基础医学与临床 ›› 2013, Vol. 33 ›› Issue (8): 1043-1049.

• 研究论文 • 上一篇    下一篇

HGF及G-CSF诱导大鼠骨髓间充质干细胞分化为肝样细胞

高勇1,梅浙川1,蒋明德2   

  1. 1. 重医医科大学附属第二医院消化内科
    2. 成都军区总医院
  • 收稿日期:2012-09-28 修回日期:2012-12-07 出版日期:2013-08-05 发布日期:2013-07-18
  • 通讯作者: 梅浙川 E-mail:meizhechuan@21cn.com

HGF and G-CSF induce the differentiation of rat bone marrow mesenchymal stem cells into hepatocyte-like Cells

  • Received:2012-09-28 Revised:2012-12-07 Online:2013-08-05 Published:2013-07-18

摘要: 目的 探讨肝细胞生长因子(HGF)以及粒细胞集落刺激因子(G-CSF)可否诱导大鼠骨髓间充质干细胞(BM-MSCs)分化为肝样细胞。方法 以贴壁法分离、培养大鼠BM-MSCs,流式细胞术鉴定表面标志。实验分为:对照组(10%FBS),0.1μmol/L G-CSF干预组,20ng/mL HGF干预组,HGF联合G-CSF共同干预组(0.1μmol/L G-CSF+20ng/mL HGF),诱导后第7天、14天和21天提取细胞总RNA以及总蛋白后进行RT-PCR和Western blot检测白蛋白(ALB)和甲胎蛋白(AFP)基因mRNA转录和蛋白水平。结果:20ng/mLHGF干预组和HGF联合G-CSF共同干预组P3代BM-MSCs诱导后细胞形态似肝样细胞。BM-MSCs表面标志物:CD34-PE 0.3%,CD45-FITC 0.1%,CD90-FITC 99.6%,CD105-PE99.8%。20ng/mLHGF干预组和HGF联合G-CSF共同干预组检测到白蛋白(ALB)和甲胎蛋白(AFP)基因mRNA转录水平及蛋白水平表达。在第7天、14天、21天,随时间增加ALB表达量递增、而AFP表达量递减,且同一时间两组表达量均有差异(P<0.05)。结论 HGF能诱导大鼠BM-MSCs分化为肝样细胞,G-CSF对HGF诱导大鼠BM-MSCs分化为肝样细胞有协同作用。

关键词: 【关键词】骨髓间充质干细胞, 肝细胞生长因子, 粒细胞集落刺激因子, 肝样细胞

Abstract: Objective To investigate if the hepatocyte growth factor(HGF)and granlocyte-colony stimulating factor (G-CSF) could induce differentiation of rat bone marrow mesenchymal stem cells(BM-MSCs)into hepatocyte-like cells .Methods The BM-MSCs were isolated and cultured in- vitro by adherent method,then identified by flow cytometry.The BM-MSCs of the third passage were used as four groups - control group(10%FBS)and intervation groups(0.1μmol/L G-CSF;20ng/mLHGF;0.1μmol/L G-CSF+20ng/mLHGF).The mRNA and protein were extracted on 7,14,21 days,then detected by RT-PCR and Western-blot. Results.The third passage BM-MSCs of intervation groups (20ng/mLHGF and 0.1μmol/L G-CSF+20ng/mLHGF) got hepatocyte like feafure after induction.The results tested by Flow cytometry analysis were as follows: CD34-PE0.3%,CD45-FITC0.1%,CD90-FITC99.6%,CD105-PE99.8%. The mRNA and protein of ALB and AFP were detected respetively by RT-PCR and Western-blot on day 7,14,and 21 from inervation groups (10%FBS+20ng/mLHGF;10%FBS+0.1μmol/L G-CSF+20ng/mLHGF), however ,the amount of ALB increased as time went by ,while AFP decreased to extremely low on day 21.Meaningfully,the difference between the two groups is significant(P<0.05). Conclusion HGF can induce the differentiation of BM-MSCs into hepatocyte-like cells, which can be significantly enhanced by G-CSF.

Key words: 【Keywords】Bone marrow mesenchymal stem cells, Hepatocyte growth factor, granlocyte-colony stimulating factor, Hepatocyte-like cells