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An Investigation of the Innate Immune Response in Bovine Mammary Epithelial Cells Challenged by Prototheca zopfii

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Abstract

Prototheca zopfii is an important bovine mastitis pathogen, which could result in severe mammary infection. However, the innate immune response in bovine mastitis associated with P. zopfii was not clear. Therefore, the aim of this study is to investigate in vitro innate immune responses implicated by P. zopfii. Bovine mammary epithelial cells (bMECs) were infected with 5.0 × 104 cells/ml P. zopfii genotypes I and II independently, and the mRNA expression of TLR-2, TLR-4, TNF-α, IL-1β, IL-8, NOD-1, NOD-2 and β-defensin-5 was determined by real-time polymerase chain reaction (RT-PCR) over a time course of 1, 3, 6, 12 and 24 h. The detection of the NF-κB p65 protein in nucleus and cytoplasm of infected bMECs over the same time course was evaluated. Results showed that P. zopfii genotype II has ability to up-regulate the expression of TLR-2, TLR-4, TNF-α, IL-1β, IL-8, NOD-1, NOD-2 and β-defensin-5 'more strongly than genotype I. Western blot results showed that when bMECs were challenged by P. zopfii genotype II, the expression of NF-κB p65 protein in the nucleus was up-regulated, while in cytoplasm it appeared to be repressed, which indicated that bMECs partly regulate the innate immune responses and inflammation by the NF-κB signaling pathway while being infected by P. zopfii genotype II. It was concluded that adhesion of genotype II was stronger than genotype I, and therefore the genotype II regulatory ability is more robust than that of the genotype I, which causes inflammation of bovine mammary tissue.

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Acknowledgments

This research was supported by the Chinese Twelfth ‘‘Five-year’’ National Science and Technology Support Project (No. 2012BAD12B03), National Education Ministry’s Major Project (No. 313054), Specialized Research Fund for the Doctoral Program of Higher Education (SRFDP) State Education Ministry (No. 20120008110042), High-end Foreign Experts Recruitment Program (No. GDT20141100043) and the National Natural Science Foundation of China (No. 31572587).

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    Authors and Affiliations

    1. Department of Clinical Veterinary Medicine, College of Veterinary Medicine, China Agricultural University, Yuan Ming Yuan West Road No. 2, Haidian District, Beijing, 100193, People’s Republic of China

      Zhaoju Deng, Muhammad Shahid, Limei Zhang, Jian Gao, Xiaolong Gu, Shiyao Zhang, Jiaqi Zou & Bo Han

    2. UCD-Centre for Food Safety, School of Public Health, Physiotherapy and Population Science, University College Dublin, Belfield, Dublin 4, Ireland

      Séamus Fanning

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    Zhaoju Deng and Muhammad Shahid have contributed equally to this work.

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    Deng, Z., Shahid, M., Zhang, L. et al. An Investigation of the Innate Immune Response in Bovine Mammary Epithelial Cells Challenged by Prototheca zopfii . Mycopathologia 181, 823–832 (2016). https://doi.org/10.1007/s11046-016-0053-0

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