T-2 toxin inhibits murine ES cells cardiac differentiation and mitochondrial biogenesis by ROS and p-38 MAPK-mediated pathway
T-2 toxin inhibits murine ES cells cardiac differentiation and mitochondrial biogenesis by ROS and p-38 MAPK-mediated pathway
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T-2 毒素通过 ROS 和 p-38 MAPK 介导的途径抑制小鼠 ES 细胞心脏分化和线粒体生物合成。
DOI：
10.1016/j.toxlet.2016.06.2103
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发表时间：
2016-09-06
影响因子：
3.5
通讯作者：
Peng, Shuangqing
中科院分区：
文献类型：
--
作者：
Fang, Haiqin;Cong, Liangzi;Peng, Shuangqing
目的：探讨T-2毒素对体外培养的小鼠胚胎干细胞(ESCs)心脏分化和线粒体生物发生的影响。将EBS暴露于0.5 ng/mlT-2毒素中24、72和120h，每天观察培养物中收缩簇的出现，并用Western印迹和免疫细胞化学方法检测心肌特异性蛋白(α-Aciniin)。用激光共聚焦扫描显微镜和透射电子显微镜观察线粒体超微结构。用H_2二氯荧光素-二乙酸酯(H_2DCF-DA)监测细胞内活性氧(ROS)。Western印迹分析p38(p-p38)和p38丝裂原活化蛋白激酶(MAPK)的磷酸化及线粒体生物发生蛋白的表达，包括PGC-1α(PGC-1α)、核呼吸因子1(NRF-1)、线粒体转录因子A(MtTFA)和线粒体呼吸链复合体IV(COXIV)。在一些实验中，MESCs用抗氧化剂Trolox(200 MU M)预处理30min，然后再暴露于Trolox(200 MU M)和T-2毒素(0.5 ng/ml)72h。结果：显微镜下观察到收缩的簇状结构，心脏特异性蛋白(α-actinin)阳性表达表明MESCs直接分化为心肌细胞。而T-2毒素处理72和120h可抑制小鼠ES细胞的心脏分化。P-p38的表达在24 h组显著升高，72、120 h组显著降低。T-2毒素处理后，线粒体数量减少，PGC-1α、NRF-1、mtTFA和COXIV等线粒体生物发生相关蛋白表达减少，且呈时间依赖性。结论：T-2毒素通过降低PGC-1α、NRF-1和mtTFA的表达，抑制线粒体的生物发生，进而抑制小鼠ES细胞的心脏分化，其作用部分与ROS介导的p38MAPK有关。(C)2016爱思唯尔爱尔兰有限公司。保留所有权利。
Objective: To investigate the effect of T-2 toxin on murine embryonic stem cells (ESCs) cardiac differentiation and mitochondrial biogenesis in vitro.Methods: Cardiac differentiation of the mouse ESCs was initiated by embryoid bodies (EBs) formation in hanging drops. EBs were exposed to 0.5 ng/ml T-2 toxin for 24, 72 and 120 h. Cultures were observed daily for the appearance of contracting clusters, and cardiac-specific protein (alpha-actiniin) were measured by Western blot and immunocytochemistry. Mitochondrial ultrastructure was observed by confocal laser scanning microscopy and transmission EM photography. Reactive oxygen species (ROS) was monitored by H2-dichlorofluorescein- diacetate (H2DCF-DA). The phosphorylation of the p38 (p-p38) and p38 mitogen-activated protein kinase (MAPK) and the expression of mitochondrial biogenesis proteins, including peroxisome proliferator activated receptor coactivator-1 alpha (PGC-1 alpha), nuclear respiratory factor 1 (NRF-1), mitochondrial transcription factor A (mtTFA), and mitochondrial respiratory chain complex IV (COXIV) were analyzed using Western blot. In some experiments, mESCs were pre-treated with the antioxidant Trolox (200 mu M) for 30 min, then exposed to Trolox (200 mu M) and T-2 toxin (0.5 ng/ml) for 72 h.Results: Contracting clusters were observed under the microscope light and cardiac-specific protein (alpha-actinin) expressed positively indicated mESCs directly differentiated in cardiomyocytes. However, the cardiac differentiation was inhibited by T-2 toxin treatment 72 and 120 h. ROS accumulated in murine ES cells in a time-dependent manner. The expression of p-p38 significantly increased in 24 h group and decrease in 72 and 120 h groups. The decrease of mitochondrial number and the mitochondrial biogenesis-related proteins expression, including PGC-1 alpha, NRF-1, mtTFA, and COXIV decreased in a time-dependent manner with T-2 toxin treatment. However, the inhibition of mitochondrial biogenesis by T-2 toxin in differentiated mESCs was recovered significantly in the presence of the antioxidant Trolox.Conclusion: Taken together, T-2 toxin decreased the expression of PGC-1 alpha, NRF-1, and mtTFA, inhibited mitochondrial biogenesis, and then inhibited the cardiac differentiation of murine ES cells, and the effect was partly responsible for the p38 MAPK mediated by ROS. (C) 2016 Elsevier Ireland Ltd. All rights reserved.