scholarly journals Beta-Sitosterol Promotes Milk Protein and Fat Syntheses-Related Genes in Bovine Mammary Epithelial Cells

Animals ◽  
2021 ◽  
Vol 11 (11) ◽  
pp. 3238
Author(s):  
Xinlu Liu ◽  
Jinglin Shen ◽  
Jinxin Zong ◽  
Jiayi Liu ◽  
Yongcheng Jin
Keyword(s):  
Epithelial Cells ◽  
Protein Expression ◽  
Dairy Cows ◽  
Milk Fat ◽  
Milk Protein ◽  
Mammary Epithelial Cells ◽  
Mammary Epithelial ◽  
Bovine Mammary Epithelial Cells ◽  
Mrna And Protein Expression ◽  
Fat Synthesis

β-sitosterol, a phytosterol with multiple biological activities, has been used in the pharmaceutical industry. However, there are only a few reports on the use of β-sitosterol in improving milk synthesis in dairy cows. This study aimed to investigate the effects of β-sitosterol on milk fat and protein syntheses in bovine mammary epithelial cells (MAC-T) and its regulatory mechanism. MAC-T cells were treated with different concentrations (0.01, 0.1, 1, 5, 10, 20, 30, or 40 μM) of β-sitosterol, and the expression levels of milk protein and fat synthesis-related genes and proteins were analyzed. β-sitosterol at 0.1, 1, and 10 μM concentrations promoted the mRNA and protein expression of β-casein. β-sitosterol (0.1, 1, 10 μM) increased the mRNA and protein expression levels of signal transducer activator of transcription 5 (STAT5), mammalian target of rapamycin (mTOR), and ribosomal protein S6 kinase beta-1 (S6K1) of the JAK2/STAT5 and mTOR signaling pathways. It also stimulated the milk fat synthesis-related factors, including sterol regulatory element-binding protein 1 (SREBP1), peroxisome proliferator-activated receptor-gamma (PPARγ), acetyl-CoA carboxylase (ACC), lipoprotein lipase (LPL), and stearyl CoA desaturase (SCD). β-sitosterol (0.1, 1, 10 μM) also significantly increased the expression of growth hormone/insulin-like growth factor-1 (GH/IGF-1) axis and hypoxia-inducible factor-1α (HIF-1α)-related genes. Notably, the compound inhibited the expression of the negative regulator, the suppressor of cytokine signaling 2 (SOCS2) at the two lower concentrations (0.1, 1 μM), but significantly promoted the expression at the highest concentration (30 μM). These results highlight the role of β-sitosterol at concentrations ranging from 0.1 to 10 μM in improving milk protein and fat syntheses, regulating milk quality. Therefore, β-sitosterol can be used as a potential feed additive to improve milk quality in dairy cows.

Effects of Insulin on Milk Fat Synthesis by Regulating the Expressions of SREBP1 and mTOR in Bovine Mammary Epithelial Cells

2020 ◽  
Author(s):  
Nan Li ◽  
Peng-Xia Zhang ◽  
Xin Huang ◽  
Hai-Tao Yao ◽  
Dong-Pu Liu ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Milk Fat ◽  
Mammary Epithelial Cells ◽  
Mammary Epithelial ◽  
Blank Group ◽  
Bovine Mammary Epithelial Cells ◽  
Real Time Quantitative Pcr ◽  
Notable Increase ◽  
Fat Synthesis ◽  
Milk Fat Synthesis

Due to the complexity of insulin in life activities, the role of insulin in mammalian lactation has not been well explained. To investigate the influence of insulin on milk fat synthesis, bovine mammary epithelial cells (BMECs) were cultured in treatment with insulin. We determined the content of Triglyceride (TG) in cell-free culture medium and found a notable increase in TG secrection. Lipid droplet staining study showed a consistent result. We also used real-time quantitative PCR and western blotting to detect the expression of signaling molecules related to milk fat synthesis. We found that insulin resulted in an obvious increase of SREBP-1, mTOR and lipogenic gene expression compared with the blank group. Taken together, our study reveals that insulin plays a significant role in milk fat synthesis.

scholarly journals MiR-485 targets the DTX4 gene to regulate milk fat synthesis in bovine mammary epithelial cells

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Juan Liu ◽  
Ping Jiang ◽  
Ambreen Iqbal ◽  
Shaokat Ali ◽  
Zhen Gao ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Milk Fat ◽  
Target Gene ◽  
Mammary Epithelial Cells ◽  
Cholesterol Content ◽  
Mammary Epithelial ◽  
Bovine Mammary Epithelial Cells ◽  
Candidate Target Gene ◽  
Fat Synthesis ◽  
Milk Fat Synthesis

AbstractMicroRNAs (miRNAs) are mRNA suppressors that regulate a variety of cellular and physiological processes, including cell proliferation, apoptosis, triglyceride synthesis, fat formation, and lipolysis, by post-transcriptional processing. In previous studies, we isolated and sequenced miRNAs from mammary epithelial cells from Chinese Holstein cows with high and low milk fat percentages. MiR-485 was one of the significantly differentially expressed miRNAs that were identified. In the present study, the relationship between the candidate target gene DTX4 and miR-485 was validated by bioinformatics and real-time fluorescent quantitative PCR (qRT-PCR) and Western blot (WB) analyses in bovine mammary epithelial cells (bMECs). The results indicated that miR-485 negatively regulated the mRNA expression of the target gene DTX4. Furthermore, an shRNA interference vector for the target gene DTX4 was constructed successfully, and it increased the triglyceride content and reduced the cholesterol content of transfected cells. These results suggest that miR-485 may affect the contents of triglycerides (TGs) and cholesterol (CHOL) by targeting the DTX4 gene. This study indicates that miR-485 has a role in regulating milk fat synthesis and that miR-485 targets the DTX4 gene to regulate lipid metabolism in bMECs. These findings contribute to the understanding of the functional significance of miR-485 in milk fat synthesis.

scholarly journals Bta-miR-34b controls milk fat biosynthesis via the Akt/mTOR signaling pathway by targeting RAI14 in bovine mammary epithelial cells

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Yujuan Wang ◽  
Xiaoyu Wang ◽  
Meng Wang ◽  
Li Zhang ◽  
Linsen Zan ◽  
...  
Keyword(s):  
Dairy Cows ◽  
Signaling Pathway ◽  
Milk Fat ◽  
Mammary Epithelial Cells ◽  
Mammary Epithelial ◽  
Mtor Signaling ◽  
Mtor Signaling Pathway ◽  
Bovine Mammary Epithelial Cells ◽  
Fat Synthesis ◽  
Milk Fat Synthesis

Abstract Background The biosynthesis of milk fat affects both the technological properties and organoleptic quality of milk and dairy products. MicroRNAs (miRNAs) are endogenous small non-coding RNAs that inhibit the expression of their mRNA targets and are involved in downstream signaling pathways that control several biological processes, including milk fat synthesis. miR-34b is a member of the miR-34 miRNA cluster, which is differentially expressed in the mammary gland tissue of dairy cows during lactation and dry periods. Previous studies have indicated miR-34b is a potential candidate gene that plays a decisive role in regulating milk fat synthesis; therefore, it is important to focus on miR-34b and investigate its regulatory effect on the biosynthesis of milk fat in bovine mammary epithelial cells (BMECs). Results In this study, elevated miR-34b levels reduced milk fat synthesis, upregulated 1,999 genes, and downregulated 2,009 genes in BMECs. Moreover, Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis of differentially expressed genes suggested that miR-34b may play an inhibitory role in milk fat synthesis via the protein kinase B (Akt)/mammalian target of rapamycin (mTOR) signaling pathway by reducing phosphorylation levels. Notably, the mTOR activator MHY1485 rescued the inhibitory effect of miR-34b. Furthermore, we demonstrated that retinoic acid-induced protein 14 (RAI14) is a target of miR-34b via TargetScan and immunofluorescence assays. RAI14 mRNA and protein levels were significantly decreased by the miR-34b mimic and increased by the miR-34b inhibitor. Moreover, the reduction in RAI14 levels led to the inhibition of the Akt/mTOR signaling pathway. Conclusions Overall, our results identified a miR-34b-RAI14-Akt/mTOR regulatory network, while also providing a theoretical basis for the molecular breeding of dairy cows.

scholarly journals Comparative proteome analysis reveals VPS28 regulates milk fat synthesis through ubiquitylation in bovine mammary epithelial cells

PeerJ ◽  
2020 ◽  
Vol 8 ◽  
pp. e9542
Author(s):  
Lily Liu ◽  
Qin Zhang
Keyword(s):  
Epithelial Cells ◽  
Milk Fat ◽  
Mammary Epithelial Cells ◽  
Proteome Analysis ◽  
Mammary Epithelial ◽  
Absolute Quantitation ◽  
Bovine Mammary Epithelial Cells ◽  
Fat Synthesis ◽  
Vacuolar Protein ◽  
Milk Fat Synthesis

In our previous study, we found that VPS28 (vacuolar protein sorting 28 homolog) could alter ubiquitylation level to regulate milk fat synthesis in bovine primary mammary epithelial cells (BMECs). While the information on the regulation of VPS28 on proteome of milk fat synthesis is less known, we explored its effect on milk fat synthesis using isobaric tags for relative and absolute quantitation assay after knocking down VPS28 in BMECs. A total of 2,773 proteins in three biological replicates with a false discovery rate of less than 1.2% were identified and quantified. Among them, a subset of 203 proteins were screened as significantly down-(111) and up-(92) regulated in VPS28 knockdown BMECs compared with the control groups. According to Gene Ontology analysis, the differentially expressed proteins were enriched in the “proteasome,” “ubiquitylation,” “metabolism of fatty acids,” “phosphorylation,” and “ribosome.” Meanwhile, some changes occurred in the morphology of BMECs and an accumulation of TG (triglyceride) and dysfunction of proteasome were identified, and a series of genes associated with milk fat synthesis, ubiquitylation and proteasome pathways were analyzed by quantitative real-time PCR. The results of this study suggested VPS28 regulated milk fat synthesis was mediated by ubiquitylation; it could be an important new area of study for milk fat synthesis and other milk fat content traits in bovine.

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