趋化因子SDF-1趋化作用的实验研究

发布时间：2018-04-03 02:07

本文选题：骨髓间质干细胞　切入点：胶质细胞　出处：《福建医科大学》2005年硕士论文

【摘要】：目的研究趋化因子SDF-1α在体内外对原代培养的骨髓间质干细胞、星形胶质细胞和N9 小胶质细胞系以及分离的中性粒细胞的趋化作用,从而探讨脑损伤和细胞移植时SDF-1α在细胞迁移中的可能机制。方法体外实验利用SDF-1α作为趋化因子加入到48孔微迁移板下室,把上述四种细胞加入到上室,抗体阻断实验把这四种细胞与抗CXCR4 多抗共孵育后加入到上室,观察细胞迁移,计算细胞的迁移指数。同时用RT-PCR 方法观察N9 小胶质细胞CXCR4 基因表达的情况。体内实验采用大鼠大脑中动脉线栓模型,把骨髓间质干细胞、中性粒细胞以及N9 小胶质细胞与抗CXCR4 多抗共孵育后的这3 种细胞分别通过大鼠尾静脉移植入体内,脑组织切片后荧光显微镜下观察脑损伤部位植入细胞的趋化情况。结果体外实验时,SDF-1α作用后,骨髓间质干细胞、N9 小胶质细胞和中性粒细胞跨膜迁移的数目较对照组明显增多,而星形胶质细胞迁移的数目未见明显变化。抗CXCR4 多抗孵育后骨髓间质干细胞、N9 小胶质细胞和中性粒细胞跨膜迁移的数目则明显减少。RT-PCR 方法证实在正常情况下N9 小胶质细胞有明显的CXCR4 表达。体内实验时,有较多的骨髓间质干细胞、N9 小胶质细胞和中性粒细胞迁移到病灶部位,而空白对照组未见到迁移的细胞,阴性对照组仅见个别迁移的细胞。结论趋化因子SDF-1α能通过受体CXCR4 发挥对骨髓间质干细胞、N9 小胶质细胞和中性粒细胞的趋化作用,对星形胶质细胞没有趋化作用。在体内,损伤的脑组织可能主要通过分泌SDF-1α增多作用于CXCR4 从而对炎症细胞、小胶质细胞或移植的骨髓间质干细胞发挥趋化作用。
[Abstract]:Objective to investigate the chemotactic effects of chemokine SDF-1 伪 on primary cultured bone marrow mesenchymal stem cells, astrocytes, N9 microglial cells and isolated neutrophils in vitro and in vivo.To explore the possible mechanism of SDF-1 伪 in cell migration during brain injury and cell transplantation.Methods in vitro, SDF-1 伪 was used as chemokine in the 48 well micromigration chamber, and the above four cells were added to the upper chamber. The antibody blocking experiment incubated the four kinds of cells with anti CXCR4 polyclonal antibodies and then added them to the upper chamber to observe the migration of the cells.Cell migration index was calculated.At the same time, the expression of CXCR4 gene in N 9 microglial cells was observed by RT-PCR method.Bone marrow mesenchymal stem cells, neutrophilic granulocytes and N9 microglial cells were transplanted into the rat caudal vein after incubating with anti CXCR4 polyclonal antibodies in vivo.The chemotaxis of implanted cells in brain injury site was observed under fluorescence microscope after brain tissue section.Results the number of N9 microglia and neutrophil transmembrane migration of bone marrow mesenchymal stem cells was significantly increased after SDF-1 伪 treatment in vitro, but the number of astrocytes did not change.The number of transmembrane migration of N9 microglia and neutrophils in bone marrow mesenchymal stem cells (BMSCs) incubated with anti CXCR4 polyclonal antibodies was significantly decreased. RT-PCR showed that N9 microglia expressed CXCR4 in normal condition.In vivo, more bone marrow mesenchymal stem cells (BMSCs) N9 microglia and neutrophils migrated to the lesion site, but no migrating cells were found in the blank control group, while only individual migrating cells were found in the negative control group.Conclusion chemokine SDF-1 伪 can exert chemotaxis on N9 microglia and neutrophils in bone marrow mesenchymal stem cells through receptor CXCR4, but has no chemotactic effect on astrocytes.In vivo, the injured brain tissue may play a chemotaxis effect on inflammatory cells, microglia cells or bone marrow mesenchymal stem cells by secreting SDF-1 伪 increase in CXCR4.
【学位授予单位】：福建医科大学
【学位级别】：硕士
【学位授予年份】：2005
【分类号】：R341

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