PEI介导 VEGFR-3 siRNA 转染对 VEGF-C诱导LEPCs分化的影响

目的：应用聚乙烯亚胺（ polyethyleneimine ， PEI ）介导淋巴管内皮祖细胞（ lymphatic endothelial progenitor cells ， LEPCs ） VEGFR-3基因 siRNA，体外沉默 LEPCs VEGFR-3基因表达，观察VEGFR-3siRNA对VEGF-C诱导LEPCs分化的影响。方法采用Ficoll法分离人脐血单个核细胞，流式细胞仪、免疫荧光分选并鉴定 LEPCs。 PEI 包裹前期实验抑制效果最好的 VEGFR-3siRNA 体外转染入LEPCs，荧光显微镜和流式细胞仪检测转染效率。流式细胞仪检测经 VEGF-C 诱导7天各组 LEPCs CD133＋细胞所占比例，14天后各组LYVE-1＋细胞所占比例。结果 VEGFR-3siRNA成功转染入LEPCs，转染效率30％，与lipofectimine转染效率无差异，VEGFR-3siRNA转染组7天的LEPCs CD133＋细胞所占比例明显高于VEGF-C诱导组（P＜0．05），而14天后两组之间LYVE-1＋细胞所占比例无显著差异（P＞0．05）。结论体外经由PEI介导的siRNA能有效的沉默LEPCsVEGFR-3的表达，从而抑制VEGF-C诱导的LEPCs分化。

Effect of interference mediated by PEI/VEGFR-3 siRNA on differentiation of lymphatic endothelial progenitor cell after inducted with VEGF-C

Objective To study the gene silencing efficiency of siRNA targeted against human VEGFR-3 （VEGFR-3 siRNA） of lymphatic endothelial progenitor cells （LEPCs） in vitro mediated by polyethyleneimine （PEI）,and to observe the effect on differentiation of LEPCs after inducted with VEGF-C.Methods The mononuclear cells of Umbilieal cord blood were isolated using density centrifugation with Ficoll solution .LEPCs were sorted and viewed using flow cytometer and confocal laser scanning microscope ,respectively.PEI mediated VEGFR-3 siRNA transfecting LEPCs .Fluorescence microscopy and flow cytometry for transfection efficiency .The expression of CD133 was detected in each groups by flowcytometer after inducted with VEGF-C for 7 days, the same method was used in the expression of LYVE-1after 14 days.Results VEGFR-3 siRNA/PEI were transected to LEPCs , the transfection efficiency was 30%, there was no difference compared with lipofectimine . The expression of CD133 was higher than control groups inducted with VEGF-C, The expressions of LYVE-1 have no difference between two groups .Conclusions The PEI could effectively transfect siRNA into cells and silence the VEGFR-3 gene expression of LEPCs , then differentiation of LEPCs was inhibited by VEGFR-3 siRNA after inducted with VEGF-C.