三氯乙烯对斑马鱼胚胎心脏发育的毒性作用及机制*

doi:10.12047/j.cjap.5787.2019.073

中国应用生理学杂志 ›› 2019, Vol. 35 ›› Issue (4): 346-350.doi: 10.12047/j.cjap.5787.2019.073

三氯乙烯对斑马鱼胚胎心脏发育的毒性作用及机制^*

史佳林¹, 夏颖¹, 花月¹, 张可佳¹, 陈涛^2△, 姜岩^1△

1. 苏州大学医学部, 江苏苏州 215123;
2. 江苏省老年疾病预防转化医学重点实验室, 苏州大学, 江苏苏州 215123

收稿日期:2018-11-22 出版日期:2019-07-28 发布日期:2019-11-06
通讯作者: ^△,Tel: 18260195811, 15950069815; E-mail: tchen@suda.edu.cn, yjiang@suda.edu.cn
基金资助:
^*国家自然科学基金项目(81570284);江苏省级大学生创新创业训练计划项目(201610285098X);苏州市应用基础研究计划(SYS201519)

Trichloroethylene interferes with heart development of zebrafish via inhibiting Wnt signal pathway

SHI Jia-lin ¹, XIA Ying ¹, HUA Yue ¹, ZHANG Ke-jia ¹, CHEN Tao ^2△, JIANG Yan ^1△

1. Medical College of Soochow University, Suzhou 215123;
2. Jiangsu Key Laboratory of Preventive and Translational Medicine for Geriatric Diseases, Soochow University, Suzhou 215123, China

Received:2018-11-22 Online:2019-07-28 Published:2019-11-06

摘要/Abstract

摘要： 目的:研究三氯乙烯(TCE)对斑马鱼胚胎心脏发育的毒性作用及其机制,为寻找干预靶点提供实验依据。方法:斑马鱼胚胎来自于国家斑马鱼资源中心,分为DMSO组(对照组)、DMSO+CHIR组、DMSO+XAV组、TCE处理组、TCE+CHIR组和TCE+XAV组(TCE设置为1、10、100 ppb三个不同的浓度;DMSO:二甲基亚砜;CHIR:CHIR-99021,Wnt信号通路激活剂;XAV:XAV-939,Wnt信号通路抑制剂),每组60条。斑马鱼胚胎饲养于系统养殖水中,恒温28℃,每隔24 h更换养殖水,并分别加入相应药物。连续培养72 h,收集斑马鱼胚胎的心脏组织,提取RNA进行转录组芯片分析,并以荧光定量PCR验证Wnt信号通路相关基因的表达。结果:与对照组相比,三氯乙烯暴露导致斑马鱼心脏畸形显著增加,以心房心室比例异常、环化不全以及心包水肿等为主要表型。芯片分析结果显示,TCE处理组Wnt信号通路相关基因(Axin2、Sox9b、Nkx2.5)表达受到显著影响。qPCR结果进一步验证,TCE处理组与DMSO对照组相比,Wnt通路靶基因Axin2、Sox9b及Nkx2.5的mRNA水平显著下调(P＜0.05),提示Wnt信号通路被抑制。Wnt激活剂CHIR降低TCE导致的斑马鱼胚胎心脏发育异常,而添加Wnt通路抑制剂XAV后,斑马鱼胚胎心脏畸形率显著增加(P＜0.05)。结论:三氯乙烯暴露导致斑马鱼胚胎心脏畸形,Wnt信号通路参与三氯乙烯的心脏发育毒性。

关键词: 斑马鱼胚胎, 三氯乙烯, Wnt信号通路, 心脏发育

Abstract: Objective: To investigate the molecular mechanism of trichloroethylene (TCE) cardiac developmental toxicity on zebrafish embryos and to try to provide experimental data for related intervention. Methods: Zebrafish embryos were purchased from the National Zebrafish Resource Center. The embryos were divided into DMSO(control group), DMSO+CHIR, DMSO+XAV, TCE, TCE+CHIR and TCE+XAV groups(TCE at the concentration of 1, 10 and 100 ppb, with the DMSO as control; DMSO: Dimethyl suldoxide; CHIR: CHIR-99021, Wnt agonist; XAV: XAV-939, Wnt antagonist), 60 embryos per group. Zebrafish embryos were fed in systematic aquaculture water, 28℃. The water was replaced every 24 h and drugs were added according to the grouping scheme. The cardiac tissues were dissected and analyzed by transcriptome microarray after RNA extraction. The expressions of Wnt signaling pathway related genes were verified by q-PCR. Wnt atagonist XAV and activator CHIR were used alone or in combination to further evaluate the possibility of the Wnt signaling participating in the cardiac developmental toxicity induced by TCE. Results: Compared with control, Zebra fish embryos exposed to TCE showed a significant increase in heart defects, and the main phenotypes were abnormal atrioventricular ratio, looping defects and pericardial edema. The results of microarray profiling showed that the expressions of genes related to Wnt signaling pathway were affected significantly. The results of qPCR further confirmed that TCE inhibited the expressions of Wnt pathway target genes Axin2, Sox9b and Nkx2.5(P＜0.05). Wnt agonist CHIR reduced the TCE-induced cardiac malformation rate significantly, while the addition of Wnt antagonist XAV markedly enhanced the cardiac developmental toxicity of TCE. Conclusion: Exposure to TCE leads to heart malformation in zebrafish embryos. Wnt signaling pathway may be involved in the cardiac developmental toxicity induced by TCE.

Key words: zebrafish embryos, trichloroethylene, Wnt signal pathway, cardiac development

史佳林, 夏颖, 花月, 张可佳, 陈涛, 姜岩. 三氯乙烯对斑马鱼胚胎心脏发育的毒性作用及机制^*[J]. 中国应用生理学杂志, 2019, 35(4): 346-350.

SHI Jia-lin, XIA Ying, HUA Yue, ZHANG Ke-jia, CHEN Tao, JIANG Yan. Trichloroethylene interferes with heart development of zebrafish via inhibiting Wnt signal pathway[J]. CJAP, 2019, 35(4): 346-350.

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三氯乙烯对斑马鱼胚胎心脏发育的毒性作用及机制^*

Trichloroethylene interferes with heart development of zebrafish via inhibiting Wnt signal pathway

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