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| Study on the Mechanism of ARPC3/ARPC4 Regulating Barrier Damage and Oxidative Stress in Bovine (Bos taurus) Mammary Epithelial Cells |
| YANG Ting-Ji1,2, BAO Dan1,2, DUAN Zhi-Wei1,2, BAI Wen-Kai1,2, QI Xing-Cai2,3, ZHAO Xing-Xu1,2, HE Yu-Xuan1,2, DONG Wei-Tao1,2,*, ZHANG Yong1,2,* |
1 College of Veterinary Medicine, Gansu Agricultural University, Lanzhou 730070, China;
2 Gansu Key Laboratory of Animal Generational Physiology and Reproductive Regulation, Lanzhou 730070, China;
3 College of Animal Science and Technology, Gansu Agricultural University, Lanzhou 730070, China |
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Abstract Actin-related protein 2/3 complex subunit 3 (ARPC3) and ARPC4, as important signaling molecules regulating actin cytoskeleton, plays an important role in maintaining the integrity of epithelial barrier function during the development of inflammation. In order to explore the mechanism of ARPC3/4 regulating barrier injury and oxidative stress in bovine (Bos taurus) mammary alveolar cells-large T antigen (MAC-T), cell counting kit-8 (CCK-8) was used to screen the optimal concentration of lipopolysaccharide (LPS) and ARPC3/ARPC4 inhibitor (CK666) in this study. qPCR and Western blot were used to detect the expression of nuclear factor-κB (NF-κB) signaling pathway, antioxidant and tight junction related proteins and genes. The results showed that LPS significantly increased the content of reactive oxygen species (ROS) and up-regulated the expression of myeloid differentiation factor 88 (MyD88), Toll-like receptor 4 (TLR4), nuclear factor-κB inhibitor kinase β (IKKβ) and NF-κB. The expression of catalase (CAT), superoxide dismutase 1 (SOD1), glutathione peroxidase 1 (GPx1), Claudin1, Occludin and zonula occludens-1 (ZO-1) were down-regulated. After inhibiting ARPC3/ARPC4 by CK666 treatment, the expression of heat shock protein 70 (HSP70) was further up-regulated compared with LPS group, and the up-regulation of NF-κB, MyD88, TLR4 and IKKβ induced by LPS was reversed. The expression of CAT, SOD1, GPx1, Claudin1, ZO-1 and Occludin was down-regulated, and the content of ROS was reduced. The results showed that in the inflammatory model of bovine mammary epithelial cells, ARPC3/ARPC4 impaired the epithelial cell barrier by destroying the tight junction, thereby activating the NF-κB signaling pathway and inhibiting the expression of antioxidant genes to promote inflammation and oxidative stress, indicating its potential as a therapeutic target for mastitis. This study provides new ideas and theoretical basis for the prevention and treatment of dairy cow mastitis.
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Received: 30 December 2024
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Corresponding Authors:
*dongwt@gsau.edu.cn; zhangyong@gsau.edu.cn
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