维甲酸对早期发育斑马鱼的心脏与骨骼毒性研究

Study on Cardiac and Skeletal Toxicity of Retinoic Acid to Danio rerio at the Early Life Stage

目的：研究维甲酸对早期发育斑马鱼心脏及骨骼的毒性。方法：将受精后24 h的斑马鱼胚胎作为毒性模型,分别暴露于含维甲酸0.1、1、10、25、100μmol/L的培养基中,分别于培养24、48 h后观察胚胎和幼鱼心脏形态的变化,并计算半数致死浓度（LC50）;将受精后4 d的斑马鱼幼鱼作为骨骼致畸模型,分别暴露于含维甲酸0.1、1、10、25、50μmol/L的培养基中,5 d后处死,采用茜素红对各组斑马鱼骨骼染色,并以显微镜采集图像,观察骨骼染色区域差异。结果：维甲酸对斑马鱼胚胎的孵化能力造成了明显不利影响,且胚胎在维甲酸暴露过程中均产生了明显的心脏致畸现象。1~100μmol/L维甲酸均对胚胎和幼鱼产生心脏致畸效应,胚胎和幼鱼表现出的主要畸形特征有心脏形态线性化、心包水肿、卵黄囊肿胀、血细胞在心区堆积。100μmol/L维甲酸可抑制胚胎的孵化能力,且对胚胎和幼鱼造成致死效应,暴露24、48 h后LC_（50）分别为36.44、23.69μmol/L。0.1~50μmol/L维甲酸均使斑马鱼幼鱼脊柱提前骨化,且骨化程度与维甲酸浓度呈正相关。结论：维甲酸对斑马鱼胚胎和幼鱼具有心脏和骨骼毒性,且与维甲酸的浓度呈正相关。

OBJECTIVE：To study the cardiac and skeletal toxicity of retinoic acid（RA） in Danio rerio at early life stage.METHODS：Danio rerio embryos of 24 hours post fertilization（hpf）were used as toxicity model and were exposed under medium with various concentrations of RA（0.1,1,10,25,100 μ mol/L）.The morphology of embryos and larvae hearts were observed24,48 h after exposed.LC50 was calculated.Danio rerio larvae of 4 days post fertilization（dpf）were used as skeletal deformity model and were exposed with a series of RA at various concentration（0.1,1,10,25,50 μmol/L）.They were sacrificed 5 d later,and then Danio rerio skeleton were fixed for staining with alizarin red.The microscopic was used to observe the difference of stained skeleton area.RESULTS：RA caused significant adverse effects on hatching capabilities of Danio rerio embryos,and the obvious malformation features were produced during the culture process.1-100 μmol/L RA could cause heart malformation in Danio rerio embryos and larvae,and the main heart malformation characteristics included heart linearization,pericardial edema,yolksacedema,hemocytes accumulation incardiac region.100 μmol/L RA could inhibit the hatching capabilities of Danio rerio embryos,and caused lethal effects on embryos and larvae.The LC_（50） were 36.44,23.69 μmol/L after exposed for 24,48 h.0.1-50 μmol/L RA induced vertebral column sclerotization of Danio rerio embryos and larvae in advance,which was positively associated with the concentration of RA.CONCLUSIONS：RA can cause cardiac and skeletal toxicity in Danio rerio embryo and larvae,which is positively associated with the concentration of RA.