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Selenomethionine Inhibits NF-κB-mediated Inflammatory Responses of Bovine Mammary Epithelial Cells Caused by Klebsiella pneumoniae by Increasing Autophagic Flux

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Abstract

Klebsiella pneumoniae (K. pneumoniae) is one of the major pathogens causing bovine clinical mastitis. Autophagy maintains cellular homeostasis and resists excessive inflammation in eukaryotic organisms. Selenomethionine (Se-Met) is commonly used as a source of selenium supplementation for dairy cows. This study aimed to investigate the effects of Se-Met on inflammatory responses mediated by nuclear factor-kappa B (NF-κB) through autophagy. We infected bovine mammary epithelial cell line (MAC-T) with K. pneumoniae and examined the expression of autophagy-related proteins and changes in autophagic vesicles, LC3 puncta, and autophagic flux at various intervals. The results showed that K. pneumoniae activated the early-stage autophagy of MAC-T cells. The levels of LC3-II, Beclin1, and ATG5, as well as the number of LC3 puncta and autophagic vesicles, increased after 2 h post-treatment. However, the late-stage autophagic flux was blocked. Furthermore, the effect of autophagy on NF-κB-mediated inflammation was investigated with different autophagy levels. The findings showed that enhanced autophagy inhibited the K. pneumoniae-induced inflammatory responses of MAC-T cells. The opposite results were found with the inhibition of autophagy. Finally, we examined the effect of Se-Met on NF-κB-mediated inflammation based on autophagy. The results indicated that Se-Met alleviated K. pneumoniae-induced autophagic flux blockage, inhibited NF-κB-mediated inflammation, and decreased the adhesion of K. pneumoniae to MAC-T cells. The inhibitory effect of Se-Met on NF-κB-mediated inflammation could be partially blocked by the autophagy inhibitor chloroquine (CQ). Overall, Se-Met attenuated K. pneumoniae-induced NF-κB-mediated inflammatory responses by enhancing autophagic flux.

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Data Availability

The datasets generated and analyzed during the current study are not publicly available but are available from the corresponding author on reasonable request.

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Funding

This work was supported by the National Natural Science Foundation of China (32273070), the Open Project of International Research Laboratory of Prevention and Control of Important Animal Infectious Diseases and Zoonotic Diseases of Jiangsu Higher Education Institutions, the Jiangsu Agricultural Industry Technology System (No. JATS [2022] 499), the 333 High-level Talent Training Project of Jiangsu Province (CN), the Priority Academic Program Development of Jiangsu Higher Education Institutions (PAPD), and the 111 Project (D18007).

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Authors and Affiliations

  1. College of Veterinary Medicine, Yangzhou University, 225009, Yangzhou, China

    Siyan Xu, Yixue Miao, Junsheng Dong, Luying Cui, Kangjun Liu, Jianji Li, Xia Meng, Guoqiang Zhu & Heng Wang

  2. Jiangsu Co-Innovation Center for the Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Yangzhou, 225009, Jiangsu, China

    Siyan Xu, Yixue Miao, Junsheng Dong, Luying Cui, Kangjun Liu, Jianji Li, Xia Meng, Guoqiang Zhu & Heng Wang

  3. International Research Laboratory of Prevention and Control of Important Animal Infectious Diseases and Zoonotic Diseases of Jiangsu Higher Education Institutions, Yangzhou University, Yangzhou, 225009, Jiangsu, China

    Siyan Xu, Yixue Miao, Junsheng Dong, Luying Cui, Kangjun Liu, Jianji Li, Xia Meng, Guoqiang Zhu & Heng Wang

  4. Joint International Research Laboratory of Agriculture and Agri-Product Safety, The Ministry of Education of China, Yangzhou, 225009, Jiangsu, China

    Siyan Xu, Yixue Miao, Junsheng Dong, Luying Cui, Kangjun Liu, Jianji Li & Heng Wang

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  1. Siyan Xu
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  2. Yixue Miao
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Contributions

SX and HW designed the study. SX performed the main experiments and drafted the manuscript. YM, JD, and KL helped with the experiments. HW, JL, and JD acquired funding and supervised the study. HW, LC, XM, and GZ participated in the review and revision of the manuscript. All authors read and approved the final manuscript.

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Correspondence to Heng Wang.

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Xu, S., Miao, Y., Dong, J. et al. Selenomethionine Inhibits NF-κB-mediated Inflammatory Responses of Bovine Mammary Epithelial Cells Caused by Klebsiella pneumoniae by Increasing Autophagic Flux. Biol Trace Elem Res 202, 1568–1581 (2024). https://doi.org/10.1007/s12011-023-03757-2

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