摘要
目的探讨槲皮素对人宫颈癌He La细胞株的放射增敏作用及槲皮素放射增敏作用机理。方法克隆形成法研究不同浓度槲皮素(20%IC_(50)和IC_(50))及不同给药时序(先放后药和先药后放)对宫颈癌He La细胞的放射增敏作用,根据实验设计分为单放组、20%IC_(50)槲皮素先药后放组、20%IC_(50)槲皮素先放后药组、IC_(50)槲皮素先药后放组、IC_(50)槲皮素先放后药组,检测各组经不同量X线照射(12、8、6、4、2、0 Gy)后的细胞存活率(SF)。选择上述实验结果最佳给药时序,将实验设计为未加药对照组、单放组、单药1组(20%IC_(50))、单药2组(IC_(50))、药放1组(20%IC_(50))及药放2组(IC_(50)),放疗剂量为4Gy,分别采用流式细胞法、DAPI染色、Western blotting(仅采用20%IC_(50))检测各组处理24 h后的细胞周期分布、细胞凋亡变化及Bcl-2、Bax蛋白水平。结果不同浓度槲皮素及不同剂量射线按不同时序处理后的各组细胞与单放组比较SF均下降(P<0.05)。SF与槲皮素浓度呈反比,药物浓度越高,槲皮素的放射增敏作用越强。相同浓度药物和相同剂量射线不同处理时序相比较,先放后药组的SF均比先药后放组低。DAPI染色后荧光显微镜显示,放射线及槲皮素均可引起He La细胞核不同程度的皱缩,两者联合作用后胞核皱缩、碎裂更为明显。流式细胞仪检测显示,与其他各组比较,药放联合组G2/M期阻滞更显著(P<0.05)。Western blotting检测显示,药放联合组Bcl-2蛋白表达下调较其他各组更加显著(P<0.05),但Bax蛋白表达上调与槲皮素单药组比较差异无统计学意义。结论槲皮素对He La细胞具有放射增敏作用,其增敏作用具有浓度及时序效应;放疗增敏作用的机制可能与诱导细胞凋亡、阻滞细胞于G2/M期及下调Bcl-2/Bax比例有关。
Objective To investigate the radiosensitization effect of quercetin on cervical cancer HeLa cells and to explore its possible mechanism. Methods Clone forming assay was used to observe the radiosensitizing effect of quercetin on HeLa cells after ra-diation with drug by various schedules and different drug concentrations（20% IC50 and IC50）. According to the experimental protocol, the experiments were carried out in radiation alone group, 20% IC50 quercetin＋radiation group, radiation＋20% IC50 quercetin group, IC50 quercetin＋radiation group and radiation＋IC50 quercetin group. The surviving fraction（ SF） of above all groups after X-ray radiation of 12, 8, 6, 4, 2 and 0 Gy were calculated and the optimum schedule was selected. The below experiments were carried out in negative control group, quercetin group（20% IC50 and IC50）, radiation group and quercetin＋irradiation group（20% IC50 and IC50）, and the dose of radiation was 4 Gy. Flow cytometry, DAPI staining, Western blotting（20%IC50 quercetin only） were explored to detect the cell cycle distribution, apoptotic changes and expression of Bcl？2 and Bax protein at 24 h after treatment. Results Compared with radiation alone group, the SF were reduced by the treatment of different concentration of radiation, quercetin in different sequence（P〈0？05）. The SF were inversely proportional to the concentration of quercetin, while the radiosensitization of quercetin was in concentration de-pended manner. Under the same concentration of quercetin and dose of X-ray, the SF in radiation＋quercetin groups were lower than those in quercetin＋radiation groups. DAPI staining showed that nucleus shrinkage were observed with the treatment of quercetin or radi-ation, and the combination of them could cause nucleus shrinkage and fragmentation more marked. Flow cytometry demonstrated that compare with other groups, quercetin＋radiation groups could arrest cell cycle in G2/M phase（ P〈0？05） . Western blotting showed that＆nbsp;the levels of Bcl？2 protein was lowed in quercetin＋radiation group than other groups（ P〈0？05）;Bax levels increased in quercetin＋radi-ation group, but it had no significance compared with quercetin group. Conclusion Quercetin has radiosensitization effect on HeLa cells and its effect is influenced by schedule and concentration of quercetin. The mechanism may be related to arrest of cell cycle in G2/M phase, cell apoptosis and the down-regulating the ratio of Bcl？2/Bax.
出处
《临床肿瘤学杂志》
CAS
2016年第3期208-212,共5页
Chinese Clinical Oncology
基金
十堰市科技局指导项目(14Y33)
湖北医药学院"2011开放基金"项目(2011JH-2014CXTT01)
