Kang Xin-Qin, Zang Wei-Jin, Bao Li-Jun, Li Dong-Ling, Xu Xiao-Li, Yu Xiao-Jiang
Department of Genetics and Molecular Biology, School of Medicine, Xi'an Jiaotong University, Xi'an 710061, PR China.
Cell Biol Int. 2006 Jul;30(7):569-75. doi: 10.1016/j.cellbi.2006.02.007. Epub 2006 Mar 6.
It has been demonstrated that the number and differentiating potential of bone marrow mesenchymal stem cells (MSCs) decrease with age. Therefore, the search for alternative sources of MSCs is of significant value. In the present study, MSCs were isolated from umbilical cord blood (UCB) by combining gradient density centrifugation with plastic adherence. Cultured cells were treated with ascorbate acid-2-phosphate, dexamethasone, beta-glycerophosphate dexamethasone, insulin, 1-methyl-3-isobutylxamthine, indomethacin, beta-mercaptoethanol, butylated hydroxyanisole, FGF-4 and HGF. Differentiating characterization of UCB-derived MSCs were detected by cytochemistry, immunocytochemistry, radioimmunoassay, RT-PCR and urea assay. The results showed UCB-derived MSCs could differentiate into osteoblasts, adipocytes and neuron-like cells. When MSCs were cultured with FGF-4 and HGF, approximately 63.6% of cells became small, round and epithelioid on day 28 by morphology. Compared with the control, levels of AFP in the supernatant liquid increased significantly from day 12 and were higher on day 28 (P<0.01). Albumin increased significantly from day 16 (P<0.01). Urea was first detected on day 20 (P<0.01), and continued to increase on day 28 (P<0.01). Cells first expressed CK-18 on day 16 through immunocytochemistry analysis. RT-PCR analysis showed that differentiated cells could express a number of hepatocyte-specific genes in a time-dependent manner. Glycogen storage was first seen on day 24. Our results suggest that UCB-derived MSCs can differentiate not only into osteoblasts, adipocytes and neuron-like cells, but also into hepatocytes. Human UCB-derived MSCs are a new source of cell types for cell transplantation and therapy.
已有研究表明，骨髓间充质干细胞（MSCs）的数量和分化潜能会随着年龄的增长而降低。因此，寻找MSCs的替代来源具有重要价值。在本研究中，通过梯度密度离心与塑料贴壁相结合的方法，从脐带血（UCB）中分离出MSCs。将培养的细胞用抗坏血酸-2-磷酸、地塞米松、β-甘油磷酸地塞米松、胰岛素、1-甲基-3-异丁基黄嘌呤、吲哚美辛、β-巯基乙醇、丁基羟基茴香醚、FGF-4和HGF进行处理。通过细胞化学、免疫细胞化学、放射免疫分析、RT-PCR和尿素测定法检测UCB来源的MSCs的分化特征。结果显示，UCB来源的MSCs能够分化为成骨细胞、脂肪细胞和神经元样细胞。当MSCs与FGF-4和HGF一起培养时，在第28天通过形态学观察，约63.6%的细胞变得小而圆且呈上皮样。与对照组相比，上清液中AFP的水平从第12天开始显著升高，并在第28天更高（P<0.01）。白蛋白从第16天开始显著增加（P<0.01）。尿素在第20天首次检测到（P<0.01），并在第28天持续增加（P<0.01）。通过免疫细胞化学分析，细胞在第16天首次表达CK-18。RT-PCR分析表明，分化的细胞能够以时间依赖性方式表达多种肝细胞特异性基因。糖原储存最早在第24天出现。我们的结果表明，UCB来源的MSCs不仅可以分化为成骨细胞、脂肪细胞和神经元样细胞，还可以分化为肝细胞。人UCB来源的MSCs是细胞移植和治疗的新型细胞类型来源。
