水流动力学注射方法建立肝内胆管癌动物模型

朱方全; 秦中强; 李红俊; 陈勇; 李永海; 谈燚

doi:10.3760/cma.j.issn.1001-9030.2017.04.055

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中华实验外科杂志

• 动物模型 •

ENGLISH ABSTRACT

水流动力学注射方法建立肝内胆管癌动物模型

作者及单位信息

出版日期： 2017 -04 -08 ·

DOI： 10.3760/cma.j.issn.1001-9030.2017.04.055

One mouse model of intrahepatic cholangiocarcinoma by hydrodynamic injection of sleeping beauty vectors

Authors Info & Affiliations

Zhu Fangquan

Department of Hepatobiliary Surgery, the First Affiliated Hospital, Bengbu Medical College, Bengbu 233004, China

Qin Zhongqiang

Department of Hepatobiliary Surgery, the First Affiliated Hospital, Bengbu Medical College, Bengbu 233004, China

Li Hongjun

Laboratory of Pathophysiology, Bengbu Mmedical College, Bengbu 233030, China

Chen Yong

Laboratory of Pathogeny Biology, Bengbu Medical College, Bengbu 233030, China

Li Yonghai

Laboratory of Pathophysiology, Bengbu Mmedical College, Bengbu 233030, China

Tan Yi

Department of Hepatobiliary Surgery, the First Affiliated Hospital, Bengbu Medical College, Bengbu 233004, China

Published： 2017 -04 -08 ·

DOI： 10.3760/cma.j.issn.1001-9030.2017.04.055

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摘要

目的建立一种小鼠肝内胆管癌动物模型。

方法采用水流动力学注射方法将表达Notch1基因胞内区活性形式pT3-EF1a-Notch1分子的胞内结构域(NICD1)转座子和表达myr-蛋白激酶B(Akt)基因转座子质粒溶液通过小鼠尾静脉注射至肝脏。注射生理盐水作为对照。注射4周后处死，病理学检查肿瘤发生情况，免疫组织化学、免疫荧光检测细胞角蛋白19(CK19)及标签蛋白流感病毒血凝素蛋白表面抗原决定簇衍生的蛋白标签序列(HA)标记的Akt表达。

结果实验组小鼠均形成肿瘤，成癌率为100%(14/14)；病理学检测为胆管细胞性肝癌，免疫组织化学检测CK19，14例中有9例强阳性表达(64.3%)，证实瘤细胞是胆管细胞性肝癌。同样，在瘤细胞中检测到HA强阳性表达率为42.9%(6/14)；免疫荧光双标记CK19和HA升高。

结论水流动力学注射方法成功构建肝内胆管癌小鼠模型且诱癌时间短，方法简单，重复性好。

关键词：肝内胆管癌;水流动力学注射;转座子;模型，动物

ABSTRACT

ObjectiveTo establish one mouse model of intrahepatic cholangiocarcinoma (ICC) for liver cancer reseach.

MethodsThe transposon vectors of sleeping beauty were constructed harboring the activated protein kinase B (myr-Akt) or the activated Notch1 intracellular domain (NICD1), respectively. These vectors were hydrodynamic injected through tail veil. Mice injected with saline was used as the control. Mice were sacrificed around 4 weeks. Tumors were featured by macro-and micro-histological observation. Immunohistological staining and fluorescence staing were used for cytokeratin (CK)19 and Akt cellular expression detected by the sequence Tag of hemagglutinin surface antigen determinant of influenza virus (HA)-tag.

ResultsCompared with the control, mice in the experimental group progressed to tumor with the rate of 100% (14/14). The features of macro-and micro-histological observation indicate the formation of intraheptic cholangiocarcinoma. High percentage (64.3%) of CK19-positive staining in the malignant cells of liver confirmed the tumorigenesis of ICC. Similarly, high rate of HA-positive(42.9%) expression was observed in malignant cells. Colocalization of CK19 and HA staining by immunofluorescence staining also verify high expression.

ConclusionWe provide one method to generate ICC murine model with the advantages of easy-manupilation and time-saving and recapitulation.

KEYWORDS： Intrahepatic cholangiocarcinoma;Hydrodynamic Injection;Sleeping beauty transposons;Mouse, model

Corresponding author: Li Yonghai, Email: mocdef.labiamgtnemirepxecmbb

Corresponding author:Tan Yi, Email: mocdef.aabnis7002iynatrotcod

FUNDING:

National Natural Science Foundation of China (81472656); Innovation Program of Bengbu Medical College (Byycx1539)

COPYRIGHTS:

No content published by the journals of Chinese Medical Association may be reproduced or abridged without authorization. Please do not use or copy the layout and design of the journals without permission.

All articles published represent the opinions of the authors, and do not reflect the official policy of the Chinese Medical Association or the Editorial Board, unless this is clearly specified.

引用本文

朱方全,秦中强,李红俊,等. 水流动力学注射方法建立肝内胆管癌动物模型[J]. 中华实验外科杂志,2017,34(4)：703-705.

DOI:10.3760/cma.j.issn.1001-9030.2017.04.055

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未经授权，不得转载、摘编本刊文章，不得使用本刊的版式设计。

除非特别声明，本刊刊出的所有文章不代表中华医学会和本刊编委会的观点。

肝内胆管癌(ICC)是指左右肝管汇合部以上的胆管上皮细胞起源的罕见的肝脏恶性肿瘤，其恶性程度高、症状隐匿，预后差。目前建立肝内胆管癌动物模型多用化学诱导的方法，有文献报道氨基比林和亚硝酸钠溶液作为叙利亚地鼠的饮用水，每周饮用6 d，连续饮用24周，可诱发肝内胆管癌，成癌率为50% ^{[
1
]}。梁平等 ^{[
2
]}报道手术结扎胆总管联合皮下注射二异丙醇亚硝胺(DPNI)，剂量500 mg/kg，每周1次，经过15周可诱发金仓鼠肝内胆管癌动物模型，成癌率为60%。用化学诱导方法制备肝内胆管癌动物模型周期长、操作繁琐，且成癌率不高。我们采用水动力转染的方法将过表达Notch1基因和蛋白激酶B(Akt)基因转座子裸质粒通过小鼠尾静脉注射，激活Notch信号通路及Akt信号通路，从而诱发小鼠的正常肝细胞转变成胆管癌细胞，建立小鼠的肝内胆管癌模型。成瘤周期为4周，操作简单，成癌率为100%，特别是本模型能够模拟人类肝内胆管癌发生发展的过程，是理想的研究肝内胆管癌实验动物模型。

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摘要
参考文献

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备注信息

通信作者：李永海，Email： mocdef.labiamgtnemirepxecmbb

通信作者：谈燚，Email： mocdef.aabnis7002iynatrotcod

基金项目：

国家自然科学基金 (81472656) 查看更多基金信息

蚌埠医学院创新计划 (Byycx1539) 查看更多基金信息

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