贵州医科大学学报

2017, v.42;No.199(04) 377-382+388

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靶向沉默HIF-1α和Survivin基因对鼻咽癌裸鼠移植瘤放疗敏感性的影响
Effect of Targeted Silencing HIF-1α and Survivin Gene on Radiation Sensitivity of Transplanted Nasopharyngeal Carcinoma in Nude Mice

向少龙;刘娟娟;赵艳;赵厚育;
XIANG Shaolong;LIU Juanjuan;ZHAO Yan;ZHAO Houyu;Guizhou Medical University;The Key Laboratory of Molecular Biology,Guizhou Medical University;Department of Oterhinolaryngology,the Affiliated Hospital of Guizhou Medical University;

摘要(Abstract):

目的:探讨RNAi靶向沉默人鼻咽癌细胞株CNE-2细胞缺氧诱导因子-1α(HIF-1α)和/或生存素(Survivin)基因对裸鼠移植瘤放疗敏感性的影响。方法:设计合成靶向沉默HIF-1α和/或Survivin的microRNA干扰质粒,采用脂质体转染法将HIF-1α和/或Survivin的microRNA干扰质粒转染到CNE-2细胞;取35只4~6周龄的雌性BALA/c裸鼠,随机均分为HIF-1α干扰组、Survivin干扰组、HIF-1α&Survivin干扰组(联合干扰组)、空载组和阴性对照组;分别给予相应转染后的CNE-2细胞注射至裸鼠右侧腋窝皮下,待各组裸鼠成瘤后,给予5Gy X-射线照射,每隔3 d放疗1次,共15Gy X-射线,观察各组裸鼠放疗前后的肿瘤体积,照射结束3 d后处死裸鼠,测量瘤体体积,计算肿瘤体积增加率;RT-q PCR法检测瘤体组织中的HIF-1α和Survivin mRNA表达水平,Western Blot法检测HIF-1α和Survivin蛋白表达水平,TUNEL试剂盒检测鼻咽癌裸鼠移植瘤中细胞凋亡。结果:HIF-1α干扰组、Survivin干扰组与联合干扰组经过放射治疗后肿瘤体积增加率明显小于空载体组与阴性对照组(P<0.05);HIF-1α干扰组、Survivin干扰组与联合干扰组瘤体组织中HIF-1α、Survivin mRNA和蛋白质表达水平显著低于空载组与阴性对照组,细胞凋亡率高于空载组与阴性对照组(P<0.05);HIF-1α干扰组、Survivin干扰组及联合干扰组瘤体组织中HIF-1α、Survivin mRNA和蛋白质表达水平及细胞凋亡率比较,差异无统计学意义(P>0.05)。结论:单独干扰HIF-1α、Survivin基因与联合干扰HIF-1α和Survivin基因均能够增加鼻咽癌裸鼠移植瘤对放疗敏感性,但联合干扰并没有进一步提高鼻咽癌裸鼠移植瘤放疗敏感性。
Objective: To investigate the effects of Hypoxia inducible factor 1-alpha( HIF-1α) and/or Survivin interference in human nasopharyngeal carcinoma cell line CNE-2 on the radiotherapy sensitivity of xenografted tumors in nude mice. Methods: The recombinant plasmids for HIF-1α and/or Survivin interference were constructed. Then the plasmids were transfected into CNE-2 cells using lipofection; 35 female nude mice aged 4 to 6 weeks were randomly divided into HIF-1α interference group,Survivin interference group,HIF-1α & Survivin interference group,the empty carrier group,and negative control group in equal numbers. CNE-2 cells after transfection were injected to nude mice respectively. Once xenografted tumors formed,each mouse would accept 5 Gy of X-ray radiations in every three days for a total 15 Gy of X-ray radiations. The volume of tumors would be observed at different times. The mice would be put to death three days later after radiation. Then the volume of tumors would be measured and the radiotherapy sensitivity would be calculated. Moreover,the mRNA expression levels and protein levels of HIF-1α and Survivin in the xenografted tumors would be detected by RT-q PCR and Western blotting respectively. The cell apoptosis in xenografted tumors would be evaluated using TUNEL kit. Results: After radiation therapy,the increase rates of tumor volume ration in HIF-1α interference group,Survivin interference group and HIF-1α & Survivin interferences group were significantly lower than that in empty carrier group and the negative control group( P < 0. 05).Furthermore,the mRNA and protein expression levels of HIF-1α,Survivin mRNA and protein expression level in HIF-1α,Survivin interference groups and HIF-1α & Survivin interference group were significantly lower than that in the empty carrier group and the negative control group( P < 0. 05). The cell apoptosis of tumors in HIF-1α and/or Survivin interference groups were significantly higher than that in the empty carrier group and the negative control group( P < 0. 05). However,there was no statistical significance for the mRNA and protein expression levels of HIF-1α and Survivin and the cell apoptosis of tumors among HIF-1α interference group,Survivin interference group and HIF-1α & Survivin interferences group( P > 0. 05). Conclusion: Independently HIF-1α interference,Survivin interference or HIF-1α & Survivin interferences could increase the radiotherapy sensitivity of Nasopharyngeal carcinoma xenografted tumors in nude mice. However,HIF-1α & Survivin interferences did not further improve radiotherapy sensitivity of Nasopharyngeal carcinoma xenografted tumors.

关键词(KeyWords): 鼻咽肿瘤;缺氧诱导因子-1α;生存素;放射疗法,计算机辅助;放疗敏感性
nasopharyngeal carcinoma;hypoxia-inducible factor-1a;Survivin;radiotherapy;computer-assisted;radiotherapy sensitivity

Abstract:

Keywords:

基金项目(Foundation): 国家自然科学基金地区科学基金项目(81260353)

作者(Author): 向少龙;刘娟娟;赵艳;赵厚育;
XIANG Shaolong;LIU Juanjuan;ZHAO Yan;ZHAO Houyu;Guizhou Medical University;The Key Laboratory of Molecular Biology,Guizhou Medical University;Department of Oterhinolaryngology,the Affiliated Hospital of Guizhou Medical University;

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DOI: 10.19367/j.cnki.1000-2707.2017.04.004

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