14-3-3γaffects eIF5 to regulateβ-casein synthesis in bovine mammary epithelial cells

2016 ◽  
Vol 96 (4) ◽  
pp. 478-487
Author(s):  
Cuiping Yu ◽  
Chaochao Luo ◽  
Xinyu Gu ◽  
Yanli Zang ◽  
Bo Qu ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Regulatory Mechanism ◽  
Mammary Epithelial Cells ◽  
Mammary Epithelial ◽  
Initiation Factor ◽  
Biological Processes ◽  
Bovine Mammary Epithelial Cells ◽  
Protein Levels ◽  
Casein Synthesis

The 14-3-3γ protein participates in many biological processes; however, its regulatory mechanism in milk protein synthesis is not well studied. We hypothesized that 14-3-3γ might affect eIF5 (an initiation factor) to regulate β-casein synthesis in dairy cows. In this study, a possible interaction between 14-3-3γ and eIF5 was investigated using bovine mammary epithelial cells (BMECs). The expression levels of 14-3-3γ and eIF5 in the mammary gland tissues from cows producing higher quality milk were higher than those from cows producing low-quality milk. Moreover, the expression of 14-3-3γ, eIF5, and β-casein were increased at both mRNA and protein levels in BMECs cultured in vitro with methionine (Met) supplementation. Coimmunoprecipitation, colocalization, and FRET analysis further showed the evidences that 14-3-3γ physically bound to eIF5 in BMECs. Gene function studies revealed that 14-3-3γ positively regulated eIF5 through alteration of eIF2α/p-eIF2α ratio. Collectively, our data suggest that 14-3-3γ regulates β-casein translation in BMECs through interaction with eIF5.

Exogenous phospholipase A2 affects inflammatory gene expression in primary bovine mammary epithelial cells

2019 ◽  
Vol 86 (2) ◽  
pp. 177-180
Author(s):  
Jacqueline P. Kurz ◽  
Mark P. Richards ◽  
Matthew Garcia ◽  
Zhongde Wang
Keyword(s):  
Epithelial Cells ◽  
Phospholipase A2 ◽  
Inflammatory Responses ◽  
Mammary Epithelial Cells ◽  
Constitutive Expression ◽  
Mammary Epithelial ◽  
Inflammatory Factors ◽  
Bovine Mammary Epithelial Cells ◽  
Lps Challenge

AbstractThis Research Communication addresses the hypothesis that exogenously administered phospholipase A2 (PLA2) affects the inflammatory responses of bovine mammary epithelial cells (bMEC) in vitro with the aim of providing preliminary justification of investigation into the uses of exogenously administered PLA2 to manage or treat bovine mastitis. Primary bMEC lines from 11 lactating Holstein dairy cows were established and the expression of 14 pro-inflammatory genes compared under unchallenged and lipopolysaccharide (LPS)-challenged conditions, with and without concurrent treatment with bovine pancreatic PLA2G1B, a secreted form of PLA2. No differences in the expression of these genes were noted between PLA2-treated and untreated bMEC under unchallenged conditions. Following LPS challenge, untreated bMEC exhibited significant downregulation of CXCL8, IL1B, CCL20, and CXCL1. In contrast, PLA2-treated bMEC exhibited significant downregulation of IL1B and CCL20 only. These findings indicate that exogenous PLA2 affects the expression of some pro-inflammatory factors in immune-stimulated bMEC, but does not influence the constitutive expression of these factors. Further investigation of the influence of exogenous PLA2 in the bovine mammary gland is justified.

scholarly journals Dipeptide (Methionyl-Methionine) Transport and Its Effect on β-Casein Synthesis in Bovine Mammary Epithelial Cells

2018 ◽  
Vol 49 (2) ◽  
pp. 479-488 ◽  
Author(s):  
Caihong Wang ◽  
Fengqi Zhao ◽  
Jianxin Liu ◽  
Hongyun Liu
Keyword(s):  
Epithelial Cells ◽  
Cell Viability ◽  
Transport Properties ◽  
Mammary Epithelial Cells ◽  
Mammary Epithelial ◽  
Janus Kinase ◽  
Initiation Factor ◽  
Peptide Transporter ◽  
Small Interference Rna ◽  
Bovine Mammary Epithelial Cells

Background/Aims: The aim of this study was to investigate the transport properties and utilization of methionyl-methionine dipeptide (Met-Met) in β-casein (β-CN) synthesis in bovine mammary epithelial cells (BMECs). Methods: The transport properties were studied for the effects of time, pH, concentration, temperature and inhibitors using Met-Met-FITC in BMECs. BMECs were treated with different concentrations of Met-Met (0, 20, 40, 80, 120 and 160 µg/ml). In several experiments, the cells were treated with Janus kinase 2 (JAK2) inhibitor (tyrphostin AG-490, 50 µM) and mammalian target of rapamycin (mTOR) inhibitor (rapamycin, 100 ng/ml). Results: The uptake of Met-Met-FITC by BMECs was rapid during the first fifteen minutes and became saturated after 15 minutes. The transport of Met-Met-FITC in BMECs exhibited a Michaelis constant of 52.4 µM and maximum transport velocity of 14.8 pmol/min/mg protein. The uptake of Met-Met-FITC in BMECs was pH-dependent, peaked at pH 6.5 and was significantly inhibited by other peptides, including Met-Lys, Lys-Lys, Gly-Met, Gly-Leu and Met-Leu. Knocking down the peptide transporter 2 (PepT2) with small interference RNA markedly decreased Met-Met-FITC uptake. Met-Met concentration-dependently increased the PepT2 expression and β-CN synthesis in BMECs with an optimal concentration of 80 µg/ml. At 80 µg/ml, Met-Met also enhanced the cell viability and cyclin D1 expression and promoted cell cycle transition from G1 phase to S phase. In addition, 80 µg/ml Met-Met increased the mRNA abundance of JAK2 and signal transducer and activator of transcription 5 (STAT5) and enhanced the phosphorylation of JAK2, STAT5, mTOR, p70 ribosomal S6 kinase 1 and eukaryotic initiation factor 4E binding protein 1. The inhibition of JAK2 and mTOR significantly decreased Met-Met-induced increase in cell viability and β-CN synthesis in BMECs. Conclusion: Our data elucidated the properties of peptide transporter and its effect on β-CN synthesis in BMECs. Met-Met, taken up by PepT2, enhances cell proliferation and promotes β-CN synthesis by activating JAK2-STAT5 and mTOR signaling pathways in BMECs.

scholarly journals Bacteriophages isolated from dairy farm mitigated Klebsiella pneumoniae-induced inflammation in bovine mammary epithelial cells cultured in vitro

2021 ◽  
Vol 17 (1) ◽  
Author(s):  
Yuxiang Shi ◽  
Wenpeng Zhao ◽  
Gang Liu ◽  
Tariq Ali ◽  
Peng Chen ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Klebsiella Pneumoniae ◽  
Inflammatory Responses ◽  
Mammary Epithelial Cells ◽  
Therapeutic Potential ◽  
Morphological Changes ◽  
Mammary Epithelial ◽  
Bovine Mammary Epithelial Cells ◽  
Tnf Α

Abstract Background Klebsiella pneumoniae, an environmental pathogen causing mastitis in dairy cattle, is often resistant to antibiotics. K. pneumoniae was used as the host bacteria to support bacteriophage replication; 2 bacteriophages, CM8-1 and SJT-2 were isolated and considered to have therapeutic potential. In the present study, we determined the ability of these 2 bacteriophages to mitigate cytotoxicity, pathomorphological changes, inflammatory responses and apoptosis induced by K. pneumoniae (bacteriophage to K. pneumoniae MOI 1:10) in bovine mammary epithelial cells (bMECs) cultured in vitro. Results Bacteriophages reduced bacterial adhesion and invasion and cytotoxicity (lactate dehydrogenase release). Morphological changes in bMECs, including swelling, shrinkage, necrosis and hematoxylin and eosin staining of cytoplasm, were apparent 4 to 8 h after infection with K. pneumoniae, but each bacteriophage significantly suppressed damage and decreased TNF-α and IL-1β concentrations. K. pneumoniae enhanced mRNA expression of TLR4, NF-κB, TNF-α, IL-1β, IL-6, IL-8, caspase-3, caspase-9 and cyt-c in bMECs and increased apoptosis of bMECs, although these effects were mitigated by treatment with either bacteriophage for 8 h. Conclusions Bacteriophages CM8-1 and SJT-2 mitigated K. pneumoniae-induced inflammation in bMECs cultured in vitro. Therefore, the potential of these bacteriophages for treating mastitis in cows should be determined in clinical trials.

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