靶向敲降Sox4基因抑制宣威地区女性肺癌细胞系XWLC-05裸鼠移植瘤的生长  

Targeted Sox4 gene-knock-down suppresses xenograft growth of Xuanwei female lung cancer cell lineXWLC-05 cells in nude mice

作　　者：周永春[1] 王熙才[1] 陈艳[1] 黄尤光[1] 刘馨[1] 金从国[1] 陈晓群[1] 李佳[1] 黄云超[1] 

机构地区：[1]昆明医科大学第三附属医院云南省肿瘤研究所,650118

出　　处：《中华肿瘤杂志》2014年第1期11-16,共6页Chinese Journal of Oncology

基　　金：国家自然科学基金重点项目（U1202224）;云南省应用基础研究面上项目（2010CD081）

摘　　要：目的探讨抑制宣威地区女性肺癌细胞系XWLC-05中转录因子Sox4的表达对裸鼠皮下移植瘤生长的影响。方法构建靶向抑制Sox4基因的shRNA干扰质粒pGFP-V-RS-Sox4 shRNA，并转染XWLC-05细胞；采用实时荧光定量PCR和Western blot检测Sox4的表达水平；将稳定转染重组质粒的XWLC-05细胞接种于裸小鼠皮下，建立移植瘤模型；观测裸小鼠一般情况、成瘤情况和肿瘤生长情况，计算肿瘤抑制率；对小鼠行CT扫描以观察肿瘤的转移情况；免疫组化染色检测移植瘤中Sox4和ki-67的表达。结果成功构建了高效抑制Sox4基因表达的pGFP-V-RS-Sox4 shRNA重组干扰质粒，并筛选出稳定转染的XWLC-05细胞。除接种生理盐水组外，接种稳定转染干扰质粒pGFP-V-RS-Sox4 shRNA的XWLC-05细胞组、接种稳定转染pGFP-V-RS-seraln shRNA的XWLC-05细胞组和接种亲本XWLC-05细胞组小鼠的成瘤率均为100％。空白对照组（不加任何试剂的亲本细胞）、阴性对照组（转染质粒pGFP-V-RS-scram shRNA）和实验组（转染质粒pGFP-V-RS-Sox4 shRNA）小鼠的体重均有不同程度的增长，但差异无统计学意义（P〉0．05）。与空白对照组和阴性对照组比较，抑制Sox4基因表达后的实验组小鼠移植瘤的增长趋势明显受到抑制，差异均有统计学意义（均P〈0．05）。实验组离体移植瘤体积为（2．304±0．34）cm^3，与阴性对照组[（3．994±0．45）cm^3]和空白对照组[（4．03±0．42）cm^3]比较，差异均有统计学意义（均P〈0．05）。实验组离体移植瘤重量为（0．864±0．14）g，低于阴性对照组[（1．844±0．27）g]和空白对照组[（1．864±0．22）g]，差异均有统计学意义（均P〈0．05）。Sox4和ki-67蛋自主要表达于细胞核，实验组移植瘤细胞中Sox4蛋白的表达低于阴性对照组和空白对照组，差异均有统计学意义（均P〈0．05）。影像学和病理学检查结果显示，在实验期内成瘤小鼠Objective To study the effect of targeted Sox4 gene-knock-down on the growth of xenografts of Xuanwei female lung cancer cell line XWLC-05 cells in nude mice. Methods Recombinant plasmid pGFP-V-RS-Sox4 shRNA was constructed and transfected into XWLC-05 cells. Real-time quantitative PCR and Western blot were applied to confirm the effect of Sox4 gene-knock-down. XWLC-05 cells stably transfected with the plasmids were inoculated into nude mice to establish the xenograft model. The nude mouse status, tumor formation and tumor growth were observed, and the tumor inhibition rate was calculated. CT scan was performed to assess the metastasis of xenografts. Immunohistochemical staining was applied to detect Sox4 and ki-67 protein expression. Results Recombinant plasmid pGFP-V-RS-A-Sox4 shRNA which can effectively knocking-down Sox4 gene was successfully constructed and the stable transfected cells were selected by puromycin-screening. The success rate of tumor cell inoculation was 100% in the mice of all groups except those inoculated with saline. The body weight of all mice inoculated withparental XWLC-05 cells （ blank control）, pGFP-V-RS-scram shRNA trsfeeted XWLC-05 cells （ negative control）, and pGFP-V-RS-Sox4 shRNA transfected XWLC-05 cells was increased to a varying degree, but there was no significant difference among the groups （P 〉 0.05 ）. The growth of xenografts was significantly inhibited after silencing the Sox4 gene expression when compared with that of the blank and negative controls （P 〈 0.05 ）. The volume of removed tumors of the Sox4 gene-inhibited mice was （2.30 ± 0. 34 ） cm^3, significantly smaller than that of the negative control （ 3.99 ± 0.45） cm^3 and the blank control （4.03 ± 0.42） cm^3 （P 〈 0.05 ）. The weight of removed tumors of Sox4 gene-inhibited mice was （0.86 ± 0. 14 ） g, significantly lower than that of the negative control （ 1.84 ± 0.27 ） g and blank control （ 1.86 ± 0.22） g, （P 〈0.05）. Immunohistochemical staining showed that

关 键 词：肺肿瘤 肿瘤移植 Sox4 基因 小鼠 

分 类 号：R734.2[医药卫生—肿瘤]