原核表达人MMP-2PEX片段对乳腺癌细胞BICR-H1的生长及转移的抑制

中国生物工程杂志

2011, Vol. 31

Issue (03): 13-17

研究报告

原核表达人MMP-2PEX片段对乳腺癌细胞BICR-H1的生长及转移的抑制

赵威, 韩海勃, 张志谦

北京大学临床肿瘤学院北京肿瘤医院暨北京市肿瘤防治研究所恶性肿瘤发病机制及转化研究教育部重点实验室北京 100142

The Effects of Human PEX, a C-terminal Hemopexin-like Domain of MMP-2, on the Growth and Metastasis of Human Breast Cancer BICR-H1 Cells

ZHAO Wei, HAN Hai-bo, ZHANG Zhi-qian

Key Laboratory of Carcinogenesis and Translational Research(Ministry of Education),School of Oncology,Peking University, Beijing Cancer Hospital & Institute,Beijing 100142, China

全文: PDF(868 KB) HTML

摘要：

目的:克隆表达人源基质金属蛋白酶-2(MMP-2)的C－端类血红素结合域片段PEX,在鸡胚脲囊膜模型上研究PEX对血管发生,乳腺癌BICR-H1的生长及转移抑制作用。方法:构建人源PEX的原核表达载体pET-28a(+)-PEX-His,转化大肠杆菌BL21DE3-pLys,异丙基β-D硫代半乳糖苷(IPTG)诱导PEX蛋白;包涵体蛋白经尿素变性后,通过Ni-NTA 琼脂糖鏊合柱纯化、复性蛋白;观察其对人脐静脉血管内皮细胞增殖和鸡胚脲囊膜血管生长的影响;用带有绿色荧光蛋白(GFP)的腺病毒感染高转移人乳腺癌细胞BICR-H1,接种细胞到10日鸡胚脲囊膜上致瘤,通过静脉注射不同剂量PEX后,观察瘤重、体积和肺转移。结果:5~30μg经原核表达纯化的人PEX蛋白能有效抑制人脐静脉血管内皮细胞增殖能力,表现为时间和剂量依赖效应,并可抑制鸡胚脲囊膜血管发生。BICR-H1的生长及转移在10μg PEX作用时可得到有效抑制,30μg时则完全抑制,未见有肿瘤在接种部位的形成,更未观察到肺脏的转移灶。结论:原核表达纯化的人源PEX具有抑制血管生成、进而抑制乳腺癌BICR-H1细胞的生长和转移作用,是潜在的抗血管发生治疗肿瘤药物,有进一步研发价值。

关键词： PEX; 血管发生; BICR-H1; 乳腺癌

Abstract:

Objective:This study was designed to express human MMP-2 C-terminal hemopexin-like domain (PEX) in E. coli, and then to test its biological effects on angiogenesis, as well as on the growth and metastasis of human breast cancer BICR-H1 cells. Method:PEX was cloned into prokaryotic expression vector pET28a (+) and was induced to express in BL21(DE3)-pLys by IPTG. PEX-His fusion protein, which existed in the inclusion bodies, was denatured by urea method, then was renatured and purified though Ni-NTA agarose beads. The purified PEX-His was further characterized for its biological effects on angiogenesis inhibition, as well as on the suppression of the growth and metastases of human breast cancer BICR-H1 cells by employing the CAM assay. Results: Human PEX expressed in E. coli was able to inhibit the growth of human umbilical vein endothelial cells in a time and dose dependent manner, and could effectively inhibit the CAM angiogenesis. Furthermore, the growth and metastases of BICR-H1 cells were suppressed significantly by injection of 10μg PEX through the vein of chicken embryos, and they were completely disappeared when 30μg PEX was administrated. Conclusion: Human PEX expressed in E. coli is an effective angiongenesis inhibitor. It can inhibit the growth and metastases of BICR-H1 cells, and it might be useful in the treatment of angiongenesis associated diseases such as cancer.

Key words: PEX Angiogenesis BICR-H1 Breast cancer

收稿日期: 2010-11-25 出版日期: 2011-04-01

ZTFLH:

Q786

基金资助:

国家"863"计划(2006AA02A402)、国家自然科学基金(30572160, 30971494)、北京市自然科学基金(7002009,7021001)教育部新世纪优秀人才专项基金(NCET-07-0031)资助项目

通讯作者: 张志谦 E-mail: zlzqzhang@bjmu.edu.cn

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引用本文:

赵威, 韩海勃, 张志谦. 原核表达人MMP-2PEX片段对乳腺癌细胞BICR-H1的生长及转移的抑制[J]. 中国生物工程杂志, 2011, 31(03): 13-17.

ZHAO Wei, HAN Hai-bo, ZHANG Zhi-qian. The Effects of Human PEX, a C-terminal Hemopexin-like Domain of MMP-2, on the Growth and Metastasis of Human Breast Cancer BICR-H1 Cells. China Biotechnology, 2011, 31(03): 13-17.

链接本文:

https://manu60.magtech.com.cn/biotech/CN/ 或 https://manu60.magtech.com.cn/biotech/CN/Y2011/V31/I03/13

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