In the present study, we demonstrated that bombyxin, an insect insulin-like peptide, modulated ecdysteroidogenesis in Bombyx mori prothoracic glands (PGs) through redox signaling. Our results showed that bombyxin treatment resulted in a transient increase in intracellular reactive oxygen species (ROS) concentration, as measured using 2',7'-dichlorofluorescin diacetate (DCFDA), an oxidation-sensitive fluorescent probe. The antioxidant N-acetylcysteine (NAC) abolished the bombyxin-induced increase in fluorescence in Bombyx PGs. Furthermore, bombyxin-induced ROS production was inhibited by mitochondrial oxidative phosphorylation inhibitors (rotenone and antimycin A), indicating mitochondria-mediated ROS production. The stimulation of ROS production in response to bombyxin appears to undergo development-specific changes. We further investigated the action mechanism of bombyxin-stimulated ROS signaling. Results showed that in the presence of either NAC, rotenone, or antimycin A, bombyxin-stimulated phosphorylation of insulin receptor, Akt, and 4E-binding protein (4E-BP) was blocked and bombyxin-stimulated ecdysteroidogenesis in PGs was greatly inhibited. From these results, we conclude that ROS signaling appears to be involved in bombyxin-stimulated ecdysteroidogenesis of PGs in B. mori by modulating the phosphorylation of insulin receptor, Akt, and 4E-BP. To our knowledge, this is the first demonstration of redox regulation in insulin signaling in an insect system.

中文翻译：

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活性氧介导的家蚕中的弹力毒素信号传导。

在本研究中，我们证明了虫毒素胰岛素样肽bombyxin通过氧化还原信号调节了家蚕前胸腺（PGs）中蜕皮甾类的生成。我们的研究结果表明，如使用氧化敏感的荧光探针2'，7'-二氯荧光素二乙酸盐（DCFDA）测得的那样，bombyxin处理导致细胞内活性氧（ROS）浓度瞬时增加。抗氧化剂N-乙酰半胱氨酸（NAC）消除了Bombyx PG中由虫毒素引起的荧光增强。此外，线粒体氧化磷酸化抑制剂（鱼藤酮和抗霉素A）抑制了虫毒素诱导的ROS生成，表明线粒体介导的ROS生成。响应蜂毒素的刺激ROS产生的刺激似乎发生了发育特定的变化。我们进一步研究了虫毒素刺激的ROS信号转导的作用机制。结果表明，在存在NAC，鱼藤酮或抗霉素A的情况下，胰高血糖素刺激的胰岛素受体，Akt和4E结合蛋白（4E-BP）的磷酸化被阻滞，并且胰高血糖素刺激的PG蜕皮类固醇生成被大大抑制。从这些结果，我们得出结论，ROS信号似乎通过调节胰岛素受体，Akt和4E-BP的磷酸化而参与了由棉球菌刺激的桑蚕中PGs的蜕皮类固醇生成。据我们所知，这是昆虫系统胰岛素信号传导中氧化还原调节的第一个证明。并阻止了4E结合蛋白（4E-BP）的产生，并极大地抑制了虫毒素刺激的PGs蜕皮类固醇生成。从这些结果，我们得出结论，ROS信号似乎通过调节胰岛素受体，Akt和4E-BP的磷酸化而参与了由棉球菌刺激的桑蚕中PGs的蜕皮类固醇生成。据我们所知，这是昆虫系统胰岛素信号传导中氧化还原调节的第一个证明。并阻止了4E结合蛋白（4E-BP）的产生，并极大地抑制了虫毒素刺激的PGs蜕皮类固醇生成。从这些结果，我们得出结论，ROS信号似乎通过调节胰岛素受体，Akt和4E-BP的磷酸化而参与了由棉球菌刺激的桑蚕中PGs的蜕皮类固醇生成。据我们所知，这是昆虫系统胰岛素信号传导中氧化还原调节的第一个证明。