复方扶芳藤合剂含药血清促进骨髓间充质干细胞增殖的作用机制

Mechanism of compound Fufangteng mixture-containing serum in promoting proliferation of bone marrow mesenchymal stem cell

    摘要
  • 摘要:
      目的  探讨复方扶芳藤合剂含药血清对骨髓间充质干细胞（BMSC）增殖的影响及其机制。
      方法  通过直接贴壁法体外分离、培养、纯化大鼠BMSC，对其进行细胞形态学观察，采用流式细胞仪对其表面标志物进行鉴定。复方扶芳藤合剂按3 mL/（kg·d）给予大鼠灌胃14 d后取含药血清。将BMSC分为空白对照组、含药血清组、Notch1小干扰核糖核酸（siRNA）组和Notch1 siRNA+含药血清组，检测各组BMSC的增殖率以及Notch1信号通路相关信使核糖核酸（mRNA）和蛋白的相对表达量。
      结果  镜下观察可见第1代BMSC呈长梭形，以平行排列生长为主或呈漩涡状生长。第3代BMSC阳性表达CD90、CD44，而CD45呈阴性表达。与空白对照组比较，含药血清组和Notch1 siRNA+含药血清组BMSC的增殖率均升高，Notch1 siRNA组BMSC的增殖率下降（均为P < 0.05）；与Notch1 siRNA组比较，Notch1 siRNA+含药血清组BMSC的增殖率升高（P < 0.05）。与空白对照组比较，含药血清组Hey1、Delta样配体（DLL）1的mRNA和蛋白相对表达量均升高，Notch1 siRNA组和Notch1 siRNA+含药血清组Hey1、DLL1的mRNA和蛋白相对表达量均下降，差异均有统计学意义（均为P < 0.05）；与Notch1 siRNA组比较，Notch1 siRNA+含药血清组Hey1、DLL1的mRNA和蛋白相对表达量均升高，差异均有统计学意义（均为P < 0.05）。
      结论  复方扶芳藤合剂含药血清可促进大鼠BMSC的增殖，其作用机制可能与Notch1信号通路激活有关。

     

    Abstract:
      Objective  To investigate the effect of compound Fufangteng mixture-containing serum on the proliferation of bone marrow mesenchymal stem cell (BMSC) and its mechanism.
      Methods  Rat BMSC were isolated, cultured and purified in vitro by direct adherence method. Cell morphology was observed. Surface markers were identified by flow cytometry. The rats were treated with compound Fufangteng mixture at a dose of 3 mL/(kg·d) by gavage for 14 d, and then the drug-containing serum was collected. BMSC were divided into the blank control group, drug-containing serum group, Notch1 small interfering ribonucleic acid (siRNA) group and Notch1 siRNA+drug-containing serum group. The proliferation rate of BMSC was detected and the relative expression levels of Notch1 signaling pathway-associated messenger ribonucleic acid (mRNA) and proteins were measured in each group.
      Results  Microscopic observation showed that the first generation BMSC were seen in the long spindle shape, and grown in the parallel or spiral pattern. The third generation BMSC positively expressed CD90 and CD44, whereas were negative for CD45. Compared with the blank control group, the proliferation rate of BMSC in the drug-containing serum group and Notch1 siRNA+ drug-containing serum group was significantly increased, whereas that of BMSC was significantly decreased in the Notch1 siRNA group (all P < 0.05). Compared with the Notch1 siRNA group, the proliferation rate of BMSC was significantly increased in the Notch1 siRNA+drug-containing serum group (P < 0.05). Compared with the blank control group, the relative expression levels of Hey1 and Delta-like ligand (DLL)1 mRNA and proteins were significantly up-regulated in the drug-containing serum group, whereas those were significantly down-regulated in the Notch1 siRNA group and Notch1 siRNA+drug-containing serum group (all P < 0.05). Compared with the Notch1 siRNA group, the relative expression levels of Hey1 and DLL1 mRNA and proteins were significantly up-regulated in the Notch1 siRNA+drug-containing serum group (all P < 0.05).
      Conclusions  Compound Fufangteng mixture-containing serum may promote the proliferation of rat BMSC, and its mechanism is probably associated with the activation of Notch1 signaling pathway.

     

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