放射诱导启动子介导p53上调凋亡调控因子基因靶向治疗舌鳞状细胞癌裸鼠移植瘤

Radiation-inducible promoters mediated PUMA gene in the treatment of xenograft of human tongue squamous in naked mice

目的探讨放射诱导启动子介导p53上调凋亡调控因子(PUMA)基因靶向治疗舌鳞状细胞癌(简称舌鳞癌)裸鼠移植瘤的疗效。方法构建放射诱导启动子介导PUMA基因表达的真核表达质粒pcDNA(+)3.1/E-PUMA,建立人舌鳞癌裸鼠移植瘤模型,以脂质体为载体进行瘤内转染,辅以3Gy放疗,描述肿瘤生长曲线,反转录聚合酶链反应(RT-PCR)检测PUMA的mRNA水平,免疫组化检测细胞核增殖抗原的表达,原位末端标记法检测细胞凋亡。数据分析采用SPSS 11.0统计软件,两样本均数比较采用χ2检验。结果放射诱导启动子介导的PUMA基因转染对舌鳞癌裸鼠移植瘤生长有明显抑制作用,诱导放疗显著增强了PUMA的mRNA表达水平,RT-PCR检测发现其电泳条带灰度值从(0.325±0.075)上调至(0.371±0.096),P<0.01;诱导放疗可显著提高疗效,增殖指数由32.27%下降至22.77%,P<0.01;凋亡指数从14.43%上升为27.98%,P<0.01。结论放射诱导启动子作为基因治疗分子开关可介导PUMA基因在人舌鳞癌裸鼠移植瘤中靶向表达促进细胞凋亡,为舌鳞癌放射基因治疗提供了新思路。

Objective To evluate the therapeutic effect of PUMA gene mediated by radiation- inducible promoters in the treatment of xenografi of human tongue squamous carcinoma in nude mice. Methods The recombinant pcDNA3.1 （＋）/E-PUMA was constructed, and the model of transplanted Tca8113 cell line in naked mice was founded. The plasmids peDNA（＋）3.1/E-PUMA were transfected into tumors by lipofectamine. 24 h later, the tumors were treated by 3 Gy irradiations. Animals were monitored for volume of tumors at regular intervals. PUMA mRNA was detected by RT-PCR. PCNA and Apoptosis were detected by immuno-histiochemical method and in situ end-labeling respectively. The data was analyzed by SPSS 11.0 software package for chi-square test. Results The tumors were suppressed obviously by radiation-inducible promoters mediated PUMA gene. The 3 Gy radiation could up-regulated the PUMA expression in xenograft of human tongue squamous in naked mice. There was significant difference between the apoptosis proliferation index （PI） or index （AI） in tumors between therapy group and control groups （P 〈 0.01 ）. Conclusions The radiation-inducible promoters can be served as a molecular switch to upgrade PUMA expression in transplanted human tongue squamous carcinoma in naked mice, and low dose induction-radiation can significantly improve the therapeutic efficiency. It is possible to validate this strategy in the targeted treatment of tongue suqmous cell carcinoma.