[关键词]
[摘要]
目的: 探讨胶质瘤U87细胞培养上清诱导CD133+内皮细胞血管生成及其可能的机制。 方法: 磁珠法分选脐血CD133+细胞,体外诱导为CD133+内皮细胞并以共聚焦显微镜加以鉴定。U87细胞培养上清作用于CD133+内皮细胞(以DMEM作用为对照),体外血管生成实验检测其体外血管生成,Transwell法检测其迁移能力,Western blotting检测CD133+内皮细胞中VEGFR-1、VEGFR-2与基质金属蛋白酶9(matrix metalloproteinase,MMP-9)的表达,明胶酶谱检测MMP-9的活性。 结果: 脐血CD133+细胞体外诱导培养14 d后,约90%的细胞呈Dil-ac-LDL和FITC-UEA-1双阳性,具有内皮细胞特性,鉴定为CD133+内皮细胞。与DMEM对照组相比,U87细胞培养上清可诱导CD133+内皮细胞血管生成\[(40.7±3.3)vs(21.0±23),P<0.05\],促进CD133+内皮细胞迁移\[(0.60±0.04) vs (0.27±0.02),P<0.05\]。U87细胞培养上清上调CD133+内皮细胞中VEGFR-2与MMP-9的表达(P<0.05),而VEGFR-1的表达基本不变;U87细胞培养上清还可增强CD133+内皮细胞中MMP-9的酶活性。 结论: 胶质瘤U87细胞培养上清通过上调CD133+内皮细胞中VEGFR-2与MMP-9的表达促进内皮细胞血管生成。
[Key word]
[Abstract]
Objective : To investigate the angiogenesis of CD133+ endothelial cells induced by glioma U87 cell supernatant and the possible mechanisms. Methods: CD133+ cells were sorted from the cord blood by magnetic activated cell sorting system, and were further induced to CD133+ endothelial cells. CD133+ endothelial cells were indetified by confocal microscopy and treated with U87 cell supernatant (DMEM used as control), and their angiogenesis and migration were examined by in vitro angiogenesis assay and Transwell assay, respectively. VEGFR-1, VEGFR-2 and MMP-9 (matrix metalloproteinase 9) expressions were determined by Western blotting analysis, and MMP-9 activity was examined by gelatin zymography assay. Results: After in vitro cultured for 14 d, the cord blood CD133+ cells showed positive expression of Dil-ac-LDL and FITC-UEA-1 and had the features of endothelial cells, so they were named CD133+ endothelial cells. The supernatant of U87 glioma cells induced tube formation of CD133+ endothelial cells and increased migration of CD133+ endothelial cells. U87 cell supernatant induced angiogenesis (\[40.7±3.3\] vs \[21±2.3\], P<0.05), increased migration (\[0.60±0.04\] vs \[0.27±002\], P<0.05), up-regulated VEGFR-2 and MMP-9 expressions (P<0.05) in CD133+ endothelial cells, while had minimal effect on VEGFR-1 expression. Moreover, U87 cell supernatant enhanced MMP-9 activity of CD133+ endothelial cells. Conclusion: Glioma U87 cell supernatant can induce angiogenesis of CD133+ endothelial cell via up-regulating VEGFR-2 and MMP-9 expressions.
[中图分类号]
[基金项目]
江苏省医学重点人才基金资助项目(No.RC2007061)