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李倩,姜丝涛,李鲁美.2019.迷迭香酸对硫酸铜致斑马鱼胚胎毒性的抑制作用.动物学杂志,54(1):99-107.
迷迭香酸对硫酸铜致斑马鱼胚胎毒性的抑制作用
Inhibitory Effect of Rosmarinic Acid on the Developmental Toxicity Induced by Copper Sulfate in Zebrafish Embryos
投稿时间:2018-04-02  修订日期:2018-11-14
DOI:10.13859/j.cjz.201901012
中文关键词:  硫酸铜  迷迭香酸  斑马鱼  活性氧  毒性效应
英文关键词:CuSO4  Rosemary acid  Zebrafish, Danio rerio  Reactive oxygen  Toxic effect
基金项目:山东省高等学校科技立项经费项目(No. J17KA257)
作者单位E-mail
李倩 烟台大学生命科学学院 烟台 264005 ydskyshx@163.com 
姜丝涛 烟台大学生命科学学院 烟台 264005 hyshwx@163.com 
李鲁美 烟台大学生命科学学院 烟台 264005 736841269@qq.com 
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中文摘要:
      本文探讨硫酸铜(CuSO4)对斑马鱼(Danio rerio)胚胎发育的毒性效应,使用迷迭香酸(RA)抑制CuSO4对斑马鱼胚胎发育的毒性并探讨其作用机制。收集受精后1 h(1 hpf)的斑马鱼胚胎暴露于不同浓度的CuSO4溶液,或含有不同浓度迷迭香酸的CuSO4溶液,对照组培养在E3培养液中,观察胚胎死亡、孵化及畸形情况,计算胚胎死亡率、孵化率和畸形率;以活性氧(ROS)荧光探针DCFH-DA染色法检测迷迭香酸保护下胚胎的活性氧水平。对实验数据进行方差分析。结果显示:(1)CuSO4浓度超过一定量时能诱导斑马鱼胚胎死亡和畸形,胚胎孵化率也降低。CuSO4对96 hpf斑马鱼胚胎的半致死浓度(LC50)为7.7 μmol/L,半致畸浓度(EC50)为1.9 μmol/L。(2)在96 hpf,迷迭香酸与8 μmol/L CuSO4共同处理组斑马鱼胚胎的死亡率明显降低,孵化率升高。迷迭香酸与1.6 μmol/L CuSO4共同处理组斑马鱼胚胎的畸形率降低。(3)CuSO4单独处理组的活性氧含量明显高于迷迭香酸与CuSO4共同处理组和对照组。结果表明,CuSO4暴露对斑马鱼胚胎发育的毒性效应可能与活性氧升高导致的氧化应激相关;迷迭香酸抑制CuSO4对斑马鱼胚胎发育的毒性作用,可能与减少活性氧生成有关。
英文摘要:
      The purpose of this study was to investigate the developmental toxicity of copper sulfate (CuSO4) and the inhibitory effect of rosmarinic acid (RA) on the zebrafish (Danio rerio) embryos. The embryos (1 hour post fertilization, 1 hpf) were exposed to different concentrations of CuSO4 solution and CuSO4 solution containing different concentrations of RA, respectively. Control group was exposed to E3 medium. The mortality, malformation and hatching of embryos were observed under stereo microscope. The lethal rate, malformity rate and hatching rate were calculated and analysed. The embryonic reactive oxygen species (ROS) was detected using the fluorescent probe of DCFH-DA under RA defense. The experimental data were statistically analyzed with variance analysis. The results showed that the exposure to the CuSO4 solution caused serious lethal effect (Fig. 1a) and teratogentic effect (Fig. 1b, 2), decreased the hatching rate of embryos (Fig. 3) in a concentration-dependent manner. The linear regression equation between concentration of CuSO4 and mortality of zebrafish embryos at 96 hpf was Y = 5.59X + 7.1533, Y and X referred to lethal rate (%) and concentration of CuSO4 (μmol/L), respectively. The corresponding half lethal concentration (LC50) of CuSO4 on zebrafish embryos was 7.7 μmol/L. The linear regression equation between concentration of CuSO4 and deformity rate of zebrafish embryos at 96 hpf was Y = 24.213X + 3.5443, Y and X referred to lethal rate (%) and concentration of CuSO4 (μmol/L), respectively, and the half effective concentration (EC50) resulted in the malformation was 1.9 μmol/L. At 96 hpf, RA decreased lethal effect and increased hatching of zebrafish embryos induced by 8 μmol/L CuSO4 (Fig. 4). RA decreased malformation effect of zebrafish embryos induced by 1.6 μmol/L (Fig. 5). CuSO4 and RA inhibited ROS level induced by CuSO4 in zebrafish embryos (Fig. 6). The results indicate that CuSO4 has serious toxic effect on the development of zebrafish embryos, which may be associated with increased ROS level and caused oxidative stress. RA inhibits developmental toxicity of CuSO4 by reducing oxidative stress.
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