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This study aimed to verify the anti-inflammatory effect of soybean isoflavones (SI) on the inflammatory response induced by Streptococcus agalactiae (S. agalactiae) of bovine mammary epithelial cells (bMECs) and to elucidate its possible mechanism. BMECs were pretreated with SI of different concentrations (20, 40, 60, 80, 100 μg/mL) for 0.5, 3, 6, 9, 12, 15, 18, 24 h. And then, S. agalactiae was used to infect bMECs for 6 h (MOI = 50:1) to establish the inflammation model. Cell viability, growth curves of S. agalactiae, cytotoxicity, and S. agalactiae invasion rate were determined. A proteomics technique was used to further detect differential proteins and enrichment pathways. SI (40 μg/mL) improved the viability of bMECs at 12 h (p < 0.05) and 60 and 80 μg/mL of SI greater (p < 0.01). Moreover, 60 μg/mL of SI protects cells from bacterial damage (p < 0.05). SI could inhibit S. agalactiae growth and internalization into bMECs in a time-and dose-dependent manner. In addition, proteomics results showed that 133 proteins were up-regulated and 89 proteins were down-regulated significantly. The differentially significantly expressed proteins (DSEPs) were mainly related to cell proliferation, differentiation, apoptosis, and migration. GO annotation showed that 222 DSEPs were divided into 23 biological processes (BP) terms, 14 cell components (CC) terms, and 12 molecular functions (MF) terms. DSEPs were significantly enriched in 10 pathways, of which the immune pathway was the main enrichment pathway.

BackgroundThe metabolic processes of cows undergo significant changes during subclinical mastitis, but their molecular mechanisms have not been clearly elucidated. This study investigated the changes in milk metabolites after intramammary infusion of matrine, a plant alkaloid with anticancer properties, in the form of a chitosan hydrogel into bovine mammary glands with subclinical mastitis. Infusions were continued for seven days, and milk samples were collected on day 1 and day 7 for analysis of the microbiome by 16S rRNA gene sequencing and metabolites by liquid chromatography-mass spectrometry.ResultsMatrine-chitosan hydrogels (MCHs) significantly decreased the somatic cell count on day 7 and the Simpson index indicated that microbial diversity was significantly lower on day 7 than on day 1. On day 7, the numbers of Aerococcus, Corynebacterium_1 and Staphylococcus were significantly lower, while the abundance of Firmicutes was very significantly decreased. The numbers of Probacteria increased, however. In milk samples, we identified 74 differentially expressed metabolites and the MCH infusion group had the most significantly upregulated metabolites including sphingolipids, glycerophospholipids, flavonoids and fatty acyls. Principal component analysis and the orthogonal partial least squares discriminant test confirmed good separation of the milk metabolites. The identification of active milk metabolic pathways after MCH treatment supported the known antimicrobial and anti-inflammatory properties of matrine that are associated with glycerophospholipid metabolism and the sphingolipid metabolic signaling pathways.ConciusionsThese insights into the mechanisms and the corresponding biological responses to matrine demonstrate its potential immunoregulatory activity and emphasize the need for continued investigation.