Effect Of LPS On The Milk Fat Synthesis And Lipid Metabolism-related Molecules In Dairy Cow Mammary Epithelial Cells

Lipopolysaccharides?LPS?from the cell wall of Gram-negative bacteria are also referred to as a bacterial endotoxin.Dairy cows with rumen acidosis,breast and uterine infection,or heat stress can produce LPS.LPS is harmful to dairy cows,causing systemic and local inflammatory reactions,and leading to a serious decrease in milk fat percentage and yield,and consequent huge economic losses in the dairy farming and processing industries.It is known that LPS can reduce the content of milk fat synthetic precursors in the blood.LPS may affect milk fat synthesis in dairy cow breast tissue in a direct way,but the underlying mechanism is not entirely clear.Therefore,in this study,primary cultured dairy cow mammary epithelial cells?DCMECs?were used as a model to investigate the effect of LPS on the synthesis of milk fat and the related molecules of lipid metabolism in DCMECs.In order to clarify the effect of LPS on lipid metabolism Mechanism of action to provide the basis for the study.We used collagenase I and hyaluronidase digestion to culture DCMECs.Pure DCMECs were obtained by differential trypsinization.The immunofluorescence assay,morphological observation,cell growth curve mapping and milk fat and milk protein synthesis function analysis to determine the culture of DCMECs consistent with the biological laws of cell growth,with the normal synthesis and secretion of milk fat and milk protein function.The DCMECs of15^thpassage was recovered,and DCMECs growth status is still good without morphological changes.The DCMECs can be used for subsequent experiments.DCMECs were treated with different concentrations of LPS for 24 hours.Cytotoxicity detection,observation of cell growth status and expression of inflammatory cytokines mRNA showed that the optimal working concentration of LPS was 10?g/mL.DCMECs were treated with LPS?10?g/m L?for 0,1,3,6,12,24 and 48 hours.After these treatments,we examined the levels of sterol regulatory element binding protein 1?SREBP1?,Liver X receptor??LXR??and peroxisome proliferators-activated receptors??PPAR??protein expression,nuclear translocation and m RNA relative expression and its regulation of fatty acid metabolism related enzymes:fatty acid synthetase?FAS?,acetyl-CoA binding protein?ACBP?,lipoprotein lipase?LPL?,cluster differentiation 36?CD36?and Acetyl-CoA carboxylase 1?ACC1?m RNA relative expression levels and lipid droplet formation function.It was found that LPS?10?g/mL?reducedthetranscription,translationandnucleartranslocationof lipid-metabolism-related transcription factor:SREBP1,PPAR?and LXR?,and the mRNAs of fatty acid metabolism related enzymes:ACC1,FAS,ACBP and LPL in DCMECs.LPS increased m RNA relative expression of the fatty acid transporter CD36,but overall decreases milk fat production,and in a time-dependent manner.In conclusion,LPS can regulate the transcription,translation and nuclear translocation of lipid metabolism-related molecules in DCMECs,thereby affecting the de novo synthesis of fatty acids and the uptake and transport of fatty acids in DCMECs,ultimately affecting the synthesis of milk fat,and in a time-dependent manner.