MicroRNA (miRNA) are a class of small noncoding RNA that function as important posttranscriptional regulators of gene expression. The acyl-CoA synthetase long-chain family member 1 (ACSL1) is an important enzyme in the process of milk lipid synthesis. In a previous study dealing with incubations of stearic acid in bovine mammary epithelial cells, an opposite expression pattern was observed between ACSL1 and miR-181a. Bioinformatics analysis with TargetScan and PicTar revealed ACSL1 as a potential target gene of miR-181a. The objective of this work was to determine the potential function of miR-181a on milk fat synthesis by defining the regulatory relationship between miR-181a and ACSL1. Primary bovine mammary epithelial cells were harvested from mid-lactation cows and cultured in Dulbecco's modified Eagle's medium/F-12 medium with 10% fetal bovine serum, 0.5μg/mL of insulin, 10 ng/mL of epidermal growth factor, 5μg/mL of transferrin, 1μg/mL of hydrocortisone, 1μg/mL of progesterone, 5μg/mL of estradiol, and 5μg/mL of prolactin. Cells were transfected with an miR-181a mimic to increase its expression and an miR-181a inhibitor to decrease its expression before culturing for 48 h. The results revealed that the overexpression of miR-181a inhibited the expression of ACSL1, whereas the downregulation of miR-181a increased ACSL1 expression. Western blot analysis of ACSL1 revealed similar effects. Oil-red-O staining indicated that cellular lipid droplet synthesis was decreased with the overexpression of bta-miR-181a, and treatment with the bta-miR-181a inhibitor increased concentration of lipid droplets. Furthermore, overexpression of bta-miR-181a resulted in a decrease in concentration of triacylglycerol in the cells, whereas inhibition of bta-miR-181a increased concentration of triacylglycerol. Therefore, the results indicated that bta-miR-181a may contribute to negative regulation of lipid synthesis in mammary cells via targeting ACSL1.
