Study On Heavy Ion Induced Late Effects In A549 Cell Line And Hippocampus Of Balb/c Mouse

Cancer is not only a serious threat to human health, but also a significant challenge to modern medicine. Radiotherapy is one of the most commonly-used cancer treatment modalities. Owing to their advantages in terms of physics and biology, energetic heavy ions have become an ideal radiation type in radiotherapy. Heavy ions are also important because astronauts who perform deep space mission are threatened with significant central nervous system(CNS) damage due to space heavy ion radiation. Neither cancer patients treated with heavy ions nor astronauts are immune from the late effects caused by heavy ion radiation. The needs of research projects on these effects(and methods of ameliorating them) have garnered increasing amounts of attention, especially in terms of whether or not cancer cells grow genetically unstable after exposure to heavy ion radiation, as well as ion radiation exposure’s long-term influence on the central nervous system. Both aspects were intensively analyzed in this study. We first investigated the genesis and features of late effects of human nonsmall-cell lung cancer cell line(A549) caused by heavy ion exposure and examined the influence of autophagy on heavy ion-induced late effects in A549 cells. Second, we irradiated the heads of Balb/c mice with carbon ions of various linear energy transfers(LET) in order to analyze the relationship between long-term behavior pattern changes, and the expression of autophagy and Nrf2 signaling-related proteins.To investigate heavy ion-induced late effects in A549 cells, we employed carbon ions(initial energy=165 Me V/u, LET=70 ke V/μm on cell samples) and iron ions (initial energy=158 Me V/u, LET=900 ke V/μm on cell samples). To elucidate the genesis mechanism of these late effects, bio-endpoints(colony formation, spontaneous micronuclei formation, and number of chromosomes) were measured one month after radiation exposure. The heterogenous response of bystander cells to factors causing non-DNA-targeted effects were identified using the cell growth curve, number of cells per spot, and the number of chromosomes in bystander cells one month after carbon ion radiation; the carbon ion radiation-induced late effects in rapamycin/chloroquine pretreated A549 cells were measured according to the number of chromosomes, colony formation, spontaneous micronuclei formation, invasion assay, wound healing, and drug/radiation resistance to explore the influence of autophagy regulation in heavy ion radiation-induced late effects.The heads of three-week old Balb/c female mice were irradiated with carbon ions through a carefully designed radiation device. The acute autophagy and Nrf2 signaling response to carbon ion radiation were then measured by performing Western bolt tests. Motion, spatial learning and memory, and novel object recognition behavioral tests were performed to evaluate any CNS injury to the mice three months after irradiation. The expression of autophagy and Nrf2 signaling-related proteins were then measured by Western blot to examine the roles they played in the CNS injuries induced in the mice by heavy ion radiation.The main conclusions drawn from this study can be summarized as follows:1. Study on heavy ion radiation induced late effects in A549 cell line. 1) The late effects induced by heavy ion radiation in nonsmall-cell lung cancer cell line(A549) were mainly mediated by non-DNA targeted factors. The only way by which late effects were mediated was soluble bystander factors when the radiation dose was high; when the radiation dose was low, they might be mediated by gap junctional intercellular communication in confluence growth cells as well.2) A549 featured response heterogeneity to non-DNA targeted factors that caused the late effects. Cells which were more heavily influenced by non-DNA targeted factors proliferated slowly at first, and then their proliferation returned to normal. These characteristics may represent the cause of the delayed radiation effect. 3) A549 cells pretreated with chloroquine had no impact on heavy ion-induced late effects. On the contrary, pretreatment with rapamycin led to A549 cells with increased chromosome instability and enhanced invasion and metastasis ability. Pretreatment with rapamycin also sensitized A549 cells to anti-cancer drugs and X-rays. A549 cells simply irradiated with carbon ions(2Gy) did not cause increased malignancy of tumor cells(such as enhanced drug/X-ray resistance, increased invasion and migration capacity, or increased proportion of tumor stem cells.)2. Study on heavy ion radiation induced late effects in CNS of Balb/c mouse. 1) Exposure of the immature mouse hippocampus to carbon ions caused an LET-dependent behavior pattern change after maturation and an LET-dependent differential molecular response. Carbon ions with higher LET values more readily induced a reduced cognitive performance in the mice, as evidenced by their declining performance behavioral tasks. 2) Autophagy was intact in the hippocampal cells of the mature mice after irradiation. 3) The persistently high nuclear Nrf2 content in hippocampal cells might account for the unchanged behavioral patterns of mice after exposure to relatively low LET carbon ions, as well as the subsequent increased radioresistance of the hippocampal cells.