The Radioprotective Mechanism Of Flavonoid Compounds Against X-rays Induced Damage In PC12Cells

In recent years, the increasing reports about that radiation damage of normal organization caused by the radiotherapies of tumor and sudden nuclear accidents. Radiation induced overproduction of free radicals, which can be an important mediator of damage to cell structures、 DNA、protein oxidation and lipid peroxidation, leading to the dysfunction of hematopoietic and immune systems, accelerating the aging process and degenerative pathological changes, and a series of radiation damage. At present, the researches on radiation damage of the central nervous system are still less, there is no typical nerve cells as vitro cells model of radiation damage.Quercetin and Breviscapine are flavonoid compounds, as the traditional Chinese herbal medicine, researches on its antioxidant activities have made some progress in domestic and abroad. Quercetin is widely exist in vegetables、fruits、tea and the other natural flavonoids, has the remarkable effect of cleaning free radicals, which has been widely used in the antioxidant study. Breviscapine can clean free radicals effectively, blocked the chain reaction of free radicals to reduce cell lipid peroxidation. So far, the reports about quercetin and breviscapine used in protective effect of free radical damage caused by radiation have not been seen in domestic and abroad. Based on the present researches on antioxidant of quercetin and breviscapine, and selected PC12cells as the model cells of radiation damage in the nervous system. Focus on the damage mechanism of free radical, investigate the radiation damage effect of multiple or single dose of X-rays. And to explore the protective effect of quercetin and breviscapine on a single dose of X-rays induced damage. In order to provide the foundation for further research on the potential radioprotective effect of quercetin and breviscapine in the nervous system. The concrete researches as follows.(一) Oxidative damage of irradiation with multiple X-rays in PC12cellsPC12cells were irradiated with50、150、450、1350mGy of X-rays, respectively. Compared with the control group, the viability of PC12cells were above100%at50、150、450mGy of X-rays after24h, while under100%at1350mGy. With the increase frequency of irradiation, the viability were decreased, showed well correlation between the viability and the accumulation of irradiation. Followed the increase frequency of irradiation, the activity of SOD were decreased(P<0.01); while the level of MDA were increased at150,450mGy(P<0.01), respectively. The results suggested that multiple middle-low dose irradiation could induce oxidative damage in PC12cells, and there were certain dose-effect relationship between the dose of irradiation and the level of damage.(二) Damage of irradiation with a single dose of X-rays in PC12cellsPC12cells were irradiated with2、4、6、8Gy of X-rays, respectively. With the increasing dose of irradiation, the viability were decreased at24h after irradiation, Compared with the control group, there were significant decrease at4、6、8Gy of X-rays(P<0.05). Compared with the control group, the level of ROS in irradiated PC12cells were significent increased, irradiated PC12 cells were inhibited at the G1phase. Followed that the structure of cells membrance were damaged, then swelling. nuclei fragments were observed.The results suggested that4Gy X-rays induced the overproduction of ROS, which inhibited cells at the G1phase. To a certain extent, that4Gy X-rays could induced damage in PC12cells.(三) Protective effect of quercetin on X-rays induced oxidative damage in PC12cellsPC12cells incubated with the different concentrations of quercetin(0、6.25、12.5.25.50and100μmol/L), quercetin induced the preliferation of PC12cells within48h, which dependent on the concentration of quercetin. While cytotoxicity at72h, that acceleration followed by the increased concentrations of quercetin(P<0.01). PC12cells were exposed to different concentrations of quercetin(0、12.5、25、50μmol/L)for2h before being irradiated to4Gy X-rays. Compared with the control group, the levels of ROS and MDA were increased, SOD and T-AOC were decreased in PC12cells at24h aftre irradiation. We did observe significant decrease in the levels of ROS and MDA, while increase in SOD and T-AOC in PC12cells treated with quercetin(P<0.01), the radioprotective efficacy of quercetin relys on its concentration. The results suggested that quercetin could ameliorates X-rays induced oxidative damage in PC12cells. Within certain limit of concentration, the radioprotective efficacy and concentration of quercetin showed good correlation.(四) Protective effect and mechanism of Breviscapine on X-rays radiation induced oxidative damage in PC12cells1. Protective effect of Breviscapine on X-rays radiation induced oxidative damage in PC12cells PC12cells incubated with the different concentrations of breviscapine(0、3.125、6.25、12.5、25、50、100and200μmol/L), breviscapine induced the preliferation of PC12cells within48h, while cytotoxicity at higher concentration than50μmol/L (P<0.05). PC12cells were exposed to different concentrations of breviscapine (0、5、10、20、40μmol/L)for2h before being irradiated to4Gy X-rays. Compared with the control group, the levels of ROS and MDA were increased, SOD and T-AOC were decreased in PC12cells at24h aftre irradiation, while the level of8-OHdG was no significant change (P>0.05). We did observed significant decrease in the levels of ROS and MDA, while increase in SOD and T-AOC in PC12cells treated with breviscapine(P<0.01), the radioprotective efficacy of breviscapine relys on its concentration.The results suggested that breviscapine could ameliorate X-rays radiation induced oxidative damage in PC12cells. The findings supported the idea that breviscapine is a free-radical scavenger and a potent radioprotector against X-rays radiation induced oxidative damage.2, The mechanism of Breviscapine against X-rays radiation induced damage in PC12cellsPC12cells were exposed to different concentrations of breviscapine(0、5、10、20、40μmol/L)for2h before being irradiated to4Gy X-rays. Compared with the control group, the cell-cycle alterations were inhibited at G1phase at24h after irradiation, breviscapine were significant released the inhibition of G1phase. The expression of JAK1、STAT1mRNA and protein were no significant changes at24h after irradiation.The results suggested that The radioprotective mechanism of breviscapine against X-rays induced damage probability are released the inhibition of G1phase, but not activated the JAK1/STAT1pathway.