The effects of oxidative stress on blood vessels formation during zebrafish development

• Main Authors: Hai-Hong Syue, 薛海宏
• Other Authors: Chang-Yi, Wu
• Format: Others
• Language: zh-TW
• Published: 2015
• Online Access: http://ndltd.ncl.edu.tw/handle/uc29hx

碩士 === 國立中山大學 === 生物科學系研究所 === 103 === Redox and genetic signaling homeostasis are required for proper growth and patterning of blood vessels. Many antioxidant genes, such as Superoxide dismutase 1 (sod1), Glutaredoxin 2 (grx2), Peroxiredoxin 1 (prdx1) and Peroxiredoxin 2 (prdx2) have been shown important for angiogenesis. However, most studies have done in adult animals under pathological conditions, and very limited information about oxidative stress and vascular development during embryogenesis. Here, we report a novel function of prdx1 that play critical roles in vascular growth during zebrafish development. Prdx1 encodes a cytosolic thioredoxin-dependent peroxidase responsible for the remove of hydrogen peroxide and organic hydroperoxides. Amino acid sequence alignment and phylogenetic analysis of PRDX1 is highly conserved from yeast to mammalian. Our in situ hybridization results showed the expression pattern of prdx1 is in developing vessels. Knockdown of prdx1 by morpholino injection impairs the growth of intersegmental vessel (ISV) and cardinal vein plexus (CVP), suggesting the role of prdx1 in promoting ISV and CVP growth. To test the knockdown effect is prdx1 gene-specific, we examine the prdx1 morpholino knockdown efficiency and performed 2nd morpholino which block translational site of prdx1 results in similar vascular defects, suggesting the specificity of morpholino knockdown of prdx1. We further showed the reduction of ISV length and cell numbers in prdx1 morphants, indicating the vascular defects is due to a decrease of cell proliferation and migration, but not results from cell death in non-endothelial cells. To test molecular mechanisms of vascular defects that are associated with prdx1, we examined the expression of vascular markers. We found that a loss of prdx1 results in a decreased expression of vein/ ISV specific markers, flt4, mrc1, vascular markers stabilin and artery markers ephrin-B2. This indicates the regulatory role of prdx1 in controlling vascular development. Loss of prdx1 results in vascular defects, suggest the antioxidant function is important. Thus, we test if oxidative stress could cause vascular defects in H2O2-treated embryos. Our data showed H2O2 treatment only impaired CVP formation but no obvious ISV defect. Interestingly, while H2O2-treated embryo combined with knockdown of prdx1, synergetic effects is observed, suggested the oxidative stress can disturb vascular development. In addition, knockdown of prdx1causes the reduced expression of main antioxidant genes (sod1, sod2, catalase and prdx2) by qPCR analysis. Moreover, we found the increased expression of blood marker gata1 coincident with the decreased expression of endothelial markers flk1 and fli in prdx1 MO, suggested prdx1 likely regulate hemangioblast fate decision. Together, we showed a novel function of prdx1 that play critical roles in vascular growth during zebrafish development.