Chao Teng-Fei, Xiong Hui-Hua, Liu Wei, Chen Yang, Zhang Jia-Xuan
Cancer Centre, Huazhong University of Science and Technology, Wuhan, 430030, China.
Department of Neurosurgery, Beijing Tiantan Hospital, Capital Medical University, Beijing, 100050, China.
J Huazhong Univ Sci Technolog Med Sci. 2013 Aug;33(4):525-529. doi: 10.1007/s11596-013-1153-4. Epub 2013 Aug 1.
The purpose of this study was to investigate the molecular mechanism by which miR-21 and its target genes mediate radiation resistance of glioblastoma cells. Real-time PCR was employed to detect miR-21 expression in normal brain tissues, glioblastoma tissues and glioblastoma cell lines (A172, T98G and U87MG). T98G cells were transfected with anti-miR-21 oligonucleotides, or plasmids containing PDCD4 or hMSH2 (PDCD4-pcDNA3 and hMSH2-pcDNA3). The survival curve was obtained to investigate the sensitivity of T98G cells to radiation. Cell apoptosis was measured by using the Caspase-3/7 kit and cell cycle by flow cytometry. Western blotting was performed to detect the expression of hMSH2 and PDCD4 in miR-21-inhibiting T98G cells. The results showed that miR-21 expression in glioblastoma cells and tissues was conversely associated with the radiation sensitivity. Over-expression of miR-21 resulted in radiation resistance, while knockdown of miR-21 led to higher sensitivity of glioblastma cells to radiation. After miR-21 knockdown, the apoptosis of T98G cells was significantly increased and the G(2) phase arrest was more significant. In addition, miR-21 knockdown increased the expression of endogenous PDCD4 and hMSH2, which contributed to the apoptosis and G(2) arrest of T98G cells. The findings suggested that miR-21 may mediate the resistance of glioblastoma cells against radiation via its target genes PDCD4 and hMSH2. MiR-21 and its target genes may be used as potential molecular targets for clinical radiotherapy sensitization in the future.
本研究旨在探究miR-21及其靶基因介导胶质母细胞瘤细胞辐射抗性的分子机制。采用实时定量聚合酶链反应检测正常脑组织、胶质母细胞瘤组织及胶质母细胞瘤细胞系（A172、T98G和U87MG）中miR-21的表达。用抗miR-21寡核苷酸或含PDCD4或hMSH2的质粒（PDCD4-pcDNA3和hMSH2-pcDNA3）转染T98G细胞。绘制生存曲线以研究T98G细胞对辐射的敏感性。使用Caspase-3/7试剂盒检测细胞凋亡，通过流式细胞术检测细胞周期。采用蛋白质免疫印迹法检测miR-21抑制的T98G细胞中hMSH2和PDCD4的表达。结果显示，胶质母细胞瘤细胞和组织中miR-21的表达与辐射敏感性呈负相关。miR-21过表达导致辐射抗性，而敲低miR-21导致胶质母细胞瘤细胞对辐射更敏感。敲低miR-21后，T98G细胞的凋亡显著增加，G(2)期阻滞更明显。此外，敲低miR-21增加了内源性PDCD4和hMSH2的表达，这有助于T98G细胞的凋亡和G(2)期阻滞。研究结果表明，miR-21可能通过其靶基因PDCD4和hMSH2介导胶质母细胞瘤细胞的辐射抗性。miR-21及其靶基因未来可能作为临床放疗增敏的潜在分子靶点。
