胰岛素样生长因子-I对人骨髓间充质干细胞增殖的影响  

Effects of insulin-like growth factor-I on the proliferation of human mesenchymal stem cells

作　　者：李建华[1] 黄建荣[2] 樊雪萍[1] 康奕飞[1] 许继德[1] 涂永生[1] 

机构地区：[1]广州医学院生理学教研室,广东省广州市510182 [2]中山大学第二附属医院骨科,广东省广州市510120

出　　处：《中国临床康复》2004年第8期1464-1466,T004,共4页Chinese Journal of Clinical Rehabilitation

摘　　要：目的:建立人胚骨髓间充质干细胞(MSCs)分离及培养的方法,探讨胰岛素样生长因子-I(IGF-I)对MSCs增殖的影响。方法:通过密度梯度离心,贴壁法分离培养人胚MSCs,取第4代MSCs,应用流式细胞术测定细胞周期和CD44、CD34、CD45表达。采用MTT法观察不同含量和不同作用时间的IGF-I对MSCs增殖的影响。结果:体外培养的MSCs呈现成纤维细胞形态,流式细胞仪检测结果显示,MSCs阳性表达CD44,阴性表达CD34,CD45。细胞周期分析显示IGF-I使S+G2+M期细胞数增加,占细胞总数(29.9±2.3)%(t=10.4,P<0.01)。1μg/LIGF-I促MSCs增殖作用不明显,10μg/LIGF-I即可促MSCs增殖,IGF-I含量为100μg/L时,其促增殖作用达最大值,吸光度为2.95。而当IGF-I含量大于100μg/L时,促细胞增殖作用反而下降;时效关系显示,IGF-I第1天发挥促增殖作用,第5天促增殖作用达高峰,维持时间可达7d。结论:外源性IGF-I能促进MSCs增殖,为实现MSCs体外快速扩增提供适宜的条件。AIM:To establish a method of isolation and cultivation of human bone marrow mesenchymal stem cells(MSCs) and investigate the effects of insulin like growth factor I(IGF I) on the proliferation of human fetal MSCs in vitro. METHODS:Human fetal MSCs were isolated and cultured in DMEM by means of density gradient centrifugation and adhesion,and the 4th generation of MSCs were obtained.Cell cycle and the expression of CD44,CD34 and CD45 were analyzed by flow cytometry.The proliferation of MSCs in different doses and different culture time of IGF I was measured with methyl thiazolyl titrazolium(MTT) method. RESULTS:Human fetal MSCs cultured in vitro displayed a fibroblast like morphology. MSCs were uniformly positive for CD44, and negative for CD34 and CD45 displayed by flow cytometry.Cell cycle analysis showed that when IGF I was added into the culture medium,more MSCs were in S+G2+M phase,which was(29.9±2.3)%of the total amount(t=10.4,P< 0.01). MTT measurement results showed that 1 μg/L of IGF I could not enhance the proliferation of MSCs obviously,while 10 μg /L of IGF I significantly enhanced the proliferation of MSCs,and this enhancement was maximal at the concentration of 100 μg /L with a optical absorbance of 2.95.While the proliferation was decreased when the IGF I was over 100 μg /L.Aging analysis showed that 100 μg /L IGF I enhanced MSCs proliferation on the first day and reached the maximum on the fifth day,and the proliferated time lasted for 7 days. CONCLUSION:Exogenous IGF I can promote the proliferation of MSCs which provides an optimal culture condition for rapid proliferation of MSCs in vitro.

关 键 词：胰岛素样生长因子-Ⅰ 人骨髓间充质干细胞 细胞增殖 细胞周期 流式细胞仪 检测 

分 类 号：R329.2[医药卫生—人体解剖和组织胚胎学]