利用胚胎干细胞试验模型评价矮壮素的发育毒性

doi:10.3969/j.issn.1004-616x.2015.04.002

癌变·畸变·突变 ›› 2015, Vol. 27 ›› Issue (4): 260-265,271.doi: 10.3969/j.issn.1004-616x.2015.04.002

利用胚胎干细胞试验模型评价矮壮素的发育毒性

孔丹^1,2, 郝卫东¹, 蒋建军¹, 尚兰琴¹, 魏雪涛¹, 时褔礼²

1. 北京大学公共卫生学院, 食品安全毒理学研究与评价北京市重点实验室, 北京 100191;
2. 北京市海淀区卫生局卫生监督所, 北京 100037

收稿日期:2015-02-02 修回日期:2015-06-08 出版日期:2015-07-30 发布日期:2015-07-30
作者简介:孔丹,E-mail:kong_d@163.com。
基金资助:
国家自然科学基金(81473002);北京市科技创新基地培育与发展专项(Z151100001615035)

Developmental toxicity assessment of chlormequat chloride using embryonic stem cell test

KONG Dan^1,2, HAO Weidong¹, JIANG Jianjun¹, SHANG Lanqin¹, WEI Xuetao¹, SHI Fuli²

1. School of Public Health, Peking University, Beijing Key Laboratory of Toxicological Research and Risk Assessment for Food Safety, Beijing 100191;
2. The Bureau for Health Inspection and Supervision of Haidian District, Beijing 100037, China

Received:2015-02-02 Revised:2015-06-08 Online:2015-07-30 Published:2015-07-30
Contact: 郝卫东,E-mail:whao@bjmu.edu.cn;Tel:010-82802352 E-mail:whao@bjmu.edu.cn

摘要/Abstract

摘要： 目的: 利用胚胎干细胞试验模型评价矮壮素的发育毒性。方法:以DMSO为阴性对照,使用不同剂量(7.8125、15.625、31.25、62.5、125、250、500、1 000 μg/mL)的矮壮素染毒小鼠胚胎干细胞D3(ES-D3)和3T3成纤维细胞,观察矮壮素对ES-D3细胞向心肌细胞分化的影响,以及对ES-D3和3T3细胞的细胞毒性,在此基础上,根据发育毒性判别公式对矮壮素致畸性进行判定。提取ES-D3分化抑制试验中的拟胚体总RNA,进行real-time PCR,在基因水平检测矮壮素对ES-D3向各胚层分化的影响。结果:1 000 μg/mL矮壮素染毒可明显促进体外培养ES-D3、3T3细胞增殖;250 μg/mL的矮壮素作用下,观察到具有心肌细胞收缩的拟胚体数仅占接种拟胚体总数的69.4%;1 000 μg/mL矮壮素能够影响ES-D3向心肌细胞分化的基因标志物(Nkx2.5,α-MHC)的表达。结论:无法利用判别公式对矮壮素进行发育毒性评价;矮壮素在不引起3T3、ES-D3细胞毒性的水平下,影响ES-D3向心肌细胞的分化,有产生胚胎毒性的可能。

关键词: 矮壮素, 植物生长调节剂, 发育毒性, 胚胎干细胞试验模型

Abstract: OBJECTIVE: To investigate the developmental toxicity and underlying mechanism of chlormequat chloride by utilizing embryonic stem cell test in vitro. METHODS: Mouse embryonic stem cells and 3T3 cells were treated with different doses of chlormequat chloride,the effects on ES and 3T3 cell proliferation and ES cell differentiation were determined. A cascade of genes related to normal cardiac development was examined by real time PCR. RESULTS: Chlormequat chloride (1 000 μg/mL) could evidently increase the viability of mouse fibroblasts 3T3 cells and mouse embryonic stem cells. Chlormequat chloride could influence embryonic stem cell cardiac differentiation in a U shape manner. Compared to the control group,only 69.4% beating EBs was observed at 250 μg/mL concentration. Chlormequat chloride was able to cause effect on gene expression during cardiogenesis,especially Nkx2.5 and α-MHC. CONCLUSION: Chlormequat chloride could not be classified according to the prediction model recommended by ECVAM. Chlormequat chloride at 250 μg/mL concentration could retard embryonic stem cell differentiation into myocardial cell without causing any morphological malformations of 3T3 and ES-D3. Chlormequat chloride could alter the expression of crucial genes during the early stages of cardiogenesis,and these effects might result in adverse developmental outcomes.

Key words: chlormequat chloride, plant growth regulators, developmental toxicity, embryonic stem cell test model

中图分类号:

R394.6

孔丹, 郝卫东, 蒋建军, 尚兰琴, 魏雪涛, 时褔礼. 利用胚胎干细胞试验模型评价矮壮素的发育毒性[J]. 癌变·畸变·突变, 2015, 27(4): 260-265,271.

KONG Dan, HAO Weidong, JIANG Jianjun, SHANG Lanqin, WEI Xuetao, SHI Fuli. Developmental toxicity assessment of chlormequat chloride using embryonic stem cell test[J]. Carcinogenesis, Teratogenesis & Mutagenesis, 2015, 27(4): 260-265,271.

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利用胚胎干细胞试验模型评价矮壮素的发育毒性

Developmental toxicity assessment of chlormequat chloride using embryonic stem cell test

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