多靶点抑制剂西奥罗尼增强人胶质瘤细胞放射敏感性的机制  

Underlying mechanism that multi-target inhibitor chiauranib enhances radiosensitivity of human glioma cells

作　　者：何超 周玉玲 王荣 魏威 刘志刚 HE Chao;ZHOU Yu-ling;WANG Rong;WEI Wei;LIU Zhi-gang(The Cancer Center,The Fifth Affiliated Hospital of Sun Yat-sen University,Guangdong Provincial Key Laboratory of Biomedical Imaging,Zhuhai 519000,China)

机构地区：[1]中山大学附属第五医院肿瘤中心,广东省生物医学影像重点实验室,广东珠海519000

出　　处：《中国病理生理杂志》2021年第9期1537-1544,共8页Chinese Journal of Pathophysiology

基　　金：国家自然科学基金资助项目(No.81572500,No.81201982);广东省自然科学基金资助项目(No.2019A1515010274,No.2021A1515010416)。

摘　　要：目的:探索多靶点抑制剂西奥罗尼对人胶质瘤细胞放射敏感性的影响及其机制。方法:利用CCK-8实验和集落形成实验测定西奥罗尼对人胶质瘤U251细胞和T98G细胞增殖能力的影响。进一步,对U251细胞和T98G细胞进行药物和(或)X射线处理,设对照组、西奥罗尼(2.5μmol/L)组、4 Gy辐照组和西奥罗尼(2.5μmol/L)联合4 Gy辐照组,流式细胞术检测4组细胞的细胞周期和凋亡变化;通过Westernblot检测4组细胞周期和凋亡关键蛋白的表达量;通过免疫荧光实验,探索西奥罗尼联合辐照对DNA损伤和修复的影响;最后通过Western blot检测DNA损伤修复关键蛋白的表达量。结果:与对照组相比,西奥罗尼显著抑制U251细胞和T98G细胞的增殖,半数抑制浓度(IC_(50))分别为7.773μmol/L和24.68μmol/L;随着X射线剂量的增加,细胞存活率在X射线辐照组和联合组中均显著降低,并且联合组在不同射线剂量的细胞存活率均低于单独辐照组,西奥罗尼的辐射增敏比(SER)分别为1.387和1.384。U251细胞对照组、西奥罗尼(2.5μmol/L)组、4 Gy辐照组和西奥罗尼(2.5μmol/L)联合4 Gy辐照组中处于G_(2)/M期的细胞比例分别为(17.68±0.98)%、(18.93±0.03)%、(16.31±0.58)%和(23.96±1.18)%;4组细胞的总凋亡率分别为(8.04±0.24)%、(17.04±0.75)%、(9.34±0.50)%和(21.22±0.38)%。T98G细胞对照组、西奥罗尼(2.5μmol/L)组、4 Gy辐照组和西奥罗尼(2.5μmol/L)联合4 Gy辐照组中处于G_(2)/M期的细胞比例分别为(2.05±0.71)%、(5.70±0.34)%、(5.14±0.33)%和(6.54±0.13)%;4组细胞的总凋亡率分别为(11.35±0.39)%、(13.23±0.18)%、(17.43±0.30)%和(21.32±0.64)%。免疫荧光实验发现,在30 min,γH2AX荧光灶点在单纯辐照组和联合组中均十分明显,并且联合组的荧光强度更强;在6 h和24 h,γH2AX荧光灶点在X射线辐照组中逐渐减少,而在联合组中可以观察到γH2AX的荧光强度依然较强。结论:西奥罗尼通过诱导G_(2)/M期阻滞、促进凋AIM:To investigate the effect of multi-target inhibitor chiauranib on the radiosensitivity of human glioma cells and its mechanism.METHODS:Human glioma U251 cells and T98G cells were cultured,and the CCK-8 assay was used to detect the growth inhibitory effect of chiauranib on U251 cells and T98G cells.The colony formation assay was used to evaluate the radiosensitivity-enhancing effect of chiauranib on U251 cells and T98G cells.The cells were divided into 4 groups:vehicle group,chiauranib(2.5μmol/L)group,4 Gy of irradiation group,and chiauranib(2.5μmol/L)+4 Gy of irradiation group.The cell cycle distribution and apoptotic rates in the 4 groups were analyzed by flow cytometry.The protein levels of key molecules related to cell cycle and apoptosis in the 4 groups were determined by Western blot.The effect of chiauranib combined with irradiation on DNA damage and repair of U251 cells and T98G cells was determined by immunofluorescence staining.The expression of key proteins related to DNA damage and repair in the 4 groups was also measured by Western blot.RESULTS:Compared with vehicle group,the proliferation of U251 cells and T98G cells was significantly inhibited after treatment with chiauranib,and the IC_(50) was calculated to be 7.773μmol/L and 24.68μmol/L,respectively.With the increase of the radiation doses,the cell survival rate was decreased in both irradiation group and combination group,more obviously in combination group,and the sensitization enhancement ratios(SER)of chiauranib for U251 cells and T98G cells were 1.387 and 1.384,respectively.The proportions of U251 cells in G_(2)/M phase in vehicle group,chiauranib(2.5μmol/L)group,4 Gy of irradiation group and chiauranib(2.5μmol/L)+4 Gy of irradiation group were(17.68±0.98)%,(18.93±0.03)%,(16.31±0.58)% and(23.96±1.18)%,respectively.The total apoptotic rates of U251 cells in the 4 groups were(8.04±0.24)%,(17.04±0.75)%,(9.34±0.50)% and(21.22±0.38)%,respectively.The proportions of T98G cells in the 4 groups were(2.05±0.71)%,(5.70±0.34)%,(5.14�

关 键 词：西奥罗尼 胶质瘤 放射敏感性 细胞凋亡 DNA损伤修复 

分 类 号：R739.4[医药卫生—肿瘤] R363.2[医药卫生—临床医学]