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Milk fat determines the quality of milk and is also a main targeted trait in dairy cow breeding. Recent studies have revealed important regulatory roles of microRNAs (miRNA) in milk fat synthesis in the mammary gland. However, the role of miRNA in bovine mammary epithelial cells (BMEC) remains largely unknown. In this study, we found that the overexpression of miR-130a significantly decreased cellular triacylglycerol (TAG) levels and suppressed lipid droplet formation, whereas the inhibition of miR-130a resulted in greater lipid droplet formation and TAG accumulation in BMEC. MiR-130a also significantly affected mRNA expression related to milk fat metabolism. Specifically, the overexpression of miR-130a reduced the mRNA expression of PPARG, FABP3, PLIN2, and FATP1, whereas the downregulation of miR-130a increased the mRNA expression of PPARG, C/EBPβ, C/EBPα, PLIN2, FABP3, and FATP1. Furthermore, western blot analysis revealed the protein level of PPARG in miR-130a mimic and inhibitor transfection groups to be consistent with the mRNA expression response. Finally, luciferase reporter assays verified that PPARG was the direct target of miR-130a. This study provides the first experimental evidence that miR-130a directly affects TAG synthesis in BMEC by targeting PPARG, suggesting that miR-130a potentially could be used to improve beneficial milk components in dairy cows. [ABSTRACT FROM AUTHOR]

Growing evidence has revealed that microRNA are central elements in milk fat synthesis in mammary epithelial cells. A negative regulator of adipocyte fat synthesis, miR-27a has been reported to be involved in the regulation of milk fat synthesis in goat mammary epithelial cells; however, the regulatory role of miR-27a in bovine milk fat synthesis remains unclear. In the present study, primary bovine mammary epithelial cells (BMEC) were harvested from mid-lactation cows and cultured in Dulbecco's modified Eagle's medium/F-12 medium with 10% fetal bovine serum, 5 µg/mL of insulin, 1 µg/mL of hydrocortisone, 2 µg/mL of prolactin, 1 µg/mL of progesterone, 100 U/mL of penicillin, and 100 µg/mL of streptomycin. We found that the overexpression of miR-27a significantly suppressed lipid droplet formation and decreased the cellular triacylglycerol (TAG) levels, whereas inhibition of miR-27a resulted in a greater lipid droplet formation and TAG accumulation in BMEC. Meanwhile, overexpression of miR-27a inhibited mRNA expression of peroxisome proliferatoractivated receptor gamma (PPARG), CCAAT/enhancer- binding protein beta (C/EBPβ), perilipin 2 (PLIN2), and fatty acid binding protein 3 (FABP3), whereas miR-27a downregulation increased PPARG, C/EBPβ, FABP3, and CCAAT enhancer binding protein alpha (C/EBPa) mRNA expression. Furthermore, Western blot analysis revealed the protein level of PPARG in miR-27a mimic and inhibitor transfection groups to be consistent with the mRNA expression response. Moreover, luciferase reporter assays verified that PPARG was the direct target of miR-27a. In summary, these results indicate that miR-27a has the ability to control TAG synthesis in BMEC via targeting PPARG, suggesting that miR-27a could potentially be used to improve beneficial milk components in dairy cows. [ABSTRACT FROM AUTHOR]