PGN and LTA from Staphylococcus aureus Induced Inflammation and Decreased Lactation through Regulating DNA Methylation and Histone H3 Acetylation in Bovine Mammary Epithelial Cells

Abstract

Staphylococcus aureus (S. aureus) and Escherichia coli (E. coli) are the most common pathogens of mastitis, and S. aureus generally causes subclinical mastitis which is more persistent and resistant to treatment. Peptidoglycan (PGN) and lipoteichoic acid (LTA) are cell wall components of S. aureus. Although the roles of PGN and LTA in causing inflammation are well studied, the epigenetic mechanisms of the effects of PGN and LTA on the inflammation and lactation remain poorly understood. This study characterized the gene expression profiling by RNA sequencing and investigated DNA methylation and histone acetylation in relation to inflammation and lactation in the immortalized bovine mammary epithelial cell line (MAC-T). The cells were cultured for 24 h with neither PGN nor LTA (CON), PGN (30 µg/mL), LTA (30 µg/mL), and PGN (30 µg/mL) + LTA (30 µg/mL), respectively. The number of differentially expressed genes (DEGs) and the expression of proinflammatory factors including interleukin (IL)-1β, IL-6, IL-8, chemokine (C-X-C motif) ligand (CXCL)1, and CXCL6 of the treatments increased in the following order: CON < PGN < LTA < PGN + LTA, and the DEGs mainly enriched on the cytokine-cytokine receptor interaction and chemokine signaling pathway. LTA and PGN + LTA induced hypomethylation of global DNA by suppressing DNA methyltransferase (DNMT) activity. PGN and LTA, alone or combined, decreased the mRNA expression of casein genes (CSN1S1, CSN2, and CSN3) and the expression of two caseins (CSN2 and CSN3), and reduced histone H3 acetylation by suppressing histone acetyltransferase (HAT) activity and promoting histone deacetylase (HDAC) activity. Collectively, this study revealed that PGN and LTA induced inflammation probably due to decreasing DNA methylation through regulating DNMT activity, and decreased lactation possibly through reducing histone H3 acetylation by regulating HAT and HDAC activity in bovine mammary epithelial cells. Key Contribution:In the present study, we demonstrated for the first time that PGN and LTA promoted the expression of proinflammatory factors probably due to inducing DNA hypomethylation by suppressing DNMT activity, whereas PGN and LTA decreased the gene expression of caseins perhaps through reducing histone H3 acetylation by suppressing HAT or/and promoting HDAC activity.

Genome-Wide DNA Methylation Analysis of Mammary Gland Tissues From Chinese Holstein Cows With Staphylococcus aureus Induced Mastitis

“…Additionally, abnormal expression of IL6 was revealed to be due to altered DNA methylation rather than genetic mutation during clinical mastitis (Zhang et al, 2018). Moreover, methylation changes in NCKAP5 and transposon MTD were associated with the development of S. aureus mastitis (Wang et al, 2020), while hypomethylation of global DNA methylation induced by S. aureus infection repressed DNA methyltransferase activity in bovine mammary epithelial cells (Wu et al, 2020).…”

The Synergism of PGN, LTA and LPS in Inducing Transcriptome Changes, Inflammatory Responses and a Decrease in Lactation as Well as the Associated Epigenetic Mechanisms in Bovine Mammary Epithelial Cells

“…Other studies have also shown that LTA and PGN could cause the DNA hypomethylation of the key regulators of inflammatory pathways, promoting the release of a variety of inflammatory factors [37,38]. In addition, our previous studies showed that LPS, LTA, and PGN suppressed the expression of lactation-related genes of BMECs due to reducing histone H3 acetylation through regulating HAT and HDAC activity [19,39]. Thus, we speculated that co-stimulation with LPS, LTA, and PGN might have an additive effect on DNA hypomethylation and histone hypoacetylation, producing a more intense inflammatory response and decreasing casein expression to a greater degree than single stimulation of either of the PAMPs in BMECs.…”

“…LPS could cause DNA hypomethylation at many inflammatory loci by suppressing DNMT expression, and then increase the inflammatory factor expression of human dental pulp cells [32] and macrophages [33], rat brain tissue [34], bovine fibroblasts [35], and so on. Our previous study showed that LPS, LTA, and PGN enhanced the inflammatory responses of BMECs by decreasing DNA methylation levels [19,36]. Other studies have also shown that LTA and PGN could cause the DNA hypomethylation of the key regulators of inflammatory pathways, promoting the release of a variety of inflammatory factors [37,38].…”

“…Medium supernatant was collected to detect inflammatory factor concentrations by ELISA. All the materials and methods used in the Western blot and ELISA were the same as those of our previous study, in which a detailed description is available [19].…”

“…Lipopolysaccharide (LPS) is the pathogen-associated molecular pattern (PAMP) of Gram-negative bacteria, and lipoteichoic acid (LTA) and peptidoglycan (PGN) are those of Gram-positive bacteria [ 2 ]. Previous studies have reported the effects of LPS, PGN, LTA, and PGN + LTA on the gene expression profiles of bovine mammary epithelial cells (BMECs) and mainly focused on their proinflammatory activity [ 12 , 13 , 14 , 15 , 16 , 17 , 18 , 19 ]. However, the additive effects of LPS, PGN, and LTA on the gene expression profiles of BMECs are still unclear.…”