天津医药

天津医药 ›› 2015, Vol. 43 ›› Issue (6): 595-598.doi: 10.11958/j.issn.0253-9896.2015.06.005

• 细胞与分子生物学 • 上一篇    下一篇

Apelin-13促进脐带华通胶间充质干细胞分化成血管网

高彦琳 1， 陈厚良 2， 张宁坤 2， 高连如 2， 朱智明 1   

  1. 1安徽医科大学海军临床学院 ； 2海军总医院心脏中心
  • 收稿日期:2015-02-11 修回日期:2015-04-20 出版日期:2015-06-15 发布日期:2015-06-10
  • 通讯作者: 朱智明 E-mail:zhuzhiming6542@sina.com.cn
  • 基金资助:
    国家自然科学基金资助项目 （81170094）

Apelin-13 promote mesenchymal stem cells isolated from Wharton’ s jelly to differentiate into#br# vascular networks

GAO Yanlin1, CHEN Houliang2, ZHANG Ningkun2, GAO Lianru2, ZHU Zhiming1   

  1. 1 Clinical College of Navy Medicine, Anhui Medical University, Anhui 230000, China; 2 Department of Cardiology,
    Navy General Hospital
  • Received:2015-02-11 Revised:2015-04-20 Published:2015-06-15 Online:2015-06-10
  • Contact: ZHU Zhiming E-mail:zhuzhiming6542@sina.com.cn

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摘要： 目的 探索 apelin-13 对干细胞定向血管网分化的调控作用。方法 采用组织块贴壁法从人脐带华通胶组织中分离间充质干细胞， 流式细胞仪检测其免疫表型。取第 3 代人华通胶间充质干细胞， 分别接种于 4 个培养瓶中， 记为 A1、 A2、 A3、 A4 组， 用诱导液诱导 7 d， 每天向 4 组中分别加入浓度 0、 1×10－6、 10×10－6、 100×10－6 mol/L 的apelin-13 20 μL， 倒置显微镜下观察细胞形态及生长情况， 并对其进行血管性血友病因子（vWF）抗体免疫荧光染色及 CD31 流式细胞学鉴定。使用水凝胶的三维培养基对诱导后的内皮细胞进行培养， 并按照原 A1、 A2、 A3、 A4 组的顺序重新依次编号为 S1、 S2、 S3、 S4 组。每天分别向 S1、 S2、 S3、 S4 组中加入浓度 0、 1×10－6、 10×10－6、 100×10－6 mol/L的 apelin-13 20 μL。7 d 后倒置显微镜下观察细胞的形态、 生长情况及在三维培养基中形成血管样结构情况。结果 人华通胶间充质干细胞可以诱导分化为 vWF 免疫荧光染色阳性的内皮样细胞。随着 apelin-13 蛋白浓度的增加， CD31 阳性表达率也随之升高。显微镜下观察到内皮样细胞在水凝胶的三维培养基中能够形成血管样结构。结论 证实 apelin-13 参与并调控人脐带华通胶间充质干细胞分化形成血管网。

关键词: 间充质干细胞, apelin-13, 血管生成, 三维培养, 脐带

Abstract: Objective To explore the role of apelin-13 in regulating stem cell differentiation into vascular net. Methods Mesenchymal stem cells were isolated from human umbilical Wharton’ s jelly using tissue adherence method．Their immunophenotypes were detected by flow cytometry . Passage 3 of WJ-MSCs (Wharton’ s jelly-mesenchymal stem cells)were inoculated in 4 flasks, denoted as A1, A2, A3, A4 group. Twenty μL of apelin-13 at concentrations of 0, 1×10－6, 10 ×10－6 and 100 ×10－6 mol/L were added to A1, A2, A3 and A4 respectively each day. After being induced for 7 days, cell morphology and viability were observed under inverted microscope. Von Willebrand factor (vWF) was examined by immunofluorescence and CD31 was identified by flow cytometry. Upon incubating with three dimensional culture medium of hydrogel,those cultured A1, A2, A3 and A4 were renumbered as S1, S2, S3, S4. Again, twenty μL of apelin-13 at concentrations of 0,1×10－6, 10 ×10－6and 100 ×10－6 mol/L were used to treat S1, S2, S3 and S4 respectively. After 7 days, cell morphology, viability and vas -like networks were observed with inverted microscope. Results Our study showed that WJ-MSCs can be induced by apelin 13 to differentiate into endothelial cells lineage indicated by positive of vWF staining. Moreover, CD31 expression increases significantly upon apelin-13 addition in a dosage dependent manner. The endothelial cells line formed vas like networks when cultured with three-dimensional medium containing hydrogel. Conclusion This study demonstrated that apelin-13 could promote human umbilical cord-MSCs to differentiate into endothelium lineage then to form vascular networks.

Key words: mesenchymal stem cells, apelin-13, angiogenesis, 3-dimensional culture, umbilical cord