Browsing by Author "Yang, Jingyue"
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- ItemOpen AccessKlebsiella pneumoniae infection causes mitochondrial damage and dysfunction in bovine mammary epithelial cells(2021-02-10) Cheng, Jia; Zhang, Jv; Yang, Jingyue; Yi, Bing; Liu, Gang; Zhou, Man; Kastelic, John P; Han, Bo; Gao, JianAbstract Klebsiella pneumoniae, an important cause of bovine mastitis worldwide, is strongly pathogenic to bovine mammary epithelial cells (bMECs). Our objective was to determine the role of mitochondrial damage in the pathogenicity of K. pneumoniae on bMECs, by assessing several classical indicators of mitochondrial dysfunction, as well as differentially expressed genes (DEGs). Two K. pneumoniae strains (HLJ-D2 and HB-AF5), isolated from cows with clinical mastitis (CM), were used to infect bMECs (MAC-T line) cultured in vitro. In whole-transcriptome analysis of bMECs at 6 h post-infection (hpi), there were 3453 up-regulated and 3470 down-regulated genes for HLJ-D2, whereas for HB-AF5, there were 2891 up-regulated and 3278 down-regulated genes (P < 0.05). Based on GO term enrichment of differentially expressed genes (DEGs), relative to the controls, the primary categories altered in K. pneumoniae-infected bMECs included cellular macromolecule metabolism, metabolic process, binding, molecular function, etc. Infections increased (P < 0.05) malondialdehyde concentrations and formation of reactive oxygen species in bMECs. Additionally, both bacterial strains decreased (P < 0.05) total antioxidant capacity in bMECs at 6 and 12 hpi. Furthermore, infections decreased (P < 0.05) mitochondrial membrane potential and increased (P < 0.01) mitochondrial calcium concentrations. Finally, severe mitochondrial swelling and vacuolation, as well as mitochondrial rupture and cristae degeneration, were detected in infected bMECs. In conclusion, K. pneumoniae infections induced profound mitochondrial damage and dysfunction in bMECs; we inferred that this caused cellular damage and contributes to the pathogenesis of K. pneumoniae-induced CM in dairy cows.
- ItemOpen AccessMicroRNA miR-223 modulates NLRP3 and Keap1, mitigating lipopolysaccharide-induced inflammation and oxidative stress in bovine mammary epithelial cells and murine mammary glands(2023-09-14) Zhou, Man; Barkema, Herman W.; Gao, Jian; Yang, Jingyue; Wang, Yue; Kastelic, John P.; Khan, Sohrab; Liu, Gang; Han, BoAbstract Bovine mastitis, the most prevalent and costly disease in dairy cows worldwide, decreases milk quality and quantity, and increases cow culling. However, involvement of microRNAs (miRNAs) in mastitis is not well characterized. The objective was to determine the role of microRNA-223 (miR-223) in regulation of the nucleotide-binding oligomerization domain-like receptor containing pyrin domain 3 (NLRP3) inflammasome and kelch like ECH-associated protein 1 (Keap1)/nuclear factor erythroid 2-related factor 2 (Nrf2) oxidative stress pathway in mastitis models induced by lipopolysaccharide (LPS) treatment of immortalized bovine mammary epithelial cells (bMECs) and murine mammary glands. In bMECs cultured in vitro, LPS-induced inflammation downregulated bta-miR-223; the latter interacted directly with the 3’ untranslated region (3’ UTR) of NLRP3 and Keap1. Overexpression of bta-miR-223 in bMECs decreased LPS and Adenosine 5’-triphosphate (ATP)-induced NLRP3 and its mediation of caspase 1 and IL-1β, and inhibited LPS-induced Keap1 and Nrf2 mediated oxidative stress, whereas inhibition of bta-miR-223 had opposite effects. In an in vivo murine model of LPS-induced mastitis, increased miR-223 mitigated pathology in the murine mammary gland, whereas decreased miR-223 increased inflammatory changes and oxidative stress. In conclusion, bta-miR-223 mitigated inflammation and oxidative injury by downregulating the NLRP3 inflammasome and Keap1/Nrf2 signaling pathway. This study implicated bta-miR-223 in regulation of inflammatory responses, with potential as a novel target for treating bovine mastitis and other diseases.
- ItemOpen AccessStreptococcus lutetiensis Induces Autophagy via Oxidative Stress in Bovine Mammary Epithelial Cells(2022-02-07) Chen, Peng; Yang, Jingyue; Wu, Naiwen; Han, Bo; Kastelic, John P.; Gao, JianStreptococcus lutetiensis, an emerging pathogen causing bovine mastitis, has not been well characterized. We reported that S. lutetiensis was pathogenic both in vivo and in vitro and caused inflammatory reactions in the mammary gland. However, roles of autophagy and oxidative stress in the pathogenesis of S. lutetiensis-induced mastitis are unclear. In this study, an autophagy model of S. lutetiensis-infected bovine mammary epithelial cells (bMECs) was used to assess oxidative stress and autophagy flux. Expressions of Beclin1, light chain 3II, and Sequestosome 1/p62 were elevated in bMECs after S. lutetiensis infection. In addition, autophagosome and lysosome formation confirmed autophagy occurred. Based on LysoTracker Red and acridine orange, lysosome degradation was blocked, and lower expressions of lysosomal-associated membrane protein 2, cathepsins D, and cathepsins L confirmed lysosomal damage. Concurrently, the nuclear factor erythroid 2-related factor 2 (Nrf2), kelch-like ECH-associated protein 1 (Keap1), heme oxygenase 1 (HO1), and NAD (P)H: quinone oxidoreductase 1 (NQO1), and basilic proteins associated with the Nrf2/Keap1 signaling pathway, were detected. Decreased keap1 and increased Nrf2, HO1, NQO1, and reactive oxygen species (ROS) indicated increased oxidative stress. Treatment with N-Acetyl-L-cysteine (NAC), an ROS inhibitor, decreased both oxidative stress and autophagy. Therefore, we concluded that S. lutetiensis caused intracellular oxidative stress and autophagy in bMECs. In addition, crosstalk between autophagy and oxidative stress affected the autophagic flux and blocked downstream autophagy. The Nrf2-keap1-p62 pathway participated in this process, with ROS acting upstream of these effects, interfering with normal cell functions.