人脐带血淋巴管内皮祖细胞的分化及其生物学特征

目的研究脐带血中CD34^＋／CD133^＋／VEGFR-3^＋淋巴管内皮祖细胞经VEGF—C诱导向内皮细胞分化过程中生物学特征的变化，并探讨其分化的机制。方法取脐带血，用PercoU密度梯度离心法分离单形核细胞，再用流式细胞仪分选CD34^＋／CD133^＋／VEGFR-3^＋细胞，然后用VEGF—C诱导分化。在扫描电镜和透射电镜下观察细胞表面形态和细胞内结构的变化，并在激光扫描共焦显微镜下观察特征性标志物的表达变化。结果脐带血中的淋巴管内皮祖细胞表达CD34、CD133和VEGFR-3。CD34^＋／CD133^＋／VEGFR-3^＋细胞经VEGF-C诱导后7d，呈长梭形，细胞伸出板状伪足和丝状伪足，出现较多短的微绒毛。表面可见细胞小凹，细胞质中含有丰富的线粒体和粗面内质网。诱导后14d，细胞已具有内皮细胞的特征，表达淋巴管内皮特异性标志物LYVE-1和5-核苷酸酶，CD133表达消失，细胞质中可见Weibel-Palade小体。结论脐带血中存在CD34^＋／CD133^＋／VEGFR-3^＋淋巴管内皮祖细胞，这些细胞在VEGF—C诱导作用下可能通过VEGF—C／VEGFR-3信号途径分化为淋巴管内皮细胞。

DIFFERENTIATION AND BIOLOGICAL CHARACTERISTICS OF THE LYMPHATIC ENDOTHELIAL PROGENITOR CELLS ISOLATED FROM UMBILICAL CORD BLOOD

Objective To study the changes of biological characteristics of CD34~ /CD133~ /VEGFR-3~ lymphatic endothelial progenitor cells(EPCs) isolated from umbilical cord blood after induction with VEGF-C and investigate the mechanisms of EPCs differentiation toward lymphatic endothelial cells. Methods Mononuclear cells were isolated from umbilical cord blood using density centrifugation with Percoll solution.CD34~ /CD133~ /VEGFR-3~ cells were sorted with FACS.Differentiation of the cells was induced with VEGF-C.The ultrastructural changes of the cells were viewed under scanning and transmission electron microscopes.Changes in expression of markers of the cells were viewed under confocal laser scanning microscope. Results Lymphatic EPCs isolated from umbilical cord blood expressed CD34,CD133 and VEGFR-3.At 7 day after induction with VEGF-C,CD34~ /CD133~ /VEGFR-3~ cells were shuttle-like,there were a lamellipodium,numerous filopodia and many short microvilli.Caveolae were observed.Mitochondrion and rough endoplasmic reticulum were rich in cytoplasm.At 14 day after induction,the cells demonstrated appearance of the endothelial cell and expressed lymphatic endothelial specific makers LYVE-1 and 5'-nucleotidase,the expression of CD133 disappeared.Weibel-Palade body was observed in the cytoplasm of the cell.Conclusion There are CD34~ /CD133~ /VEGFR-3~ lymphatic EPCs in human umbilical cord blood.The cells may differentiate into lymphatic endothelial cells through VEGF-C/VEGFR-3 signaling pathway after induction with VEGF-C.