College of Veterinary Medicine, Shandong Agricultural University, Tai'an, 271018, P. R. China.
Research Center for Animal Disease Control Engineering, Shandong Agricultural University, Tai'an, 271018, P. R. China.
J Microbiol. 2020 Apr;58(4):320-329. doi: 10.1007/s12275-020-9182-8. Epub 2020 Feb 26.
Bovine mastitis is a common disease in the dairy industry that causes great economic losses. As the primary pathogen of contagious mastitis, Staphylococcus aureus (S. aureus) can invade bovine mammary epithelial cells, thus evading immune defenses and resulting in persistent infection. Recently, autophagy has been considered an important mechanism for host cells to clear intracellular pathogens. In the current study, autophagy caused by S. aureus was detected, and the correlation between autophagy and intracellular S. aureus survival was assessed. First, a model of intracellular S. aureus infection was established. Then, the autophagy of MAC-T cells was evaluated by confocal microscopy and western blot. Moreover, the activation of the PI3K-Akt-mTOR and ERK1/2 signaling pathways was determined by western blot. Finally, the relationship between intracellular bacteria and autophagy was analyzed by using autophagy regulators (3-methyladenine [3-MA], rapamycin [Rapa] and chloroquine [CQ]). The results showed that S. aureus caused obvious induction of autophagosome formation, transformation of LC3I/II, and degradation of p62/SQSTM1 in MAC-T cells; furthermore, the PI3K-Akt-mTOR and ERK1/2 signaling pathways were activated. The number of intracellular S. aureus increased significantly with autophagy activation by rapamycin, whereas the number decreased when the autophagy flux was inhibited by chloroquine. Therefore, this study indicated that intracellular S. aureus can induce autophagy and utilize it to survive in bovine mammary epithelial cells.
奶牛乳腺炎是奶牛养殖业中的一种常见病，会造成巨大的经济损失。金黄色葡萄球菌（S. aureus）作为传染性乳腺炎的主要病原体，能够侵入奶牛乳腺上皮细胞，从而逃避免疫防御，导致持续感染。最近，自噬被认为是宿主细胞清除细胞内病原体的重要机制。本研究检测了金黄色葡萄球菌诱导的自噬，并评估了自噬与细胞内金黄色葡萄球菌存活的相关性。首先，建立了细胞内金黄色葡萄球菌感染模型。然后，通过共聚焦显微镜和 Western blot 评估 MAC-T 细胞的自噬。此外，通过 Western blot 测定 PI3K-Akt-mTOR 和 ERK1/2 信号通路的激活情况。最后，通过使用自噬调节剂（3-甲基腺嘌呤[3-MA]、雷帕霉素[Rapa]和氯喹[CQ]）分析细胞内细菌与自噬之间的关系。结果表明，金黄色葡萄球菌在 MAC-T 细胞中引起明显的自噬体形成、LC3I/II 的转化和 p62/SQSTM1 的降解；此外，PI3K-Akt-mTOR 和 ERK1/2 信号通路被激活。当用雷帕霉素激活自噬时，细胞内金黄色葡萄球菌的数量显著增加，而当用氯喹抑制自噬流时，细胞内金黄色葡萄球菌的数量减少。因此，本研究表明，细胞内金黄色葡萄球菌可以诱导自噬，并利用自噬在奶牛乳腺上皮细胞中存活。
