Zhou Chunhui, Yang Fan, Xi Wenjin, Wei Ming, Zheng Guoxu, Wang Wei, Yang Angang, Zhang Jianning, Wen Weihong
Department of Neurosurgery, Navy General Hospital, Beijing 100037; Graduate School, Second Military Medical University, Shanghai 200433, China.
Department of Immunology, School of Basic Medicine, Fourth Military Medical University, Xi'an 710032, China.
Xi Bao Yu Fen Zi Mian Yi Xue Za Zhi. 2014 May;30(5):471-5.
To investigate the effect of RING finger protein 2 (RNF2) knockdown on the biological characteristics and radiosensitivity in glioma U87 cells.
Plasmids containing shRNA targeting RNF2 were transfected into U87 cells. Real-time quantitative PCR (qRT-PCR) and Western blotting were respectively applied to detect the mRNA and protein level of RNF2. MTT assay was used to detect cell proliferation. Cell cycle and apoptosis were measured by flow cytometry combined with annexin V-FITC/PI staining in the control and RNF2 knockdown cells. Apoptosis was also detected after X-ray radiation.
Both shRNAs efficiently inhibited RNF2 expression in U87 cells. Cell proliferation was obviously depressed in RNF2 knockdown cells. The percentage of cells decreased in S phase (shRNA-NC: 27.31 ± 1.35; shRNF2-1: 16.72 ± 2.90; shRNF2-3: 10.35 ± 1.33) and increased in G1 phase (shRNA-NC: 56.13 ± 1.80; shRNF2-1: 76.32 ± 3.11; shRNF2-3: 80.45 ± 2.83). More cell apoptosis was observed in RNF2 knockdown cells. After X ray radiation, the apoptosis rate was significantly raised in RNF2 knockdown cells (shRNA-NC: 20.88 ± 0.64; shRNF2-1: 39.69 ± 0.57; shRNF2-3: 47.82 ± 0.45).
Knockdown of RNF2 can inhibit cell proliferation, induce cell cycle arrest and promote apoptosis in U87 cells. RNF2 knockdown can obviously increase the sensitivity of U87 cells to X ray radiation.
探讨下调指环蛋白2（RNF2）对胶质瘤U87细胞生物学特性及放射敏感性的影响。
将靶向RNF2的短发夹RNA（shRNA）质粒转染至U87细胞。分别采用实时定量聚合酶链反应（qRT-PCR）和蛋白质免疫印迹法检测RNF2的mRNA和蛋白水平。采用MTT法检测细胞增殖情况。通过流式细胞术结合膜联蛋白V-异硫氰酸荧光素/碘化丙啶（annexin V-FITC/PI）染色检测对照细胞和RNF2下调细胞的细胞周期及凋亡情况。在X线照射后也检测了凋亡情况。
两种shRNA均有效抑制了U87细胞中RNF2的表达。RNF2下调细胞的细胞增殖明显受到抑制。S期细胞百分比降低（shRNA-NC：27.31±1.35；shRNF2-1：16.72±2.90；shRNF2-3：10.35±1.33），G1期细胞百分比升高（shRNA-NC：56.13±1.80；shRNF2-1：76.32±3.11；shRNF2-3：80.45±2.83）。RNF2下调细胞中观察到更多的细胞凋亡。X线照射后，RNF2下调细胞的凋亡率显著升高（shRNA-NC：20.88±0.64；shRNF2-1：39.69±0.57；shRNF2-3：47.82±0.45）。
下调RNF2可抑制U87细胞的增殖，诱导细胞周期阻滞并促进其凋亡。下调RNF2可明显提高U87细胞对X线辐射的敏感性。
