Clec14a基因调控斑马鱼胚胎血管发育

Clec14a gene contributes to zebrafish angiogenesis

目的:探索Clec14a在斑马鱼胚胎期的表达及其在血管发育过程中的功能,探讨该基因调控血管发育机制。方法:通过胚胎整体原位杂交技术,观察Clec14a基因在斑马鱼胚胎期的表达情况;在转基因斑马鱼系Tg(kdrl:mCherry)和Tg(fli1a:nGFP)中,显微注射吗啉修饰的反义寡核苷酸(morpholinoantisense oligos)下调Clec14a基因表达,同时在转基因斑马鱼系Tg(kdrl:mCherry)中注射Clec14a-mRNA调控Clec14a基因表达,利用激光共聚焦显微镜观察分析斑马鱼节间血管(intersegment vessel,ISV)表型,统计分析内皮细胞迁移速度和数目的变化,并应用qRT-PCR技术检测下调后该基因mRNA表达情况。结果:在受精后24 h、48 h和72 h,Clec14a基因在斑马鱼脑、心脏、血管系统中有表达;Clec14a基因表达下调导致ISV发育缺陷,内皮细胞数目减少,抑制其迁移速度。结论:下调Clec14a基因表达能够通过抑制内皮细胞增殖和内皮细胞迁移调控斑马鱼胚胎血管发育。

Objective:To investigate the expression of Clec14 a and its role in angiogenesis during embryonic period of the zebrafish. Methods:Whole mount in situ hybridization was performed to detect the expression pattern of Clec14 a.Clec14 aspecificmorpholino was injected respectively into the one-cell stage embryo to specifically knock-down expression of Clec14 a in Tg(kdrl:mCherry) and Tg(fli1 a:nGFP) transgenic lines. And Clec14 amRNA was injected respectively into the embryo to rescue the expression of Clec14 a in Tg(kdrl:mCherry). The morphology and endothelial cell number of intersegmental vessel(ISV) were analyzed after imaging by using the laser scanning confocal microscope. And qRT-PCR technology was used to detect the mRNA expression after the knock-down of Clec14 a. Results:Clec14 a was expressed in the brain, heart and vasculature of zebrafish larvae at 24, 48 and 72 hours post-fertilization(hpf). Knock-down of Clec14 a affected the development of ISV. Endothelial cell number was reduced after the knock-down of Clec14 a. Conclusions:Clec14 a controls zebrafish angiogenesis by affecting the endothelial cell proliferation and migration.