培美曲塞对PC9肺腺癌细胞株的放射增敏作用

发布时间：2018-05-17 22:36

本文选题：肺腺癌 + PC细胞　；参考：《中华肿瘤防治杂志》2017年09期

【摘要】：目的培美曲塞是治疗非鳞癌非小细胞肺癌(non-small cell lung cancer,NSCLC)的常用药物,其对表皮生长因子受体(pithelial growth factor receptor,EGFR)敏感突变的肺腺癌是否具有放射增敏作用尚未明确。本课题研究培美曲塞在体外对EGFR 19外显子突变的人肺腺癌细胞株PC9的放射增敏作用,并初步探讨其作用机制。方法以PC9细胞作为研究对象,采用MTT法检测培美曲塞的20%抑制浓度(20%inhibition concentration,IC_(20)),将PC9细胞分为对照组、单独照射组、培美曲塞单药组和培美曲塞+照射组4组;采用流式细胞术检测培美曲塞联合或不联合照射对PC9细胞的凋亡率及细胞周期的影响;克隆形成实验检测培美曲塞对PC9细胞的放射效应,计算存活分数,拟合存活曲线;蛋白质印迹法检测CDC20蛋白在各组的表达。结果 MTT结果显示,PC9细胞的增殖抑制与培美曲塞呈时间-剂量依赖性,取48h的IC_(20)(0.084μmol/L)的培美曲塞作为实验浓度。流式细胞术结果显示,培美曲塞单药组和单独照射组均可增加凋亡率,分别为(7.17±1.14)%和(10.78±1.52)%,而培美曲塞+照射组具有协同增效作用,凋亡率达(29.23±1.33)%,F=227.57,P0.001;细胞周期结果显示,对照组G_2/M期比例为(0.79±0.63)%,培美曲塞单药组为(0.79±0.47)%,单独照射组为(18.21±0.72)%,培美曲塞+照射组为(25.09±1.04)%,提示培美曲塞使细胞阻滞在S期,与放射线联合作用后,S期细胞比例减少,G_2/M期的比例明显增多,差异有统计学意义,F=276.85,P0.001。克隆形成实验提示,培美曲塞具有较好的放射增敏作用,其放射增敏比(sensitization enhancement ratio,SER)为1.41。培美曲塞联合照射能增加细胞周期相关蛋白CDC20的表达,F=282.12,P0.001。结论培美曲塞可提高EGFR 19外显子突变的PC9肺腺癌细胞的放射敏感性,其机制可能与照射引起CDC20的增加致对放射敏感的G_2/M期阻滞,而照射耐受的S期比例减少有关。
[Abstract]:Objective pemetrexil is a common drug in the treatment of non-small cell lung cancer (NSCLC). It is not clear whether pemetrexed has radiosensitization effect on epidermal growth factor receptor (EGFR) sensitive mutation in lung adenocarcinoma. The aim of this study was to investigate the radiosensitizing effect of pemetrexil on human lung adenocarcinoma cell line PC9 with exon 19 mutation of EGFR in vitro, and to explore its mechanism. Methods PC9 cells were used to detect 20% inhibition concentration of pemetrexide by MTT. PC9 cells were divided into 4 groups: control group, single irradiation group, pemetrexed single drug group and pemetrexed irradiation group. Flow cytometry was used to detect the apoptosis rate and cell cycle of PC9 cells induced by combined or uncombined irradiation of pemetrexide, and clone formation assay was used to detect the radiative effect of pemetrexide on PC9 cells, to calculate the survival fraction and to fit the survival curve. The expression of CDC20 protein in each group was detected by Western blot. Results the results of MTT showed that the inhibition of proliferation of PC9 cells was time-dose dependent on pemetrexide, and the concentration of pemetrexil was 48 h IC_(20)(0.084 渭 mol / L as the experimental concentration. The results of flow cytometry showed that the apoptotic rate was increased in both pemetrexide group and irradiation group, which were 7.17 卤1.14% and 10.78 卤1.52%, respectively, while that in pemetrexed group was synergistic, and the apoptotic rate was 29.23 卤1.33% (P 0.001). In the control group, the ratio of G2 / M phase was 0.79 卤0.63, that of pemetrexed group was 0.79 卤0.47g, that of single irradiation group was 18.21 卤0.72g, and that of pemetrexed irradiation group was 25.09 卤1.040.The results showed that pemetrexil made the cells block in S phase, and the proportion of cells in S phase decreased in G2M phase after combined irradiation. The difference was statistically significant (P 0.001). Clone formation experiment showed that pemetrexil had better radiosensitization, and its radiosensitization ratio was 1.41%. The combined irradiation of pemetrexide could increase the expression of cell cycle associated protein CDC20. Conclusion pemetrexil can increase the radiosensitivity of PC9 lung adenocarcinoma cells with mutation of exon 19 of EGFR, and its mechanism may be related to the increase of CDC20 induced by irradiation and the arrest of G 2 / M phase of radiation sensitive to radiation, but the decrease of S phase ratio of radiation tolerance.
【作者单位】：广西壮族自治区人民医院化疗一区;广西壮族自治区人民医院科研实验中心;
【基金】：广西科技厅项目(桂科攻0632007-1L)
【分类号】：R730.55;R734.2

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