干扰Wrap53基因调控人骨肉瘤U2OS细胞放射敏感性实验研究  

Experimental research of inhibition of Wrap53gene regulating the radiosensitivity of osteosarcoma U2OS cells

作　　者：竺炎 吴传高[1] 刘少平[1] 邱惠[1] 谢丛华[1] 周福祥[1] 

机构地区：[1]武汉大学中南医院放化疗科.肿瘤生物学行为湖北省重点实验室.湖北省肿瘤医学临床研究中心,湖北武汉430071

出　　处：《中华肿瘤防治杂志》2013年第21期1657-1660,共4页Chinese Journal of Cancer Prevention and Treatment

基　　金：湖北省杰出青年基金(2011CDA095)

摘　　要：目的:探讨干扰Wrap53基因表达对肿瘤细胞放射敏感性的作用及其分子机制。方法:构建针对Wrap53的干扰质粒;脂质体转染法转染至p53野生型(wtp53)的人骨肉瘤U2OS细胞;RT-PCR和蛋白质印迹法检测Wrap53和wtp53基因的mRNA和蛋白表达;克隆形成实验测定放射敏感性参数;流式细胞术(flow cytometry,FCM)分析细胞周期变化。结果:Wrap53的特异性shRNA质粒转染U2OS细胞后,细胞内Wrap53和wtp53的mRNA和蛋白表达水平均受到明显抑制;Wrap53基因干扰组U2OS细胞的D0值为0.44±0.01,明显高于阴性对照组0.37±0.01,P=0.001,Wrap53基因干扰组SF2值为0.72±0.10,高于阴性对照组0.44±0.02,P=0.047;Wrap53干扰细胞在放射后16h(P=0.014)和24h(P=0.011)的G2/M期阻滞更加明显,显著高于对照组;但Wrap53干扰细胞在放射后出现G1期阻滞减少,以放射后24h最为显著,P=0.001。结论:干扰Wrap53基因表达降低了U2OS细胞的放射线敏感性,该作用可能与下调wtp53表达和诱导细胞放射后G2/M期阻滞有关。OBJECTIVE To explore the effect of interference Wrap53 gene expression on the radiosensitivity of tumor cells,and its molecular mechanism. METHODS： The specific Wrap53 interference plasmid was designed and trans fected into wild type p53 osteosarcoma U2OS cells. The mRNA and protein expressions of Wrap53 and wtp53 were ana lyzed by RT-PCR and Western blotting,the radiosensitivity parameters were measured by clonogenic assay,and cell cycle progression was detected by flow cytometry. RESULTS： The specific Wrap53 shRNA plasmid was successfully designed and was transfected into U2OS cells,which inhibited the expression of Wrap53 and wtp53 in mRNA and protein levels. The Do and SF2 of Wrap53 interferen.ce group were 0.44±0.01 and 0.72 ± 0. 10 respectively, which were significantly higher than those in the negative control group （Do =0. 374-0.01,P=0. 001 SF2=0. 444-0.02,P=0. 047）. Wrap53 in-terference group significantly occurred G/M arrest after radiation in 16 hours and 24 hours respectively, in comparison with the control group （P= 0. 014, P= 0. 011）. In contrast to this, decrease of Gl-arrested cells was more prominent in Wrap53 interference group after radiation in 24 hours （P = 0.001）. CONCLUSION.. Wrap53 interference decreases the radi- osensitivity of U2OS cells,which may be related to downregulation of wtp53 expression and induction of G2/M arrest.

关 键 词：Wrap53 RNAI WTP53 放射敏感性 细胞周期 

分 类 号：R738.1[医药卫生—肿瘤]