反义hTERT/PTEN联合基因治疗恶性胶质瘤的体内外研究  

Study on combined gene therapy for malignant gliomas transfected with antisense hTERT/PTEN in vitro and in vivo

作　　者：尤永平[1] 傅震[1] 赵鹏[1] 王存祖[1] 刘宁[1] 鲁艾林[1] 

机构地区：[1]南京医科大学第一附属医院神经外科,210029

出　　处：《中华医学遗传学杂志》2006年第6期605-609,共5页Chinese Journal of Medical Genetics

基　　金：中国博士后科学基金(2003033497);江苏省自然科学基金创新人材项目(BK2004428)~~

摘　　要：目的探讨腺病毒介导的反义hTERT和野生型PTEN联合基因转染对恶性胶质瘤细胞生长的影响。方法用细菌内高效同源重组系统构建的含有hTERT反义序列及野生型PTEN的腺病毒在体内外单独或联合转染恶性胶质瘤细胞系U251,检测肿瘤细胞生长情况、端粒酶活性、蛋白表达、细胞周期变化等。结果在体外试验中单独或联合转染都能明显抑制肿瘤细胞的生长,以联合转染效果最明显,转染后第6天联合转染组的细胞生存率为37.6%,端粒酶活性水平和hTERT蛋白表达水平明显下降,分别为28·8TPG、0.2106,PTEN蛋白表达水平上升为0.9630;在体内试验中肿瘤生长也明显减慢。结论腺病毒介导的反义hTERT和野生型PTEN联合转染能明显抑制恶性胶质瘤细胞的生长。Objective To study inhibitory efficacy of combined gene therapy for malignant ghomas transfected with antisense human telomerase reverse transcriptase （hTERT）/PTEN in vitro and in vivo. Methods To construct two adenovirus recons which contained antisense hTERT and wild-type PTEN respectively with high performance homologous recombination system in bacteria. The two adenovirus recons were transfected into U251 human mahgnant ghoma cells eombinedly or respectively in vitro and in vivo. U251 cell proliferation in vitro was determined by MTT assay and flow cytometry, tumor growth in vivo was measured by the volume of ghoma in nude mice. Telomerase activity was detected by telomerie repeat amplification protocol （TRAP） assay. Expression of hTERT and PTEN protein was detected by Western blotting methods. Reeults After transfection in vitro, the growth of U251 cells was inhibited significantly. The inhibitory effect was time-dependent. The strongest inhibition was observed in combined transfection group, at the 6th day, the survival rate was 37.6%, telomerase activity （only 28.8TPG） was inhibited significantly, hTERT protein expression was inhibited significantly too, which was 0.2106, but PTEN protein expression was increased significantly, which was 0.9630. In vivo, the growth of tumors was also effectively inhibited. Conclusion Growth of malignant glioma cells is effectively inhibited after transfection with combined antisense hTERT and PTEN in vitro and in vivo.

关 键 词：反义hTERT基因 野生型PTEN基因 联合基因治疗 恶性胶质瘤 

分 类 号：R739.4[医药卫生—肿瘤]