Li Yanbo, Guo Caixia, Wang Zhicheng, Gong Pingsheng, Sun Zhiwei, Gong Shouliang
School of Public Health and Family Medicine, Capital Medical University, Beijing, 100069, China.
Key Laboratory of Radiobiology, Ministry of Health, School of Public Health, Jilin University, Changchun, 130021, China.
J Huazhong Univ Sci Technolog Med Sci. 2012 Apr;32(2):167-172. doi: 10.1007/s11596-012-0030-x. Epub 2012 Apr 20.
This study examined the effects of TRAIL-endostatin-based gene-radiotherapy on cellular growth, apoptosis and cell cycle progression in human vascular endothelial cells ECV304 in vitro. The expression of TRAIL and endostatin protein in ECV304 cells was detected by ELISA after the transfection of recombinant plasmid pshuttle-Egr1-shTRAIL-shES and X-ray irradiation. Then MTT assay was used for determining the cellular proliferation, and flow cytometry (FCM) plus Annexin V and propidium iodide (PI) double-staining or PI single-staining were employed for the detection of apoptosis and cell cycle progression. The results showed that expression of TRAIL and endostatin protein exhibited a time- and dose-dependent change in ECV304 cells after pshuttle-Egr1-shTRAIL-shES transfection in conjunction with irradiation. In the TRAIL-endostatin-based single- or double-gene-radiotherapy, the cell viability declined in a time- and dose-dependent manner, the percentage of cells at G(2)/M phase and apoptotic rate was increased, and the percentage of cells at G(0)/G(1) phase was lowered as compared with those receiving radiotherapy alone. Moreover, TRAIL-endostatin-based double-gene-radiotherapy demonstrated better effects on growth inhibition, promotion of apoptosis and induction of cell cycle arrest in ECV304 cells than single-gene-radiotherapy.
本研究检测了基于TRAIL-内皮抑素的基因放疗对人血管内皮细胞ECV304体外细胞生长、凋亡及细胞周期进程的影响。转染重组质粒pshuttle-Egr1-shTRAIL-shES并进行X线照射后,采用ELISA法检测ECV304细胞中TRAIL和内皮抑素蛋白的表达。然后用MTT法检测细胞增殖情况,采用流式细胞术(FCM)结合膜联蛋白V和碘化丙啶(PI)双染或PI单染法检测细胞凋亡及细胞周期进程。结果显示,在pshuttle-Egr1-shTRAIL-shES转染联合照射后,ECV304细胞中TRAIL和内皮抑素蛋白的表达呈现出时间和剂量依赖性变化。在基于TRAIL-内皮抑素的单基因或双基因放疗中,与单纯放疗组相比,细胞活力呈时间和剂量依赖性下降,G(2)/M期细胞百分比和凋亡率升高,G(0)/G(1)期细胞百分比降低。此外,与单基因放疗相比,基于TRAIL-内皮抑素的双基因放疗对ECV304细胞的生长抑制、凋亡促进及细胞周期阻滞诱导作用更强。