重组人粒细胞集落刺激因子促进骨髓间充质干细胞增殖

Promoting the proliferation of bone marrow mesenchymal stem cell by recombinant human granulocyte colony-stimulating factor

目的探讨重组人粒细胞集落刺激因子(rhG-CSF)对小鼠骨髓间充质干细胞(MSCs)增殖的影响。方法将昆明小鼠随机分为2组(n=15),G-CSF组采髓前皮下注射重组人粒细胞集落刺激因子(rhG-CSF)80μg·kg-1·d-1,连用5d,对照组皮下注射等量生理盐水。每组分别于最后一次注射后6h、12h及7d(168h)取小鼠骨髓,分离、培养MSCs,计数成纤维样细胞集落形成单位(CFU-F)的个数;应用流式细胞仪检测单个核细胞(MNCs)细胞周期及CFU-F细胞表面抗原特征。结果应用rhG-CSF后,培养MNCs所获得的CFU-F数目增加(P<0.01),CFU-F数与取材时间(6h、12h、7d)呈负相关(P<0.05),但12h取材者与7d取材者比较,CFU-F数目的差异无统计学意义(P>0.05)。MNCs培养形成的CFU-F,其细胞表面抗原呈CD34-、CD133-、CD90+、CD105+,分别占2.5%、3.1%、67.0%和78.0%。G-CSF组各时间点获得的MNCs,其G0/G1期细胞百分率较对照组低(P<0.05),而S+G2/M期增高(P<0.05),且6h取材者S+G2/M期细胞百分率明显高于12h和7d取材者(P<0.05)。结论rhG-CSF可促进骨髓MNCs进入细胞增殖周期,增殖的高峰在应用G-CSF后6h左右。

Objective To investigate the effects on proliferation of bone marrow mesenchymal stem cells （MSCs） by recombinant human granulocyte colony-stimulating factor in mice. Methods Kunming mice were randomly divided into G-CSF and control groups （n = 15）. The mice were subjec ted to subcutaneous injections of rhG-CSF at a dose of 80μg/kg per day and control of saline for 5 days. The bone marrow was obtained on 6th, 12th and 168th h respectively after the final administration. The MSCs were separated and cultured, and the colony forming unit-fibroblast （CFU-F） was evaluated. The cell cycle and the surface antigens were analyzed by flow cytometry. Results The number of CFU-F was increased after administration of the rhG-CSF （P〈0. 01 ）. The number of CFU-F in G-CSF groups was negatively relevant to the sampling time from bone marrow （P〈0. 05）, but there was no obvious difference between 12 h and 168 h （P〈0. 05）. Flow cytometic detection of MSCs surface marks in fibroblast colony showed CD34^- , CD133^- , CD90^＋ and CD105^＋ , with the percentage of 2.5%, 3.1%, 67. 0 % and 78. 0 %, respectively. After mobilization with rhG-CSF, the percentage of G0/G1phases in bone marrow MNCs was decreased （P〈0. 05）, while that percentage of S ＋ G2/M phases increased （P〈0. 05）, and the percentage of S ＋ G2/M phases in G-CSF groups at 6 h was significantly higher than that at 12 h and 168 h, respectively （P〈0. 05）. Conclusion rhG-CSF treatment could promote bone marrow MNCs reentering into cell cycle, and the peak value occurred at about 6 h after the final rhG-CSF administration.