华西口腔医学杂志

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表皮生长因子受体反义cDNA干预喉癌细胞信号转导的实验研究

鲜均明 周光耀 梁传余 刘世喜   

  1. 四川大学华西医院耳鼻咽喉科, 四川成都610041
  • 收稿日期:2007-12-25 修回日期:2007-12-25 出版日期:2007-12-20 发布日期:2007-12-20
  • 通讯作者: 刘世喜, Tel: 13982063567
  • 作者简介:鲜均明( 1968-),男,四川人,主治医师,博士
  • 基金资助:

    国家自然科学基金资助项目( 0040205401078);四川省青年科技基金资助项目( 04ZQ026- 024)

Study on inter fer ence ther apy induced by epidermal growth factor r eceptor - antisense cDNA in signal tr ansduction of laryngeal squamous cell car cinoma

XIAN Jun- ming, ZHOU Guang- yao, LIANG Chuan- yu, LIU Shixi   

  1. Dept. of Otolaryngology, West China Hospital, Sichuan University, Chengdu 610041, China)
  • Received:2007-12-25 Revised:2007-12-25 Online:2007-12-20 Published:2007-12-20
  • Contact: LIU Shixi, Tel: 13982063567

摘要:

目的探讨表皮生长因子受体( EGFR)反义cDNA在体外对人喉癌Hep- 2细胞信号转导的干预作用。方法以缺陷重组腺病毒为载体,构建EGFR正、反义cDNA的重组腺病毒,并在人胚肾母细胞( HEK293)中包装、纯化。将已纯化的EGFR cDNA重组腺病毒在体外转染人Hep- 2喉癌细胞,采用MTT法检测重组腺病毒对Hep- 2细胞的生长抑制作用、流式细胞术检测细胞周期DNA表达的变化、Western blot方法检测Hep- 2细胞内EGFR蛋白的表达。结果成功地构建并制备了高滴度的EGFR cDNA 1 032 bp片段的正、反义重组腺病毒。同时,它们能被高效地转移到Hep- 2细胞内,其中反义重组腺病毒能够抑制Hep- 2细胞的增殖和EGFR蛋白的表达。结论EGFR反义cDNA能有效地干预人喉癌Hep- 2细胞的细胞周期信号转导机制,从而抑制肿瘤细胞的增殖。

关键词: 下颌骨髁突, 关节软骨, 骨形成蛋白-2, 咬合, 表皮生长因子受体, 信号转导, 喉癌

Abstract:

Objective To investigate effect of interference therapy induced by epidermal growth factor receptor ( EGFR)- antisense cDNA in signal transduction of Hep - 2 laryngeal squamous cell carcinoma in vitro. Methods AdEasy Vector System was used to construct the recombinant adenovirus vector sense/antisense- pAdEasy- EGFR. The recombinant adenovirus vector introduced EGFR- sense/antisense cDNA fragment into HEK293 cell. The purified recombinant adenovirus sense/antisense- pAdEasy- EGFR transfected Hep- 2 cells in vitro. The inhibition of EGFR protein expression and proliferation of Hep- 2 cells, the changes of cell cycle and DNA content in Hep- 2 cells were examined by MTT, Western blot analysis, flow cytometry essay, and immunocytochemistry respectively. Results The higher titre sense and antisense mRNA expression recombinant adenovirus containing 1 032 bp EGFR- cDNA was constructed and prepared successfully. When antisense- pAdEasy- EGFR was transferred into Hep- 2 cells the inhibition of cell proliferation and EGFR protein expression in Hep- 2 cells were investigated effectively. Conclusion The antisense- pAdEasy- EGFR effectively interfere the Hep- 2 signal transduction pathway and induce apoptosis which results in inhibiting proliferation of laryngeal squamous cell carcinoma.

Key words: mandibularcondyle;articularcartilage;bonemorphogeneticprotein-2;occlusion;, epidermal growth factor receptor, signal transduction, laryngeal squamous cell carcinoma