation of RAC2. This phenominen was not observed in G1 cells. In addition, P38 MAPK phosphorylation increased the radioresistance of G0 cells. Feedback loop formed by P38 MAPK, phosphorylated P38 MAPK and RAC2 regulates the radioresistance of G0 cells.