靛玉红抗斑马鱼胚胎血管生成活性机制研究

Indirubin inhibits angiogenesis in zebrafish embryos and its mechanism

目的:研究靛玉红抗血管生成作用及相关机制。方法:以tgfli1:eGFP(+/-)斑马鱼为模型,实验分空白对照组、二甲基亚砜组、靛玉红组(15μM、30μM、90μM)、阳性SU5416对照组,在12hpf时加药,于48hpf时在荧光显微镜下观察其体节间血管生成情况,并进行定量分析。运用整体原位杂交的方法和RT-PCR技术检测flk1基因的表达情况。结果:与空白对照组相比,靛玉红各浓度组体节间血管长度有显著性差异(P<0.01),并呈剂量依赖性抑制;与空白对照组相比,靛玉红对斑马鱼胚胎flk1mRNA表达有明显抑制作用,并随浓度的增高而增加。结论:靛玉红(15μM～90μM)具有抑制斑马鱼胚胎体节间血管生成的作用,其机制可能是通过阻断VEGF/VEGFR2信号通路的某个环节来发挥作用。

Objective： To investigate the antiangiogenic activity and related mechanism of indirubin in zebrafish embryos. Methods： The antiangio- genic activity was tested with in vivo transgenic zebrafish embryos, and its mechanism was clarified in zebra_fish by gene expression analysis of the vascular endothelial growth factor receptor2 （VEGFR2） . The expression of VEGFR2 was detected by RT-PCR assay and whole mount in situ hybridization （WISH）. Results： The treatment of zebrafish embryos with indirubin manifested potent anti-angiogenic activity against in- tersegmental vessels formation. WISH and semi-quantitative RT-PCR analysis showed that indirubin dose-dependently reduced the mRNA ex- pression of vascular endothelial growth factor （VEGF） receptor2 （VEGFR2）. Conclusions： The study suggested that indirubin could inhibit angiogenesis of intersegmental vessels of the zebrafish embryos through inhibition of the phosphorylation of certain protein in the VEGF/VEG- FR2 pathway, which results in down-regulation of VEGFR2 mRNA expression.