高等学校化学学报 ›› 2017, Vol. 38 ›› Issue (10): 1897-1905.doi: 10.7503/cjcu20170121

• 高分子化学 • 上一篇    下一篇

用于人类脂肪干细胞的纯化和体外长期培养的聚合物涂层

邱林子1, 王桂容1, 潘艳2, 王溢文3, 李新庆4, 邓林红2, 张嵘1()   

  1. 1. 常州大学材料科学与工程学院,2. 生物医学工程与健康科学研究院, 常州 213164
    3. 江苏和泽生物科技有限公司, 常州 213164
    4. 常州市第二人民医院, 常州 213164
    5. School of Chemistry, University of Edinburgh, Edinburgh, Edinburgh EH93JJ, UK
  • 收稿日期:2017-02-27 出版日期:2017-10-10 发布日期:2017-09-07
  • 作者简介:联系人简介: 张 嵘, 男, 博士, 教授, 主要从事生物材料研究. E-mail:rzhang@cczu.edu.cn
  • 基金资助:
    国家自然科学基金(批准号: 21374012)资助

Polymer Coatings for Purification and Long-term Culture of Human Adipose-derived Stem Cells

QIU Linzi1, WANG Guirong1, PAN Yan2, WANG Yiwen3, LI Xinqing4, DENG Linhong2, Mark Bradley5, ZHANG Rong1,*()   

  1. 1. School of Materials Science and Engineering,2. Institute of Biomedical Engineering and Health Science, Changzhou University, Changzhou 213164, China
    3. Jiangsu Union Biological Co., Ltd. Changzhou 213164, China
    4. Changzhou Second People’s Hospital, Changzhou 213164, China
    5. School of Chemistry, University of Edinburgh, Edinburgh, Edinburgh EH93JJ, UK
  • Received:2017-02-27 Online:2017-10-10 Published:2017-09-07
  • Contact: ZHANG Rong E-mail:rzhang@cczu.edu.cn
  • Supported by:
    † Supported by the National Natural Science Foundation of China(No.21374012)

摘要:

采用紫外固化法制备了基于丙烯酸酯类水凝胶的聚合物涂层(PC), 并用X射线光电子能谱(XPS)、 水接触角(WCA)和原子力显微镜(AFM)分别对PC进行了化学组成和表面性能的表征. 在PC表面进行了人类脂肪干细胞(hASC)的体外长期培养扩增, 得到的第3代细胞的生物学表征结果表明, 干细胞在PC表面能正常黏附生长, 流式细胞仪检测发现干细胞对特征标记物CD49d, CD73, CD105的阳性显性比例较高, 对HLA-DR和CD31几乎不显性, 说明扩增的干细胞具有hASC特征. 对PC上扩增的干细胞进行诱导分化, 并用油红O、 茜素红和阿利新蓝分别进行染色分析, 结果表明, 该干细胞保留了hASC的多能特性: 能分化为成脂、 成骨和成软骨细胞. 含有单体甲基丙烯酰氧乙基三甲基氯化铵(DMC)、 甲基丙烯酸环己酯(CHMA)和甲基丙烯酸-2-(二乙氨基)乙酯(DEAEMA)的PC2(质量比为3:1:2)在用于hASC体外长期培养时, 比其它PC和TCP更有利于细胞的黏附和增殖, 纯化细胞, 保持其多能性. 实时荧光定量PCR(RT-qPCR)的分析表明PC2上得到的细胞更容易向成骨和成软骨细胞分化.

关键词: 聚合物涂层, 人类脂肪干细胞, 体外长期培养, 纯化, 分化, 丙烯酸酯类水凝胶

Abstract:

Polymer coating(PC) based on polyacrylate hydrogels was prepared through UV initiated polymerization and characterized with X-ray photoelectron spectroscopy(XPS), water contact angles(WCA) and atomic force microscopy(AFM) respectively to understand their chemistry and surface properties. The biological characteristics of human adipose-derived stem cell(hASC) were evaluated after long-term cultured on PC in vitro for 3 passages. The results indicated that the hASC proliferated successfully on the PC, expressed high percentage of specific membrane markers CD49d, CD73, CD105, and very low percentage of HLA-DR and CD31 by using flow cytometry. The results suggested that the proliferated stem cells had the characteristics of hASC. The hASC were differentiated to adipocytes, osteoblasts and chondrocytes and stained with Oil Red O, Alizarin Red S and Alcian Blue respectively to demonstrate the maintaining of pluripotency of hASC. The results showed that the PC2 based on methacrylatoethyltrimethyl ammonium chloride, cyclohexyl methacrylate and diethyl amino ethyl methacrylate(mass ratio of 3:1:2) was the best in hASC binding and proliferation, improved the purification, maintained the pluripotency among the three coatings and TCP. Relative gene expression of the differentiated cells was analyzed by quantitative real-time polymerase chain reaction(RT-qPCR). The results show that the harvest hASC from PC2 are apt to differentiate to osteoblasts and chondrocytes.

Key words: Polymer coating, Human adipose-derived stem cell, Long-term culture in vitro, Purification, Differentiation, Polyacrylate hydrogel

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