β-Hydroxybutyrate aggravates LPS-induced inflammatory response in bovine endometrial epithelial cells by activating the oxidative stress/NF-κB signaling pathway

Abstract

Ketosis, a metabolic disorder characterized by elevated levels of ketone bodies in the blood or urine, is known to impair the health and productivity of dairy cows, leading to substantial economic losses in the dairy industry. When ketosis occurs in dairy cows, the levels of β-hydroxybutyrate (BHBA), an abundant form of ketone bodies, in the blood increase significantly. Elevated BHBA levels have been shown to negatively impact reproductive performance and increase the incidence of periparturient diseases in dairy cows, including mastitis and endometritis. However, the role of BHBA in the development of endometritis in dairy cows and its underlying mechanisms remain largely unclear. The present study was designed to investigate the specific role of BHBA in the development of endometritis using an inflammatory response model of the bovine endometrial epithelial cell line (BENDs). Escherichia coli lipopolysaccharide (LPS) treatment (1 μg/mL) significantly increased the expression levels of interleukin (IL)-6 and IL-1β, as well as the phosphorylation of p65 and IκB in BENDs. In addition, co-treatment with BHBA (2.4 mM) and LPS (1 μg/mL) significantly increased the expression levels of proinflammatory cytokines (IL-6, IL-1β, and IL-8), as well as the phosphorylation of p65 and IκB, compared to the LPS-only treatment group. Immunofluorescence staining showed that the addition of LPS altered the nuclear localization of p65, and co-treatment with BHBA and LPS further promoted the translocation of p65 to the nucleus. Additionally, the addition of BHBA significantly increased the levels of oxidation indicators (MDA), whereas the levels of antioxidative indicators, including heme oxygenase-1 (HO-1) and catalase (CAT), were markedly decreased in BENDs. As a representative antioxidant, N-acetylcysteine (NAC) treatment significantly reduced the phosphorylation of p65 and IκB in the BHBA and LPS co-treatment group. SC75741, an NF-κB signaling pathway inhibitor, significantly decreased the expression levels of proinflammatory cytokines (IL-6, IL-1β, IL-8, and CCL5) in the BHBA and LPS co-treatment group. In summary, the current study demonstrates that BHBA aggravates LPS-induced inflammatory response in BENDs through the activation of oxidative stress/NF-κB signaling pathway, unravelling the mechanism by which BHBA exacerbates the inflammatory response in the BENDs of dairy cattle. This study elucidates the role of ketosis and its key metabolite BHBA in the pathogenesis of endometritis in dairy cows, providing valuable insights for understanding this pathological process.