[1]杜虎,吴小候,罗春丽,等.shRNA沉默survivin基因对人膀胱移行细胞癌T24细胞生物学行为的影响[J].第三军医大学学报,2009,31(02):124-127.
 DU Hu,WU Xiao-hou,LUO Chun-li,et al.Inhibitory effects of shRNA-silencing endogenous survivin gene on T24 bladder carcinoma cells[J].J Third Mil Med Univ,2009,31(02):124-127.
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《第三军医大学学报》[ISSN:1000-5404/CN:51-1095/R]

卷:
31卷
期数:
2009年第02期
页码:
124-127
栏目:
论著
出版日期:
2009-01-30

文章信息/Info

Title:
Inhibitory effects of shRNA-silencing endogenous survivin gene on T24 bladder carcinoma cells
作者:
杜虎吴小候罗春丽刘川曾柯何云锋尹志康
重庆医科大学:附属第一医院泌尿外科,检验系
Author(s):
DU Hu WU Xiao-hou LUO Chun-li LIU Chuan ZENG Ke HE Yun-feng YIN Zhi-kang
Department of Urinary Surgery, First Affiliated Hospital,Falculty of Medical Laboratory, Chongqing Medical University, Chongqing 400016, China
关键词:
survivinshRNA肿瘤侵袭膀胱肿瘤
Keywords:
survivin short hairpin RNA neoplasm invasiveness bladder neoplasm
分类号:
R73-354;R730.23;R737.14
文献标志码:
A
摘要:
目的    构建survivin特异性短发夹RNA(shRNA)的表达载体,探讨其对人膀胱移行细胞癌T24细胞的生物学行为的影响。    方法    将构建survivin特异shRNA表达载体转染T24细胞,RT-PCR、Western blot检测survivin mRNA及其蛋白的表达水平;通过细胞生长曲线、随机运动实验、Matrigel穿膜实验检测肿瘤细胞生长、运动及体外侵袭能力。    结果    经EcoRⅠ和HindⅢ双酶切和测序鉴定,成功筛选含目的 DNA片段的重组载体pshRNA-survivin2,并成功转染T24细胞;转染pshRNA-survivin2后可显著抑制细胞中survivin mRNA及其蛋白的表达,抑制率分别为61.73%和73.27%;pshRNA-survivin2组的细胞侵袭力与运动能力均有明显的下降,第3、5天时肿瘤细胞生长抑制率分别为59.13%、83.86%。    结论    应用pTZU6+1质粒载体构建survivin的shRNA表达载体,能有效抑制其survivin mRNA及其蛋白的表达,并可抑制T24细胞的增殖、运动及体外侵袭能力。
Abstract:
Objective    To construct the plasmid containing short hairpin RNA (shRNA) of survivin to suppress the expression of endogenous survivin gene in T24 cells.     Methods    The recombinant plasmid  pshRNA-survivin2 expression construct was confirmed by EcoR Ⅰ and HindⅢ double digestion and by sequencing. The plasmid pshRNA-survivin2 was stably transfected into transitional cell carcinoma of the bladder (TCCB) cells T24. Then survivin mRNA and protein expressions in the transfected T24 cells were detected by semi-quantitative reverse transcriptase-polymerase chain reaction (RT-PCR) and Western blotting. The biological effects of the plasmid pshRNA-survivin2 on T24 cells were evaluated through the detection of their anchorage-independent growth and in vitro invasion by cell migration assay and tranwell chamber assay.     Results    The expression of survivin in transfected T24 cells was markedly depressed at both mRNA and protein levels (61.73% and 73.37%, respectively) as compared with control. Anchorage-independent growth assay showed the growth of tumor cells was retarded. The inhibitory percentage of transfected T24 cells was respectively 59.13% and 83.86% on the third day and the fifth day. Transwell chamber assay showed the invasion ability of T24 cells was inhibited significantly.     Conclusion    The pshRNA-survivin2 expression plasmid, constructed from plasmid pTZU6+1, can be successfully transfected into TCCB cell line T24 and can effectively inhibit the expression of survivin mRNA and protein. Therefore, the proliferation, migration and invasion of T24 cells transfected with pshRNA-survivin2 expression plasmid are inhibited. RNAi targeting survivin has a potential value in gene therapy of bladder cancer.

参考文献/References:

杜虎, 吴小候, 罗春丽, 等. shRNA沉默survivin基因对人膀胱移行细胞癌T24细胞生物学行为的影响[J]. 第三军医大学学报, 2009, 31(2):124-127.

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更新日期/Last Update: 2009-01-08