当归红芪超滤物联合重离子^(12)C^(6+)对肝癌H22细胞辐射增敏的实验研究  

Radix Angelicae Sinensis and Radix Hedysari Enhance Radiosensitivity of Human Liver Cancer Cells to ^(12)C^(6+) Radiation

作　　者：窦春江[1] 寇炜[1] 李应东[2] 刘凯[2] 张红 

机构地区：[1]西北民族大学医学院,兰州730030 [2]甘肃中医药大学,兰州730030 [3]中国科学研究院近代物理研究所,兰州730030

出　　处：《四川大学学报（医学版）》2017年第6期840-843,861,共5页Journal of Sichuan University(Medical Sciences)

基　　金：国家自然科学基金(No.81460677;No.81160478)资助

摘　　要：目的观察当归红芪超滤物(Radix Angelicae Sinensis and Radix Hedysari,RAS-RH)增强人肝癌H22细胞对重离子^(12 )C^(6+)辐射敏感性的作用,并探讨其可能的作用机制。方法H22细胞分为空白对照组、药物组(给予RAS-RH)、辐射组(一次性给予^(12 )C^(6+)辐射)及联合组(给予RAS-RH+辐射),用CCK-8法检测RAS-RH对人肝癌H22细胞增殖抑制的影响,筛选合适的RAS-RH工作浓度;克隆集落形成实验检测RAS-RH对人肝癌H22的辐射增敏作用,筛选合适的辐射剂量;通过流式细胞仪检测RAS-RH对人肝癌H22细胞凋亡的影响,利用Western blot法检测Survivin、Caspase-9的蛋白表达水平的改变。结果RAS-RH对H22细胞的增殖抑制作用在一定的时间(0~72h)和浓度范围(0~200mg/L)内呈现时间和剂量依赖性,其细胞抑制率为20%(IC_(20))时RAS-RH的质量浓度为(117.60±2.15)mg/L,以此选定100mg/L为工作浓度;利用Prism5.0软件拟合的H22细胞生存曲线均向右下呈弧度的曲线,两条曲线明显分开呈一定的距离,联合组与辐射组相比,曲线低剂量区"肩区"缩小变窄,且各剂量点(1~10Gy)的存活率降低,2Gy时,其辐射增敏比(SER)为1.39±0.07,以此选定2Gy为辐射总剂量。联合组(100 mg/L RAS-RH+2 Gy)凋亡率高于其余3组(P<0.01);Western blot结果显示联合组的Survivin蛋白表达低于其余3组(P<0.01),而联合组的Caspase-9蛋白表达高于其余3组(P<0.01)。结论RAS-RH联合重离子^(12 )C^(6+)束对人肝癌H22细胞有辐射增敏作用,其辐射增敏机制可能通过下调凋亡抑制因子Survivin蛋白表达,上调促凋亡因子Caspase-9蛋白表达以促进细胞凋亡有关。Objective To determine the effect of Radix Angelicae Sinensis and Radix Hedysari （RAS-RH） on radiosensitivity of human liver cancer H22 cells to heavy ion ^12C6^＋ and its possible mechanism. Methods The experiment involved a comparison of proliferation of H22 cells （detected by CCK-8 assay） between four groups： control, drug （RAS-RH）, radiation, and combination （RAS-RH＋radiation）. H22 cells were treated with different doses of radiation alone or radiation followed by RAS-RH. The radiation enhancement effect of RAS-RH on H22 was detected by Colony forming assay. The effect of RAS-RH on the apoptosis of H22 cells was detected by flow cytometry. The influence of RAS-RH on the expression levels of related protein Survivin and Caspase-9 was investigated by Western blot. Results RAS-RH inhibited the proliferation of H22 cells, with a time and dose dependency 〔inhibitory concentration 20% （IC20）=（117.60±2.15） mg/L〕. The survival rate of H22 cells decreased significantly with the increase of heavy ion 12C6＋. The two survival curves produced by the Graph Pad Prism 5.0 software were clearly separated. The combination group demonstrated smaller shoulder area at low dosage and lower survival rate of cells compared with radiation group, with a sensitization enhancement ratio （SER） of 1.39±0.07. The combination group （100 mg/L RAS-RH＋2 Gy） had higher apoptosis rate and Caspase-9 protein expression level, and lower Survivin protein expression level, compared with other 3 groups （ P〈0.01）. Conclusion RAS-RH has radiation sensitization effect on human hepatocellular carcinoma H22 cells. The mechanism may be related to down-regulation of Survivin protein expression and up regulation of Caspase-9 protein expression.

关 键 词：当归红芪超滤物 H22细胞 重离子^12C6^+ 凋亡 辐射增敏 

分 类 号：R735.7[医药卫生—肿瘤]