Caveolin-1 targeting by its sh RNA enhances human hepatocyte response to X-ray radiation through p53-Mdm2 pathway

【作者】 李洪艳； 王洋； 史丹； 韩朝； 刘艳； 万京华； 张叶军； 刘晶； 邹伟；

【机构】 辽宁师范大学生命科学学院； 辽宁省生物技术与分子药物研发重点实验室； 人民解放军第210医院； 大连医科大学附属第一医院再生医学中心； 大连市第五人民医院；

【摘要】 Caveolin-1(Cav-1),as an important structural protein of caveolae,has been proved to be correlated with several signal transduction pathways.Recent studies showed that Cav-1 may play a critical role in response to DNA damage of cells.However,it is not clear about the mechanism of Cav-1 response to DNA damage.It is also not known whether downregulation of Cav-1 gene is required to enhance the sensitivity of human cells to X-ray radiation as Cav-1 is much more abundant in hepatocytes than in hepatoma cells.We have investigated the direct effect of Cav-1 in human hepatocyte Chang’s cells.Cav-1 knockdown cell line(CHL-CAV7)was stably established by the use of sh RNA,which inhibited the expression of Cav-1 by up to 65-70%in the parental cells(CHL cells).Cell survival ability,cell cycle distribution and the expression of proteins related to DNA damage and repair were examined after cells being radiated with X-ray.Cav-1 sh RNA induced a significant decrease in the surviving fraction of irradiated CHL cells when exposed to 8 Gy X-ray.CHL-CAV7 cells appeared to be more blocked in G1phase than CHL cells after being radiated by X-ray in 24 h.γH2AX,p-ATM and DNA-PKcs were expressed in both CHL and CHL-CAV7 cells treated by X-ray radiation.However,γH2AX expression level was higher,p-ATM and DNA-PKcs expression level was lower in CHL-CAV7 cells than that in CHL cells.P53 expression was increased in CHL cells and decreased in CHL-CAV-7 cells after recovery for 2 h after X-ray radiation.Meanwhile,the interaction of Mdm2 and Cav-1 was decreased in CHL-CAV7 cells compared to that in CHL cells after X-ray radiation.Taken together,the radiosensitivity of CHL-CAV7 cells was increased due to decreasing p53 dissociation from Mdm2 through down-regulation Cav-1.This mayprovide an idea for liver cancer therapy by down-regulation of Cav-1 expression in hepatoma cells but not in hepatocytes to increase the curative effect of X-ray radiation.更多还原

【Abstract】 Caveolin-1(Cav-1),as an important structural protein of caveolae,has been proved to be correlated with several signal transduction pathways.Recent studies showed that Cav-1 may play a critical role in response to DNA damage of cells.However,it is not clear about the mechanism of Cav-1 response to DNA damage.It is also not known whether downregulation of Cav-1 gene is required to enhance the sensitivity of human cells to X-ray radiation as Cav-1 is much more abundant in hepatocytes than in hepatoma cells.We have investigated the direct effect of Cav-1 in human hepatocyte Chang’s cells.Cav-1 knockdown cell line(CHL-CAV7) was stably established by the use of sh RNA,which inhibited the expression of Cav-1 by up to 65-70% in the parental cells(CHL cells).Cell survival ability,cell cycle distribution and the expression of proteins related to DNA damage and repair were examined after cells being radiated with X-ray.Cav-1 sh RNA induced a significant decrease in the surviving fraction of irradiated CHL cells when exposed to 8 Gy X-ray.CHL-CAV7 cells appeared to be more blocked in G1 phase than CHL cells after being radiated by X-ray in 24 h.γH2AX,p-ATM and DNA-PKcs were expressed in both CHL and CHL-CAV7 cells treated by X-ray radiation.However,γH2AX expression level was higher,p-ATM and DNA-PKcs expression level was lower in CHL-CAV7 cells than that in CHL cells.P53 expression was increased in CHL cells and decreased in CHL-CAV-7 cells after recovery for 2 h after X-ray radiation.Meanwhile,the interaction of Mdm2 and Cav-1 was decreased in CHL-CAV7 cells compared to that in CHL cells after X-ray radiation.Taken together,the radiosensitivity of CHL-CAV7 cells was increased due to decreasing p53 dissociation from Mdm2 through down-regulation Cav-1.This mayprovide an idea for liver cancer therapy by down-regulation of Cav-1 expression in hepatoma cells but not in hepatocytes to increase the curative effect of X-ray radiation.更多还原