Innate immune responses induced by lipopolysaccharide and lipoteichoic acid in primary goat mammary epithelial cells

Abstract

BackgroundInnate immune responses induced by in vitro stimulation of primary mammary epithelial cells (MEC) using Gram-negative lipopolysaccharide (LPS) and Gram-positive lipoteichoic acid (LTA) bacterial cell wall components are well- characterized in bovine species. The objective of the current study was to characterize the downstream regulation of the inflammatory response induced by Toll-like receptors in primary goat MEC (pgMEC). We performed quantitative real-time RT-PCR (qPCR) to measure mRNA levels of 9 genes involved in transcriptional regulation or antibacterial activity: Toll-like receptor 2 (TLR2), Toll-like receptor 4 (TLR4), prostaglandin-endoperoxide synthase 2 (PTGS2), interferon induced protein with tetratricopeptide repeats 3 (IFIT3), interferon regulatory factor 3 (IRF3), myeloid differentiation primary response 88 (MYD88), nuclear factor of kappa light polypeptide gene enhancer in B-cells 1 (NFKB1), Toll interacting protein (TOLLIP), and lactoferrin (LTF). Furthermore, we analyzed 7 cytokines involved in Toll-like receptor signaling pathways: C-C motif chemokine ligand 2 (CCL2), C-C motif chemokine ligand 5 (CCL5), C-X-C motif chemokine ligand 6 (CXCL6), interleukin 8 (CXCL8), interleukin 1 beta (IL1B), interleukin 6 (IL6), and tumor necrosis factor alpha (TNF).ResultsStimulation of pgMEC with LPS for 3 h led to an increase in expression of CCL2, CXCL6, IL6, CXCL8, PTGS2, IFIT3, MYD88, NFKB1, and TLR4 (P < 0.05). Except for IL6, and PTGS2, the same genes had greater expression than controls at 6 h post-LPS (P < 0.05). Expression of CCL5, PTGS2, IFIT3, NFKB1, TLR4, and TOLLIP was greater than controls after 3 h of incubation with LTA (P < 0.05). Compared to controls, stimulation with LTA for 6 h led to greater expression of PTGS2, IFIT3, NFKB1, and TOLLIP (P < 0.05) whereas the expression of CXCL6, CXCL8, and TLR4 was lower (P < 0.05). At 3 h incubation with both toxins compared to controls a greater expression (P < 0.05) of CCL2, CCL5, CXCL6, CXCL8, IL6, PTGS2, IFIT3, IRF3, MYD88, and NFKB1 was detected. After 6 h of incubation with both toxins, the expression of CCL2, CXCL6, IFIT3, MYD88, NFKB1, and TLR4 was higher than the controls (P < 0.05).ConclusionsData indicate that in the goat MEC, LTA induces a weaker inflammatory response than LPS. This may be related to the observation that gram-positive bacteria cause chronic mastitis more often than gram-negative infections.Electronic supplementary materialThe online version of this article (doi:10.1186/s40104-017-0162-8) contains supplementary material, which is available to authorized users.

VA inhibits LPS-induced oxidative stress via modulating Nrf2/NF-κB-signalling pathways in bovine mammary epithelial cells

“…LPS could result in the activation of NF-jB, regulating the expression of inflammatory cytokines in mice embryonic fibroblast (Yamamoto et al 2003). LPS is one of the main stress sources used in the study of oxidative damage in BMECs and goat mammary epithelial cells to induce IL-1 production (Bulgari et al 2017;Wang et al 2017). Our previous research has established LPS-induced oxidative stress model of BMECs .…”

The Mammary Microenvironment in Mastitis in Humans, Dairy Ruminants, Rabbits and Rodents: A One Health Focus

“…We have previously demonstrated that, during murine post-lactational regression, mammary epithelial cells exhibit dramatic, Stat3-dependent, up-regulation of CD14, an innate immune component [ 52 , 53 ]. In addition, cultured bovine and caprine mammary epithelial cells have been shown to express TLR2 and TLR4 mRNA, and responses of mammary epithelial cells to stimulation with bacterial cell wall components, or other agents with the potential to cause mastitis, such as Prototheca spp., result in elaboration of a plethora of cytokines and other inflammatory mediators [ 54 , 55 ]. These findings suggest that mammary epithelial cells, across species, have a critical role to play in the mammary immune microenvironment although clearly much of the current evidence for this assertion is derived from studies using rodent mammary epithelial cells in vitro, and from in vivo studies of rodent models.…”

C-C motif chemokine ligand 2 induces proliferation and prevents lipopolysaccharide-induced inflammatory responses in bovine mammary epithelial cells

“…The CCL2 generally induces inflammatory responses through monocyte infiltration, but it also acts as an anti-inflammatory agent at low concentrations (Flaishon et al, 2008). The expression of CCL2 in mammary epithelial cells of goats increases in response to LPS, but it is unknown whether CCL2 can control mastitis in cows (Bulgari et al, 2017).…”