中国组织工程研究

中国组织工程研究 ›› 2011, Vol. 15 ›› Issue (40): 7450-7454.doi: 10.3969/j.issn.1673-8225.2011.40.010

• 脐带脐血干细胞 umbilical cord blood stem cells • 上一篇    下一篇

人脐血中分离培养扩增内皮集落形成细胞及其生物相容性

陈  超，杨述华，冯  勇，陈  东，禹  虔，王小红   

  1. 华中科技大学同济医学院附属协和医院，湖北省武汉市  430022
  • 收稿日期:2011-03-02 修回日期:2011-04-06 出版日期:2011-10-01 发布日期:2011-10-01
  • 通讯作者: 杨述华，教授，主任医师，博士生导师，华中科技大学同济医学院附属协和医院骨科，湖北省武汉市430022 shuhuayabc@vip.sina.com
  • 作者简介:陈超☆，男，1984年生，湖北省枣阳市人，华中科技大学同济医学院附属协和医院在读博士，主要从事骨外科方面的研究。 chenchao027@163.com
  • 基金资助:

    国家自然科学基金资助项目(30973044)。

Culture, large-scale expansion and biocompatibility of endothelial colony forming cells

Chen Chao, Yang Shu-hua, Feng Yong, Chen Dong, Yu Qian, Wang Xiao-hong   

  1. Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan  430022, Hubei Province, China
  • Received:2011-03-02 Revised:2011-04-06 Online:2011-10-01 Published:2011-10-01
  • Contact: Yang Shu-hua, Professor, Doctoral supervisor, Chief physician, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430022, Hubei Province, China shuhuayabc@vip.sina.com
  • About author:Chen Chao☆, Studying for doctorate, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430022, Hubei Province, China chenchao027@163.com
  • Supported by:

    the National Natural Science Foundation of China, No. 30973044*

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摘要：

背景：内皮集落形成细胞是内皮祖细胞的一种亚型，体外培养扩增及其在组织工程中的应用尚不明确。
目的：探索从人脐血中分离培养扩增内皮集落形成细胞并观察其生物相容性。
方法：采用密度梯度离心法从人脐血中分离单个核细胞，接种于Ⅰ型鼠尾胶原包被的培养板上，采用内皮祖细胞专用EGM-2培养基诱导培养，早期传代后扩增，免疫荧光鉴定细胞，并检测其体外成血管和增殖能力，与纳米羟基磷灰石/磷酸三钙材料复合培养后的生物相容性。
结果与结论：脐血来源内皮集落形成细胞呈铺路石样集落生长，吞噬Dil-Ac-LDL并结合FITC-UEA-1，表达CD31，vWF，KDR和Tie-2，早期传代后拥有较强增殖潜能，可在体外大量扩增，在体外基质胶上可形成闭合管腔样结构，复合纳米羟基磷灰石/磷酸三钙材料后生物学特性不受影响。提示可从脐血中分离并体外大量扩增内皮集落形成细胞。

关键词: 内皮祖细胞, 内皮集落形成细胞, 脐血, 组织工程, 种子细胞

Abstract:

BACKGROUND: There is still uncertain about in vitro culture and expansion of endothelial colony forming cells (ECFCs) and its application in tissue engineering.
OBJECTIVE: To elucidate a method for in vitro culture, identification and large-scale expansion of ECFCs and to investigate its biocompatibility with biological material.
METHODS: Monocytes were isolated by Ficoll density gradient centrifugation from human umbilical cord blood, in vitro cultured and expanded in EGM-2 medium on tissue culture plates precoated with type I rat tail collagen. Then cells were characterized by immunofluorescence staining. The growth kinetics, tube formation capacity and biocompatibility with nano-hydroxyapatite/tricalcium phosphate (HA/TCP) were observed.
RESULTS AND CONCLUSION: During culture, the cells displayed cobble-stone morphology with outgrowth, took up DiI-Ac-LDL, bound FITC-UEA-1, and stained positively for cell markers CD31, vWF, KDR and Tie-2. These cells demonstrated high proliferation rate after passage, formed closed network structures on Matrigel and the bioactivity remained stable when co-cultured with HA/TCP. Results indicate it is feasible to isolate and large-scale expand ECFCs from human umbilical cord blood in vitro and ECFCs may serve as seed cells in tissue engineering.

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