不同培养条件对人脂肪源性干细胞生长状态的影响  

Effects of different culture conditions on the growth state of human adipose tissue-derived stem cells

作　　者：王鹏基[1] 周六化[1] 陈赟[2] 邱雪峰[1] 杨斌[1] 戴玉田[1] 

机构地区：[1]南京大学医学院附属鼓楼医院男科,210008 [2]上海市第十人民医院泌尿外科

出　　处：《中华实验外科杂志》2013年第8期1698-1700,共3页Chinese Journal of Experimental Surgery

基　　金：国家自然科学基金资助项目（81170563）

摘　　要：目的观察不同培养条件对人脂肪源性干细胞（hADSCs）体外生长状态的影响。方法取腹部手术患者皮下脂肪组织，用0．1％的I型胶原酶消化法分离出hADSCs，将获得的细胞分别在以下3组培养条件下培养：高糖DMEM培养基、高糖DMEM培养基＋碱性成纤维生长因子（bFGF）、间充质干细胞培养基（MSCM）。取第2代或者第3代细胞进行以下实验：细胞免疫荧光及流式细胞仪行细胞表面标志物CD31、CD34和Stro-1鉴定；绘制细胞生长曲线观察3组细胞的生长状态；采用不同的培养条件对3组细胞进行体外培养，观察细胞的增殖。结果通过胶原酶消化法可成功地分离出hADSCs。流式细胞仪检测DMEM组CD31、CD34、Stro-1阳性率分别为0．9％、2．9％、56．7％；DMEM＋bFGF组CD31、CD34、Stro．1阳性率分别为1．3％、2．5％、73．5％；MSCM组CD31、CD34、Stro-1阳性率分别为0．7％、2．4％、84．5％。采用不同的培养条件对hADSCs进行培养，DMEM组、DMEM＋bFGF组、MSCM组细胞的倍增时间分别为93．7、64．5、49．1h，差异有统计学意义（P〈0．01）。此外，各组中所培养的细胞在MSCM和高糖DMEM＋bFGF的培养条件下的增殖速率明显高于高糖DMEM培养条件（P〈0．05）。结论采用MSCM培养hADSCs可以在短期内获得大量细胞，在传统的高糖DMEM培养基中加入bFGF后也可以获得相似的效果，可以满足组织工程研究的种子细胞数量的要求。Objective To observe the effects of different culture conditions on growth state of hu- man adipose tissue-derived stem cells （hADSCs）. Methods The subcutaneous adipose tissue was collect- ed surgically. Primary hADSCs were isolated subsequently by the methods of stirring digestion within 1% collagenase type I. The cells were divided into three groups and cultured in high-glucose DMEM, high-glu- cose DMEM containing basic fibroblast growth factor （bFGF） and mesenchymal stem cell media （MSCM） respectively. The expression levels of CD31, CD34 and Stro-1 were detected by using immumofluorescence and flow cytometry. The growth state of cells in the three groups was evaluated. Proliferation of cells in each group was evaluated after culture in different culture conditions. Results hADSCs were successfully isolated through stirring digestion within collagenase I. The positive expression rate of CD31, CD34 and Stro-1 in hADSCs was 0. 9%, 2.9% and 56.7% respectively in DMEM, 1.3%, 2. 5% and 73.5% in DMEM containing bFGF, and 0.7%, 2. 4% and 84. 5% in MSCM, respectively. The cell doubling time in DMEM, DMEM containing bFGF and MSCM was 93.7 h, 64. 5 h and 49. 1 h respectively. MSCM and high-glucose DMEM containing bFGF could significantly promote the proliferation of cells in each group as compared with high-glucose DMEM （P 〈 0. 05）. Conclusion A great number of hADSCs could be ob- tained in a short time when cultured in MSCM and high-glucose DMEM containing bFGF, which can suffice for the cell source for tissue engineering.

关 键 词：脂肪源性干细胞 培养 组织工程 

分 类 号：R329[医药卫生—人体解剖和组织胚胎学]