ABSTRACT
Subacute ruminal acidosis (SARA) is a common nutritional metabolic disease in ruminants that causes significant economic losses to dairy farming. Strontium (Sr) is known to be involved in bone metabolism and exhibits potent anti-inflammatory effects. To evaluate the effect of Sr on inflammation in bovine ruminal epithelial cells, a model of LPS-induced inflammation was established in this study, and the cell viability of bovine ruminal epithelial cells was measured using CCK-8. The production of pro-inflammatory cytokines was measured by ELISA and real-time PCR, respectively. The related proteins of the TLR4/MyD88/NF-κB pathway were assayed through Western blotting, and the fluorescence of p-p65 and p-IκB were assayed by immunofluorescence. Molecular docking of Sr and TLR4/MyD88/NF-κB pathway-related proteins was performed using MIB2 ( http://bioinfo.cmu.edu.tw/MIB2/ ). Results showed that after treatment for 24 h, the cell viability was decreased at the high concentration of Sr (≥ 10 mmol/L). Sr significantly decreased the production of TNF-α, IL-1ß, and IL-6, downregulated the related proteins expression of the TLR4/MyD88/NF-κB pathway, and reduced the fluorescence levels of p-p65 and p-IκB. The NF-κB pathway inhibitor PDTC and molecular docking further revealed that Sr reduced LPS-induced pro-inflammatory cytokines production via the TLR4/MyD88/NF-κB pathway. These results suggest that Sr reduces LPS-induced pro-inflammatory cytokines production via the TLR4/MyD88/NF-κB pathway, thereby exerting an anti-inflammatory effect in bovine ruminal epithelial cells, providing a basis for Sr in the treatment of bovine rumen acidosis disease.
ABSTRACT
Subacute ruminal acidosis (SARA) is a prevalent disease in intensive dairy farming, and the rumen environment of diseased cows acidifies, leading to the rupture of gram-negative bacteria to release lipopolysaccharide (LPS). LPS can cause rumentitis and other complications, such as liver abscess, mastitis and laminitis. Propionate, commonly used in the dairy industry as a feed additive, has anti-inflammatory effects, but its mechanism is unclear. This study aims to investigate whether sodium propionate (SP) reduces LPS-induced inflammation in rumen epithelial cells (RECs) and the underlying mechanism. RECs were stimulated with different time (0, 1, 3, 6, 9, 18 h) and different concentrations of LPS (0, 1, 5, 10 µg/mL) to establish an inflammation model. Then, RECs were treated with SP (15, 25, 35 mM) or 10 µM PDTC in advance and stimulated by LPS for the assessment. The results showed that LPS (6h and 10 µg/mL) could stimulate the phosphorylation of NF-κB p65, IκB, JNK, ERK and p38 MAPK through TLR4, and increase the release of TNF-α, IL-1ß and IL-6. SP (35 mM) can reduce the expression of cytokines by effectively inhibiting the NF-κB and MAPK inflammatory pathways. This study confirmed that SP inhibited LPS-induced inflammatory responses through NF-κB and MAPK in RECs, providing potential therapeutic targets and drugs for the prevention and treatment of SARA.
