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目的:利用斑马鱼模型对马兜铃酸进行心血管毒性的评价。方法:设置4个不同浓度的马兜铃酸溶液,同时设置空白对照组(养鱼水处理组)和溶剂对照组(0.1%DMSO)处理一定阶段的野生型AB系斑马鱼幼鱼至实验终点,每个浓度处理30尾斑马鱼。待测药物处理斑马鱼4 h后,随机选择10尾斑马鱼对心率进行计数。待测药物处理至实验终点时,在立体解剖显微镜下观察有无心包水肿、血流速度变化、血栓及出血等,计算各表型发生率。另设置4个不同浓度的马兜铃酸溶液,并分别设置空白对照组(养鱼水处理组)和溶剂对照组(0.9%Na Cl注射液)处理组,至实验终点每个浓度收集100尾斑马鱼(-80℃贮存),提取mRNA反转成c DNA样本后,进行荧光定量RT-PCR检测斑马鱼组织中MicroRNA146a的表达。结果:当马兜铃酸浓度≥MNLC/3时,可诱发斑马鱼心血管毒性,并呈现浓度相关性增加,表现为血流变慢或(和)缺失、静脉窦处瘀血、静脉窦处水肿和心率变慢(仅LC10处理组),未见心律不齐。荧光定量RT-PCR结果显示,马兜铃酸的4个浓度MNLC10、MNLC/3、MNLC、LC10均显著提高MicroRNA146a的表达(与对照组比较P<0.01),且存在一定的浓度关系。结论:马兜铃酸具有一定的心血管毒性作用,且具有浓度相关性。
OBJECTIVE: To evaluate the cardiovascular toxicity of aristolochic acid using a zebrafish model. Methods: Four different concentrations of aristolochic acid solution were set up. At the same time, blank control group (fish-water treatment group) and solvent control group (0.1% DMSO) 30 zebra fish were treated at each concentration. After treatment of the zebrafish with the drug to be tested for 4 h, ten zebrafish were randomly selected to calculate the heart rate. Test drug treatment to the end of the experiment, observed under stereotactic microscope with or without pericardial edema, changes in blood flow velocity, thrombosis and bleeding, calculate the incidence of each phenotype. Another four different concentrations of aristolochic acid solution were set and blank control group (fish water treatment group) and solvent control group (0.9% Na Cl injection) treatment group to the end of the experiment for each concentration of 100 Zebrafish (-80 ℃ storage), mRNA extraction inverted c c DNA samples, fluorescence quantitative RT-PCR detection of zebrafish MicroRNA146a expression. Results: When aristolochic acid concentration ≥ MNLC / 3, zebrafish could induce cardiovascular toxicity and showed a concentration-related increase, manifested as slow or (or) loss of blood flow, blood stasis in sinus venous sinus Edema and heart rate slowed (LC10-only group), no arrhythmia. Fluorescent quantitative RT-PCR results showed that four concentrations of aristolochic MNLC10, MNLC / 3, MNLC and LC10 significantly increased the expression of MicroRNA146a (P <0.01 compared with the control group), and there was a certain concentration relationship. Conclusion: Aristolochic acid has certain cardiovascular toxicity and concentration-relatedness.