Department of Environmental Sciences, Hangzhou Normal University, Xuelin Road 16#, Xiasha Gaojiao Dongqu, Hangzhou, Zhejiang Province 310036, China.
Fish Shellfish Immunol. 2012 Dec;33(6):1229-37. doi: 10.1016/j.fsi.2012.08.016. Epub 2012 Aug 24.
Nodularin, a metabolite of Nodularin spumigena, is widely detected in water blooms worldwide and causes serious negative effects on fish. The apoptosis-related cytotoxic effects and mechanisms of nodularin on Carassius auratus lymphocytes were investigated. Transmission electron microscopy results showed that nodularin-treated lymphocytes display a series of morphological changes, including condensed cytoplasm, nuclear chromatin agglutination and marginalization. DNA fragmentation was verified by the DNA-ladder and formation of sub-G1 DNA peaks. These cell characteristics confirmed the occurrence of apoptosis in lymphocytes. Flow cytometric results showed that the percentages of apoptotic cells incubated with 1, 5, 10, and 100 μg/L nodularin for 12 h reached 15.76%, 17.36%, 20.34% and 44.21%, respectively; controls showed low rates of apoptosis (2.4%). The mechanism of apoptosis induced by nodularin was determined, and results showed that nodularin exposure caused a significant increase in intracellular reactive oxygen species (ROS), loss of mitochondrial transmembrane potential in a dose-dependent manner, upregulation of intracellular Ca²⁺, downregulation of Bcl-2 and upregulation of Bax expression at the mRNA and protein levels, and activation of caspase-3 and caspase-9 without caspase-8. In summary, all the results suggest that nodularin induces lymphocyte apoptosis via the mitochondrial apoptotic pathway and destroys the immune response of fish.
微囊藻毒素-N，是微囊藻属的一种代谢产物，在世界各地的水华现象中广泛存在，并对鱼类造成严重的负面影响。本研究旨在探讨微囊藻毒素-N 对鲤鱼淋巴细胞的细胞凋亡相关细胞毒性作用及机制。透射电子显微镜结果显示，经微囊藻毒素-N 处理的淋巴细胞呈现一系列形态学变化，包括细胞质浓缩、核染色质凝聚和边缘化。通过 DNA 梯状带和亚 G1 峰的形成验证了 DNA 片段化。这些细胞特征证实了淋巴细胞发生了凋亡。流式细胞术结果显示，经 1、5、10 和 100μg/L 微囊藻毒素-N 孵育 12 h 的细胞凋亡率分别达到 15.76%、17.36%、20.34%和 44.21%；对照组的凋亡率较低（2.4%）。确定了微囊藻毒素-N 诱导细胞凋亡的机制，结果表明，微囊藻毒素-N 暴露导致细胞内活性氧（ROS）显著增加，线粒体跨膜电位呈剂量依赖性下降，细胞内 Ca²⁺水平升高，Bcl-2 表达下调，Bax 表达上调，同时 caspase-3 和 caspase-9 被激活，而 caspase-8 无变化。综上所述，所有结果表明，微囊藻毒素-N 通过线粒体凋亡途径诱导淋巴细胞凋亡，破坏鱼类的免疫反应。
