中国组织工程研究

中国组织工程研究 ›› 2012, Vol. 16 ›› Issue (15): 2695-2698.doi: 10.3969/j.issn.1673-8225.2012.15.008

• 血管组织构建 vascular tissue construction • 上一篇    下一篇

胰蛋白酶消化法体外培养和鉴定人脐静脉血管内皮细胞*☆

白燕慧1，张明昌1，边  芳2   

  1. 1华中科技大学同济医学院附属协和医院眼科，湖北省武汉市   430022；2武汉市新华路协和医院眼科，湖北省武汉市  430022
  • 收稿日期:2011-10-08 修回日期:2011-12-15 出版日期:2012-04-08 发布日期:2012-04-08
  • 通讯作者: 张明昌，博士生导师，主任医师，华中科技大学同济医学院附属协和医院眼科，湖北省武汉市 430022
  • 作者简介:白燕慧☆，女，1982年生，河南省郑州市人，汉族，华中科技大学同济医学院附属协和医院在读博士，主要从事角膜病的研究。 z212006baiyanhui@126.com
  • 基金资助:

    国家自然科学基金青年基金资助项目(81000365)。

Culture and identification of human umbilical vein endothelial cells in vitro using Trypsin digestion method 

Bai Yan-hui1, Zhang Ming-chang1, Bian Fang2   

  1. 1Department of Ophthalmology, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan  430022, Hubei Province, China; Union Hospital, Wuhan  430022, Hubei Province, China
  • Received:2011-10-08 Revised:2011-12-15 Online:2012-04-08 Published:2012-04-08
  • Contact: author: Zhang Ming-chang, Doctoral supervisor, Chief physician, Department of Ophthalmology, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430022, Hubei Province, China
  • About author:Bai Yan-hui☆, Studying for doctorate, Department of Ophthalmology, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430022, Hubei Province, China z212006baiyanhui@126.com
  • Supported by:

    the National Natural Science Foundation for the Youth, No. 81000365*  

PDF

307

被引次数

6

摘要：

背景：体外建立稳定可靠的人脐静脉血管内皮细胞模型，目前培养方法缺乏统一的标准。
目的：探索脐静脉内皮细胞的体外培养的方法。
方法：采用2.5 g/L胰蛋白酶和0.02%EDTA消化、分离、体外原代培养及消化传代脐静脉内皮细胞，简化完全培养液组分(不添加血管内皮细胞生长因子、肝素等辅助因子)，当原代培养细胞80%以上汇合，根据细胞特有的形态学特征和Ⅷ因子进行内皮细胞的鉴定。
结果与结论：种植在培养瓶中的内皮细胞2 h贴壁生长，24 h换液后内皮细胞80%融合，细胞状态好，内皮细胞呈单层铺路石样外观，经过镜下观察和Ⅷ因子相关抗原鉴定证明是脐静脉内皮细胞。证实用胰蛋白酶灌注脐静脉是一种简单、实用的获得人脐静脉血管内皮细胞的方法，可靠性大，成功率高，可以构建体外研究血管内皮细胞的模型。
关键词：人脐静脉；血管内皮细胞；细胞培养；形态学；鉴定
缩略语注释：HUVECs: human umbilical vein endothelial cells，人脐静脉血管内皮细胞
doi:10.3969/j.issn.1673-8225.2012.15.008

关键词: 人脐静脉, 血管内皮细胞, 细胞培养, 形态学, 鉴定

Abstract:

BACKGROUND: The establishment of human umbilical vein endothelial cells model in vitro has significant meaning for the study of neovascularization, but human umbilical vein endothelial cells are difficult to cultivate in vitro and there is not a criterion.
OBJECTIVE: To explore how to harvest and identify human umbilical vein endothelial cells in vitro.
METHODS: 0.25 g/L Trypsin and 0.02% ethylene diamine tetraacetic acid perfusion method was used to isolate human umbilical vein endothelial cells from the fresh umbilical cord, and the cells were cultured and amplified. The composition of culture medium was simplified by not adding vascular endothelial cell growth factor and heparin. Human umbilical vein endothelial cells grown to 80% confluence were identified by morphological observation and immunofluorescence method.
RESULTS AND CONCLUSION: The endothelial cells spread on the bottom of the dishes within 2 hours, then coalesced and grew to form a confluent monolayer of polygonal cells within 24 hours. The cultured cells were identified as human umbilical vein endothelial cells. Trypsin perfusion is a simple and effective method for collection of human umbilical vein endothelial cells. Cells harvested with this protocol can be used as models on research of vascular endothelial cells in vitro.
 

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