三维培养对脐带间充质干细胞表型及造血因子基因表达的影响

本研究探讨三维立体培养条件下脐带间充质干细胞（UC-MSC）的细胞表型变化及部分造血相关因子的基因表达,为下一步研究三维立体条件下的造血扩增及诱导分化提供基础.从脐带中分离出MSC,分别在二维及三维条件下培养,通过流式细胞术检测MSC表面抗原的表达水平,通过成血管试验比较了两种细胞的成血管能力,并通过实时定量PCR检测两种培养条件下造血相关细胞因子的基因表达情况.结果表明：从脐带中成功地分离出了MSC,在三维培养条件下其内皮细胞、内皮祖细胞和间充质原始干细胞的表面标记CD31、CD133和CD271等阳性细胞百分比显著增加,体外成血管能力增强;骨架蛋白β-actin基因在三维培养条件下表达上调,G-CSF、LIF、SCF、IL-1 α、IL-1β、IL-3、IL-7和IL-11等与造血调控有关的细胞因子基因表达也有不同程度的增高,其中LIF、IL-3和IL-7升高尤为明显;免疫调控相关的细胞因子IL-10的表达也增高,而SDF-1和IL-6的表达虽有轻微的降低,但无显著性差异.结论：三维培养条件下UC-MSC的CD31、CD133和CD271表达上调,体外成血管能力增强,并且多个造血相关因子的基因表达水平上调.

Effects of Three-Dimensional Culture on Cell Phenotype and Hematopoietic Cytokine Expression of Umbilical Cord Mesenchymal Stem Cells

This study was aimed to investigate the change of cell phenotype and the expression of hematopoiesis associated cytokines in umbilical cord mesenchymal stem cells （UC-MSC） in three-dimensional （3-D） system.MSC were isolated from umbilical cord,and then cultured in 2-D and 3-D system respectively.The phenotype of MSC was detected by flow cytometry; the angiogenic capability of MSC cultured in 2-D and 3-D syitem was assessed using in vitro capillary formation assay.The cytokine expression of MSC in two kinds of culture conditions was measured by real-time PCR.The results showed that MSC were successfully isolated from umbilical cord.Flow cytometry showed that the percentage of CD31,CD133 and CD271 expressed in endothelial cells,endothelial progenitor cells and primitive mesenchymal stem cells increased significantly in 3-D culture conditions,as compared to 2-D system.Capillary formation assay showed that the angiogenic capability of UC-MSC was greatly enhanced.Quantitative PCR showed that the expression of β-actin was upregulated in 3-D system.The expression of some cytokines associated with hematopoiesis,such as G-CSF,LIF,SCF,IL-lα,IL-1β,IL-3,IL-7 and IL-11,increased,especially for LIF,IL-3,IL-7.The expression of IL-10 associated with immune regulation also increased.The expression of SDF-1,IL-6 slighdy decreased,but without significant difference.It is concluded that expression of CD31,CD133 and CD271 increases in 3-D system,the angiogenic capability of UC-MSC enhances and the expression of hematopoiesis-associated cytokines in UC-MSC increases in 3-D system.