Elevated levels of blood interleukin-1β (IL-1β),
interleukin-6 (IL-6) and tumour
necrosis factor-α (TNF-α) increase
insulin resistance and result in
inflammation. It is not clear whether elevated blood level of
acetoacetate (ACAC) and decreased blood level of
glucose, which are the predominant characteristics of clinical biochemistry in ketotic dairy cows, increase proinflammatory
cytokines and subsequent
inflammation. The objective of this study was to test the hypothesis that ACAC and
glucose activate the NF-κB signalling pathway to regulate
cytokines expression in bovine hepatocytes. Bovine hepatocytes were cultured with ACAC (0-4.8 mm) and
glucose (0-5.55 mm) with or without NF-κB inhibitor
PDTC for 24 h. The secretion and
mRNA levels of
cytokines were determined by
enzyme-linked
immunosorbent assay (ELISA) and real-time fluorescence quantitative polymerase chain reaction (qRT-PCR). The NF-κB signalling pathway activation was evaluated by western blotting. Results showed that the secretion and expression of IL-1β,
IL-6 and TNF-α increased in an ACAC dose-dependent manner. Additionally, there was an increase in the secretion and
mRNA expression of these three
cytokines in
glucose treatment group, which increased significantly when the
glucose concentrations exceed 3.33 mm. Furthermore, both ACAC and
glucose upregulated NF-κB p65
protein expression and IκBα phosphorylation levels. However, these effects were reduced by
PDTC. These results demonstrate that elevated levels of ACAC and
glucose increase the synthesis and expression of proinflammatory factors by activating NF-κB signalling pathway in hepatocytes, which may contribute to
inflammation injury in ketotic dairy cows.