Zhang Shenshen, Liu Chuanrui, Li Yang, Imam Mustapha U, Huang Hui, Liu Haohao, Xin Yongjuan, Zhang Huizhen
Department of Environmental Health, College of Public Health, Zhengzhou University Zhengzhou, China.
Front Physiol. 2016 Nov 8;7:527. doi: 10.3389/fphys.2016.00527. eCollection 2016.
Microcystin-LR (MC-LR) is a ubiquitous peptide that exhibits strong reproductive toxicity, although the mechanistic basis for such toxicity remains largely unknown. The present study was conducted to investigate the mechanisms underlying the adverse effects of exposure to MC-LR in Chinese hamster ovary (CHO) cells. The results showed that MC-LR inhibited the proliferation of CHO cells significantly, with an IC of 10 μM. Moreover, MC-LR-treated CHO cells revealed strong induction of cell cycle arrest and apoptosis. Additionally, exposure of CHO cells to MC-LR resulted in excess reactive oxygen species production and intracellular calcium release, with resultant endoplasmic reticulum stress (ERs). There was also extensive accumulation of autophagic vacuoles with the highest concentration of MC-LR used (10 μM). Furthermore, the expression of ERs (GRP78, ATF-6, PERK, IRE1, CHOP) and autophagy (Beclin1 and LC3II) proteins was increased, with concomitantly reduced expression of LC3I suggesting that ERs and autophagy were induced in CHO cells by MC-LR treatment. Conversely, pretreatment of CHO cells with 4-Phenyl butyric acid, the ERs inhibitor reduced the MC-LR-induced apoptotic cell death and cellular autophagy as evidenced by the reduced expression of Beclin1 and LC3II. Similarly, MC-LR treatment in combination with an autophagy inhibitor (3-methyladenine) increased apoptotic cell death compared with MC-LR alone, and induced ERs upregulating ERs proteins. The overall results indicated that activation of ERs and autophagy are both associated with MC-LR-induced apoptosis in CHO cells. ERs may be a trigger of autophagy in this process.
微囊藻毒素-LR（MC-LR）是一种普遍存在的肽，具有很强的生殖毒性，尽管这种毒性的作用机制在很大程度上仍不清楚。本研究旨在探讨中国仓鼠卵巢（CHO）细胞暴露于MC-LR产生不良反应的机制。结果表明，MC-LR显著抑制CHO细胞的增殖，半数抑制浓度为10 μM。此外，经MC-LR处理的CHO细胞显示出强烈的细胞周期阻滞和凋亡诱导。另外，CHO细胞暴露于MC-LR会导致活性氧过量产生和细胞内钙释放，从而引发内质网应激（ERs）。在使用的最高浓度MC-LR（10 μM）下，自噬泡也大量积累。此外，ERs（GRP78、ATF-6、PERK、IRE1、CHOP）和自噬（Beclin1和LC3II）蛋白的表达增加，同时LC3I的表达降低，这表明MC-LR处理可诱导CHO细胞中的ERs和自噬。相反，用内质网应激抑制剂4-苯基丁酸预处理CHO细胞可减少MC-LR诱导的凋亡细胞死亡和细胞自噬，Beclin1和LC3II表达降低证明了这一点。同样，与单独使用MC-LR相比，MC-LR与自噬抑制剂（3-甲基腺嘌呤）联合处理增加了凋亡细胞死亡，并诱导ERs上调ERs蛋白。总体结果表明，ERs和自噬的激活均与MC-LR诱导的CHO细胞凋亡有关。在此过程中，ERs可能是自噬的触发因素。
