解剖学报

›› 2009, Vol. 40 ›› Issue (3): 414-418.doi: 10.3969/j.issn.0529-1356.2009.03.014

• 论著 • 上一篇    下一篇

反义细胞周期蛋白B1重组AAV抑制骨肉瘤U2OS细胞增殖和诱导凋亡

卓建新1 ;李翔2 ;项大业1; 陈日曙1; 张伶3 *    

  1. 1.浙江省乐清市第二人民医院，浙江 温州 325608; 2.浙江省人民医院，杭州 310014; 3.中国人民解放军成都军区总医院肿瘤科，成都 610083
  • 收稿日期:2008-11-17 修回日期:2008-12-03 出版日期:2009-06-06
  • 通讯作者: 张伶

Recombinant adeno-associated virus containing antisense nucleotide against cyclin b1 inhibits osteosarcoma cell U2OS growth and induces apoptosis

  1. 1.Yueqing Second People’s Hospital of Zhejiang, Zhejing Wenzhou 325608, China; 2.Zhejiang Province People’s Hospital, Hangzhou 310014, China; 3.Department of Oncology, General Hospital of Chengdu Military Command, PLA, Chengdu 610083, China
  • Received:2008-11-17 Revised:2008-12-03 Online:2009-06-06
  • Contact: ZHANG Ling

摘要：

关键词: 反义cDNA, 细胞周期蛋白B1, 骨肉瘤, 腺相关病毒, 流式细胞术, 人

Abstract: To investigate the effect of recombinant adenoassociated virus (rAAV) containing antisense nucleotide against cyclin B1 (ASCCNB1) on osteosarcoma cells in vitro. Methods The rAAV was produced by Cotransfecting HEK293 cell line with the rAAV plasmid pd16-95-AS-CCNB1/neo and AAV/Adenoviral helper plasmid pSH3. The rAAV particles expressing ASCCNB1 were purified and concentrated. Subsequently the virus titer was assayed by dotbloting analysis. Then, human osteosarcoma U2OS cells were infected by rAAV at the pfu of 10SUP>5/SUP>-10SUP>6/SUP>/cell titer. CCNB1 expression was detected by Western blotting and RT-PCR methods, human osteosarcoma U2OS cells growth curve was assayed by MTT, cell cycle and apoptosis were detected by FACS. Results The rAAV-AS-CCNB1 plasmid was successfully constructed and the virus titer is up to 1.0×10SUP>11/SUP> v.g./ml. After infected with rAAV EM>in vitro/EM>, U2OS cells growth was inhibited significantly. Both Western blotting and RT-PCR assay showed that CCNB1 expression of U2OS cells was downregulated by rAAV-AS-CCNB1,and cell apoptosis and GSUB>1/SUB> arrest were observed. Conclusion The recombinant adenoassociated virus

Key words: Antisense cDNA, Cyclin B1, Osteosarcoma, Adenoassociated virus, Flow cytometry, Human

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