Although high-concentrate diet feeding can temporarily increase milk production, it can cause a series of metabolic diseases, such as subacute ruminal acidosis (SARA) and milk fat depression. The main purpose of this experiment was to study the effects of a high-concentrate diet on the inflammatory response, oxidative stress, and milk fat synthesis in the mammary gland of dairy cows. Twelve Holstein cows equipped with rumen fistulas were randomly divided into 2 groups, each with 6 cows, fed a low-concentrate diet (LC) and a high-concentrate diet (HC). On d 20 and 21 of the experiment, rumen fluid was collected to measure pH, and milk samples were collected for milk component analysis and lipopolysaccharide (LPS) concentration testing. On d 21, mammary vein blood was collected to detect the LPS concentration. At the end of the 21-d experimental period, mammary gland tissue was collected, and the expression of inflammatory response-, oxidative stress-, and milk fat synthesis-related genes and proteins in the mammary gland was analyzed by real-time quantitative PCR and western blot. The pH of rumen fluid in the HC group was significantly lower than that in the LC group, and the pH of 2 time points in the HC group was lower than 5.6, indicating that a high-concentrate diet induced SARA. The LPS concentration of the peripheral blood in HC group increased significantly compared with that in the LC group. For the inflammatory response, the proinflammatory cytokines (IL-6 and IL-1α) and innate immune factors (lingual antimicrobial peptide and tracheal antimicrobial peptide) in the mammary gland of the HC group were significantly increased, and the TLR4-NF-κB signaling pathway was activated. For oxidative stress, after HC diet feeding, the content of malondialdehyde in mammary vein blood and mammary gland tissue increased, the content of glutathione in mammary vein blood decreased, the activity of superoxide dismutase and the total antioxidant capacity in mammary gland tissue and mammary vein blood decreased, and the expression of antioxidant enzymes and antioxidant transcription factor nuclear factor, erythroid 2 like 2 (NFE2L2) in mammary gland decreased. For milk fat metabolism, HC diet feeding reduced the milk fat content in milk samples and the triacylglycerol content in the mammary gland and inhibited the expression of de novo synthase (ACACA and FASN), long-chain fatty acid converting enzymes (ACSL1 and SCD), fatty acid transporters (CD36, FATP, FABP3, and LPL), triacylglycerol synthase (AGPAT6, DGAT1, and LPIN1), lipid droplet releasing enzyme (PLIN1), and transcription factors sterol regulatory element binding protein (SREBP1) and peroxisome proliferator activated receptor gamma (PPARG). In summary, a HC diet can induce SARA with increased concentration of LPS in the peripheral vein, stimulate inflammatory reactions and oxidative stress, and inhibit milk fat synthesis in the mammary gland of dairy cows.
