Dataset Information

Deep Sequencing and Screening of Differentially Expressed MicroRNAs Related to Milk Fat Metabolism in Bovine Primary Mammary Epithelial Cells.

ABSTRACT: Milk fat is a key factor affecting milk quality and is also a major trait targeted in dairy cow breeding. To determine how the synthesis and the metabolism of lipids in bovine milk is regulated at the miRNA level, primary mammary epithelial cells (pMEC) derived from two Chinese Holstein dairy cows that produced extreme differences in milk fat percentage were cultured by the method of tissue nubbles culture. Small RNA libraries were constructed from each of the two pMEC groups, and Solexa sequencing and bioinformatics analysis were then used to determine the abundance of miRNAs and their differential expression pattern between pMECs. Target genes and functional prediction of differentially expressed miRNAs by Gene Ontology and the Kyoto Encyclopedia of Genes and Genomes analysis illustrated their roles in milk fat metabolism. Results show that a total of 292 known miRNAs and 116 novel miRNAs were detected in both pMECs. Identification of known and novel miRNA candidates demonstrated the feasibility and sensitivity of sequencing at the cellular level. Additionally, 97 miRNAs were significantly differentially expressed between the pMECs. Finally, three miRNAs including bta-miR-33a, bta-miR-152 and bta-miR-224 whose predicted target genes were annotated to the pathway of lipid metabolism were screened and verified by real-time qPCR and Western-blotting experiments. This study is the first comparative profiling of the miRNA transcriptome in pMECs that produce different milk fat content.

SUBMITTER: Shen B

PROVIDER: S-EPMC4783934 | biostudies-literature |

REPOSITORIES: biostudies-literature

ACCESS DATA

Similar Datasets

Project description:Mastitis is one of the most serious diseases that causes losses in the dairy industry, seriously impairing milk production and milk quality, and even affecting human health. Menthol is a cyclic monoterpene compound obtained from the stem and leaves of peppermint, which has a variety of biological activities, including anti-inflammatory and antioxidant activity. The purpose of this study was to investigate the preventive effect of menthol on the lipopolysaccharide-induced inflammatory response in primary bovine mammary gland epithelial cells (BMECs) and its anti-inflammatory mechanism. First, BMECs were isolated and amplified from the udders of Holstein cows by enzymatic hydrolysis. BMECs were treated with menthol (10, 50, 100, 200 μM) for 1h, followed by lipopolysaccharide (5μg/ml) for 12 h. Lipopolysaccharide treatment upregulated the protein levels of cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (INOS) and the mRNA abundance of tumor necrosis factor α (TNF-α), interleukin-6 (IL-6), and interleukin-1β (IL-1β), while menthol was able to inhibit this effect. The inhibitory effect of menthol on proinflammatory factors was significantly reduced when autophagy was blocked using 3-Methyladenine (5μg/ml), an inhibitor of autophagy. Furthermore, lipopolysaccharide treatment reduced the expression levels of milk lipids and milk proteins, which were inhibited by menthol. In addition, menthol (200 μM) treatment was able to significantly upregulate the expression level of autophagy-related protein LC3B, downregulate the expression level of P62, promote the expression abundance of autophagy-related gene mRNA, and enhance significantly enhance autophagic flux. Interestingly, treatment of BMECs with menthol (200 μM) promoted the phosphorylation of AMP-activated protein kinase (AMPK) and unc-51 like kinase 1 (ULK1) and increased the nuclear localization of nuclear factor-E2 associated factor 2 (Nrf-2). When the AMPK pathway was blocked using compound C (10μg/ml), an inhibitor of AMPK, autophagy was significantly inhibited. Autophagy levels were significantly decreased after blocking the Nrf-2 pathway using ML385 (5μg/ml), an inhibitor of Nrf-2. Overall, the data suggest that menthol activates the AMPK-ULK1 pathway to initiate the onset of autophagy and maintains the level of autophagy through the AMPK-Nrf-2 pathway. In conclusion, the findings suggest that menthol may alleviate the inflammatory response in BMECs via the AMPK/ULK1/Nrf-2/autophagy pathway.

Dataset Information

Deep Sequencing and Screening of Differentially Expressed MicroRNAs Related to Milk Fat Metabolism in Bovine Primary Mammary Epithelial Cells.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets