华西口腔医学杂志 ›› 2012, Vol. 30 ›› Issue (4): 364-367.doi: 10.3969/j.issn.1000-1182.2012.04.008

• 基础研究 • 上一篇    下一篇

RNA干扰酪氨酸激酶受体2对人脐静脉内皮细胞增殖的影响

吴世卿1 曾曙光2 温志欣1 彭细毛2 李玉兰1 卿安蓉1   

  1. 1.南方医科大学附属医院·顺德第一人民医院口腔科, 顺德528300;2.广东省口腔医院口腔颌面外科, 广州510280
  • 收稿日期:2011-04-22 修回日期:2011-09-21 出版日期:2012-08-01 发布日期:2012-08-01
  • 通讯作者: 曾曙光,Tel:020-84426974
  • 作者简介:吴世卿(1982—),女,广东人,住院医师,硕士
  • 基金资助:

    广东省科技计划基金资助项目(2008B030301190)

Tyrosine kinase 2 with immunoglobulin-like and epidermal growth factor homology domains RNA interference inhibit the proliferation of human umbilical vein endothelial cells

Wu Shiqing1, Zeng Shuguang2, Wen Zhixin1, Peng Ximao2, Li Yulan1, Qing Anrong1.     

  1. 1. Dept. of Stomatology, The First People’s Hospital of Shunde, The Affiliated Stomatological Hospital of Southern Medical University, Shunde 528300, China; 2. Dept. of Oral and Maxillofacial Surgery, Guangdong Provincial Stomatological Hospital, Guangzhou 510280, China
  • Received:2011-04-22 Revised:2011-09-21 Online:2012-08-01 Published:2012-08-01

摘要:

目的研究抑制酪氨酸激酶受体2(Tie2)对内皮细胞凋亡和增殖活性的影响。方法采用RNA干扰技术,将含有Tie2基因的特异性短发夹状RNA(shRNA)片段的质粒转染入人脐静脉内皮细胞(HUVECs),采用实时定量逆转录聚合酶链反应和免疫印迹法检测细胞中Tie2 mRNA和蛋白表达的改变;噻唑蓝比色分析(MTT)法检测细胞的生
长情况;显微镜下观察细胞凋亡情况。以转染了pGenesil-hk质粒组为阴性对照,未转染质粒组为空白对照。结果质粒转染入HUVECs后,实验组细胞中Tie2 mRNA和蛋白表达水平较阴性对照组和空白对照组下调,尤以转染后48 h Tie2的mRNA和蛋白表达水平下调更为明显(P<0.05)。MTT检测结果显示:实验组细胞的增殖活性受到明显抑制(P<0.05)。转染48 h后,实验组的细胞凋亡率明显高于两个对照组(P<0.05)。结论RNA干扰技术沉默Tie2基因可致Tie2基因表达下调后诱导HUVECs凋亡,并抑制HUVECs的增殖。

关键词: 酪氨酸激酶受体2, 细胞凋亡, 人脐静脉内皮细胞, 增殖, 抗血管生成

Abstract:

Objective The purpose of this study was to investigate the regulatory role of tyrosine kinase 2 with immunoglobulin-like and epidermal growth factor homology domains(Tie2)on apoptosis and proliferation in the endothelial cells. Methods RNA interference(RNAi) technique was used to silence Tie2 gene expression by transfecting an expression vector containing short hairpin RNA(shRNA) for Tie2 into human umbilical vein endothelial cells(HUVECs). Real time quantitation reverse transcriptase polymerase chain reaction(QRT-PCR) and Western blot were used to monitor Tie2 mRNA, as well as protein expression. The proliferation of HUVECs was examined by methyl thiazolyl tetrazolium(MTT), and the apoptosis was detected under microscope. HUVECs transfected with pGenesil-hk was negative control, and HUVECs transfected with nothing was empty control. Results Tie2 mRNA expression was down-regulated 24 h and 48 h after transfection, and Tie2 protein expression was significantly down-regulated at 24 h and 48 h(P<0.05), especially 48 h after transfection. The apoptosis rate was conspicuously higher in experimental group than in negative control and empty control group after 48 h(P<0.05). The growth monitoring showed that proliferation was also markedly inhibited in experimental group(P<0.05) compared with two control groups. Conclusion Down-regulated expression
of Tie2 by RNAi can promotes apoptosis of HUVECs and has an anti-proliferation activity effect on them.

Key words: tyrosine kinase 2 with immunoglobulin-like and epidermal growth factor homology domains, apoptosis, human umbilical vein endothelial cells, proliferation, anti-angiogenesis