Effects of selenium on selenoprotein synthesis and antioxidant parameters of bovine mammary epithelial cells

This study aimed to investigate the effects of selenium (Se) on the selenoproteins synthesis and antioxidant parameters of bovine mammary epithelial cells (BMECs). The experiment was conducted as a single factor completely randomized design to explore the effect of different levels of Se supplementation (0, 10, 20, 50, and 100 nmol/l) on selenoproteins synthesis and antioxidant parameters of BMECs, and to screen the appropriate dose of Se supplementation ensuring a better antioxidant function. Se supplementation increased cell proliferation, the activities of glutathione peroxidase (GPx) and superoxide dismutase, total antioxidant capacity and seleoprotein P (SelP) content, and decreased reactive oxygen species and malondialdehyde levels in a dose-dependent manner. Se supplementation of 50–100 nmol/l had a better effect. Se supplementation also increased thioredoxin reductase (TrxR) activity in a dose-dependent manner, and Se supplementation of 20–50 nmol/l had a better promoting effect. The dose-dependent response between Se supplementation and mRNA and protein expression of GPx1 and TrxR1, as well as SelP mRNA expression was also observed in this experiment. The mRNA and protein expression of GPx1 was up-regulated with the addition of 50–100 nmol/l Se, and the mRNA expression of TrxR1 and SelP was up-regulated with the addition of 20–100 nmol/l Se. Results indicated that Se supplementation of 50 nmol/l had a better promoting effect on the selenoproteins synthesis and antioxidant parameters of BMECs.

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

Animals ◽

10.3390/ani11113238 ◽

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.

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Bacterial Lipopolysaccharide Induced Alterations of Genome-Wide DNA Methylation and Promoter Methylation of Lactation-Related Genes in Bovine Mammary Epithelial Cells

Toxins ◽

10.3390/toxins11050298 ◽

2019 ◽

Vol 11 (5) ◽

pp. 298 ◽

Cited By ~ 9

Author(s):

Jingbo Chen ◽

Yongjiang Wu ◽

Yawang Sun ◽

Xianwen Dong ◽

Zili Wang ◽

...

Keyword(s):

Dna Methylation ◽

Immune Response ◽

Mrna Expression ◽

Promoter Methylation ◽

Mammary Epithelial Cells ◽

Mammary Epithelial ◽

Bovine Mammary Epithelial Cells ◽

Genome Wide ◽

Immune Response Pathway ◽

Gene Mrna

Bacterial lipopolysaccharide (LPS) could result in poor lactation performance in dairy cows. High methylation of DNA is associated with gene repression. However, it is unclear whether LPS could suppress the expression of lactation-related genes by inducing DNA methylation. Therefore, the objective of this study was to investigate the impact of LPS on genome-wide DNA methylation, using methylated DNA immunoprecipitation with high-throughput sequencing (MeDIP-seq) and on the promoter methylation of lactation-related genes using MassArray analysis in bovine mammary epithelial cells. The bovine mammary epithelial cell line MAC-T cells were treated for 48 h with LPS at different doses of 0, 1, 10, 100, and 1000 endotoxin units (EU)/mL (1 EU = 0.1 ng). The results showed that the genomic methylation levels and the number of methylated genes in the genome as well as the promoter methylation levels of milk genes increased when the LPS dose was raised from 0 to 10 EU/mL, but decreased after further increasing the LPS dose. The milk gene mRNA expression levels of the 10 EU/mL LPS treatment were significantly lower than these of untreated cells. The results also showed that the number of hypermethylated genes was greater than that of hypomethylated genes in lipid and amino acid metabolic pathways following 1 and 10 EU/mL LPS treatments as compared with control. By contrast, in the immune response pathway the number of hypomethylated genes increased with increasing LPS doses. The results indicate LPS at lower doses induced hypermethylation of the genome and promoters of lactation-related genes, affecting milk gene mRNA expression. However, LPS at higher doses induced hypomethylation of genes involved in the immune response pathway probably in favor of immune responses.

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Caffeic Acid Prevented LPS-Induced Injury of Primary Bovine Mammary Epithelial Cells through Inhibiting NF-κB and MAPK Activation

Mediators of Inflammation ◽

10.1155/2019/1897820 ◽

2019 ◽

Vol 2019 ◽

pp. 1-12 ◽

Cited By ~ 4

Author(s):

Mingjiang Liu ◽

Guoqing Fang ◽

Shaojie Yin ◽

Xin Zhao ◽

Chi Zhang ◽

...

Keyword(s):

Epithelial Cells ◽

Cell Viability ◽

Signaling Pathways ◽

Caffeic Acid ◽

Proinflammatory Cytokines ◽

Mammary Epithelial Cells ◽

Mammary Epithelial ◽

Signal Pathways ◽

Dependent Manner ◽

Bovine Mammary Epithelial Cells

In our previous study, lipopolysaccharide (LPS) significantly reduced the cell viability of primary bovine mammary epithelial cells (bMEC) leading to cell apoptosis, which were prevented by caffeic acid (CA) through inhibiting NF-κB activation and reducing proinflammatory cytokine expression. While the underlying mechanism remains unclear, here, we determined that LPS induced the extensive microstructural damage of bMEC, especially the mitochondria and endoplasmic reticulum. Then, the obvious reduction of mitochondrial membrane potential and expression changes of apoptosis-associated proteins (Bcl-2, Bax, and casepase-3) indicated that apoptosis signaling through the mitochondria should be responsible for the cell viability decrease. Next, the high-throughput cDNA sequencing (RNA-Seq) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis were employed to verify that the MAPK and JAK-STAT signaling pathways also were the principal targets of LPS. Following, the critical proteins (ERK, JNK, p38, and c-jun) of the MAPK signaling pathways were activated, and the release of proinflammatory cytokines (TNF-α, IL-1β, IL-6, and IL-8) regulated by NF-κB and MAPKs was significantly increased, which can promote a cascade of inflammation that induces cell injury and apoptosis. Meanwhile, CA significantly inhibited the activation of MAPKs and the release of proinflammatory cytokines in a dose-dependent manner, which were similar to its effects on the NF-κB activation that we previously published. So we concluded that CA regulates the proteins located in the upstream of multiple cell signal pathways which can reduce the LPS-induced activation of NF-κB and MAPKs, thus weakening the inflammatory response and maintaining cell structure and function, which accordingly inhibit apoptosis.

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Inhibition of Staphylococcus aureus Invasion into Bovine Mammary Epithelial Cells by Contact with Live Lactobacillus casei

Applied and Environmental Microbiology ◽

10.1128/aem.03323-12 ◽

2012 ◽

Vol 79 (3) ◽

pp. 877-885 ◽

Cited By ~ 59

Author(s):

Damien S. Bouchard ◽

Lucie Rault ◽

Nadia Berkova ◽

Yves Le Loir ◽

Sergine Even

Keyword(s):

Staphylococcus Aureus ◽

Epithelial Cells ◽

Lactobacillus Casei ◽

Mammary Epithelial Cells ◽

Control Strategies ◽

Mammary Epithelial ◽

Dairy Herds ◽

Dependent Manner ◽

Content Type ◽

Bovine Mammary Epithelial Cells

ABSTRACTStaphylococcus aureusis a major pathogen that is responsible for mastitis in dairy herds.S. aureusmastitis is difficult to treat and prone to recurrence despite antibiotic treatment. The ability ofS. aureusto invade bovine mammary epithelial cells (bMEC) is evoked to explain this chronicity. One sustainable alternative to treat or prevent mastitis is the use of lactic acid bacteria (LAB) as mammary probiotics. In this study, we tested the ability ofLactobacillus caseistrains to prevent invasion of bMEC by twoS. aureusbovine strains, RF122 and Newbould305, which reproducibly induce acute and moderate mastitis, respectively.L. caseistrains affected adhesion and/or internalization ofS. aureusin a strain-dependent manner. Interestingly,L. caseiCIRM-BIA 667 reducedS. aureusNewbould305 and RF122 internalization by 60 to 80%, and this inhibition was confirmed for two otherL. caseistrains, including one isolated from bovine teat canal. The protective effect occurred without affecting bMEC morphology and viability. Once internalized, the fate ofS. aureuswas not affected byL. casei. It should be noted thatL. caseiwas internalized at a low rate but survived in bMEC cells with a better efficiency than that ofS. aureusRF122. Inhibition ofS. aureusadhesion was maintained with heat-killedL. casei, whereas contact between liveL. caseiandS. aureusor bMEC was required to preventS. aureusinternalization. This first study of the antagonism of LAB towardS. aureusin a mammary context opens avenues for the development of novel control strategies against this major pathogen.

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Effects of Sodium Octanoate on Innate Immune Response of Mammary Epithelial Cells duringStaphylococcus aureusInternalization

BioMed Research International ◽

10.1155/2013/927643 ◽

2013 ◽

Vol 2013 ◽

pp. 1-8 ◽

Cited By ~ 6

Author(s):

Nayeli Alva-Murillo ◽

Alejandra Ochoa-Zarzosa ◽

Joel E. López-Meza

Keyword(s):

Immune Response ◽

Epithelial Cells ◽

Mrna Expression ◽

Innate Immune Response ◽

Mammary Epithelial Cells ◽

Short Chain Fatty Acids ◽

Mammary Epithelial ◽

Innate Immune ◽

Sodium Octanoate ◽

Bovine Mammary Epithelial Cells

Bovine mammary epithelial cells (bMECs) are capable of initiating an innate immune response to invading bacteria. Short chain fatty acids can reduceStaphylococcus aureusinternalization into bMEC, but it has not been evaluated if octanoic acid (sodium octanoate, NaO), a medium chain fatty acid (MCFA), has similar effects. In this study we determined the effect of NaO onS. aureusinternalization into bMEC and on the modulation of innate immune elements. NaO (0.25–2 mM) did not affectS. aureusgrowth and bMEC viability, but it differentially modulated bacterial internalization into bMEC, which was induced at 0.25–0.5 mM (~60%) but inhibited at 1-2 mM (~40%). Also, bMEC showed a basal expression of all the innate immune genes evaluated, which were induced byS. aureus. NaO induced BNBD4, LAP, and BNBD10 mRNA expression, but BNBD5 and TNF-αwere inhibited. Additionally, the pretreatment of bMEC with NaO inhibited the mRNA expression induction generated by bacteria which coincides with the increase in internalization; only TAP and BNDB10 showed an increase in their expression; it coincides with the greatest effect on the reduction of bacterial internalization. In conclusion, NaO exerts a dual effect onS. aureusinternalization in bMEC and modulates elements of innate immune response.

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FGF21 Ameliorates Hepatic Fibrosis by Multiple Mechanisms

10.21203/rs.3.rs-418625/v1 ◽

2021 ◽

Author(s):

Fanrui Meng ◽

Mir Hassan Khoso ◽

Kai Kang ◽

Qi He ◽

Yukai Cao ◽

...

Keyword(s):

Protein Expression ◽

Western Blot ◽

Hepatic Fibrosis ◽

Blot Analysis ◽

Western Blot Analysis ◽

Cell Model ◽

Dependent Manner ◽

Rt Pcr ◽

Mrna And Protein Expression ◽

Dose Dependent

Abstract Previous study reports that FGF21 could ameliorate hepatic fibrosis, but its mechanisms have not been fully investigated. In this study, three models were used to investigate the mechanism by which FGF21 alleviates liver fibrosis. CCL4 and DMN were respectively used to induce hepatic fibrosis animal models. Our results demonstrated that liver index and liver function were deteriorated in both models. HE and Masson’s staining showed that the damaged tissue architectonics were observed in the mice of both models. Treatment with FGF21 significantly ameliorated these changes. ELISA analysis showed that the serum levels of IL-1β, IL-6 and TNF-α were significantly elevated in both models. However, administration of FGF21 significantly reduced these inflammatory cytokines. RT-PCR and Western blot analysis showed that mRNA and protein expression of collagenI, α-SMA and TGF-β were significantly decreased by treatment with FGF21. PDGF-BB stimulant was used to establish the experimental cell model in HSCs. RT-PCR and Western blot analysis demonstrated that the expression of collagenI and α-SMA were significantly upregulated by this stimulant in model group. Interestingly, our results showed that mRNA and protein expression of leptin were also significantly induced in PDGF-BB treated HSCs. Administration of FGF21 could significantly reduce leptin expression in a dose dependent manner and these effects were reversed in siRNA (against β-klotho) transfected HSCs. Furthermore, the leptin signaling pathways related protein p-ERK/t-ERK, p-STAT3/STAT3 and TGF-β were significantly downregulated by FGF21 treatment in a dose dependent manner. The expression of SOCS3 and Nrf-2 were enhanced by treatment with FGF21. The underlying mechanism may be that FGF21 regulates leptin-STAT3 axis via Nrf-2 and SOCS3 pathway in activated HSCs.

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The role of native bovine α-lactalbumin in bovine mammary epithelial cell apoptosis and casein expression

Journal of Dairy Research ◽

10.1017/s0022029908003403 ◽

2008 ◽

Vol 75 (3) ◽

pp. 319-325 ◽

Cited By ~ 4

Author(s):

Lisa G Riley ◽

Peter C Wynn ◽

Peter Williamson ◽

Paul A Sheehy

Keyword(s):

Epithelial Cells ◽

Mrna Expression ◽

Mammary Epithelial Cells ◽

Cell Types ◽

Mammary Epithelial ◽

Major Influence ◽

Expression Levels ◽

Bovine Mammary Epithelial Cells ◽

Epithelial Cell Apoptosis

Folding variants of α-lactalbumin (α-la) are known to induce cell death in a number of cell types, including mammary epithelial cells (MEC). The native conformation of α-la however has not been observed to exhibit this biological activity. Here we report that native bovine α-la reduced the viability of primary bovine mammary epithelial cells (BMEC) and induced caspase activity in mammospheres, which are alveolar-like structures formed by culturing primary BMEC on extracellular matrix in the presence of lactogenic hormones. These observations suggest a possible role for bovine α-la in involution and/or maintaining the luminal space in mammary alveoli during lactation. In addition, co-incubation of bovine α-la in an in-vitro mammosphere model resulted in decreased β-casein mRNA expression and increased αs1- and κ-casein mRNA expression. This differential effect on casein expression levels is unusual and raises the possibility of manipulating expression levels of individual caseins to alter dairy processing properties. Manipulation of α-la levels could be further investigated for its potential to enhance milk protein expression and/or improve lactational persistency by influencing the balance between proliferation and apoptosis of BMEC, which has a major influence on the milk-producing capacity of the mammary gland.

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Growth hormone acts on the synthesis and secretion of α-casein in bovine mammary epithelial cells

Journal of Dairy Research ◽

10.1017/s0022029905000889 ◽

2005 ◽

Vol 72 (3) ◽

pp. 264-270 ◽

Cited By ~ 20

Author(s):

Kazuhito Sakamoto ◽

Tokushi Komatsu ◽

Takuya Kobayashi ◽

Michael T Rose ◽

Hisashi Aso ◽

...

Keyword(s):

Growth Hormone ◽

Epithelial Cells ◽

Mrna Expression ◽

Mammary Epithelial Cells ◽

Mammary Epithelial ◽

Gh Treatment ◽

Bovine Mammary Epithelial Cells ◽

Lactogenic Hormone ◽

Mammary Epithelia

To study the effect of growth hormone (GH) on the functions of mammary epithelia, we examined the effect of GH on the synthesis and secretion of α-casein in a bovine mammary epithelial cell (BMEC) clonal line, which was established from a 26-d-pregnant Holstein heifer. GH receptors (GHR) were observed in the BMEC on the membrane as well as in the cytoplasm. After BMEC were plated onto cell culture inserts, GH stimulated α-casein release in both the presence and absence of the lactogenic hormone complex, which included dexamethasone, insulin and prolactin (DIP). DIP enhanced the effect of GH on α-casein release. Although αs1-casein mRNA expression was not detected in untreated control cells, its expression was observed in BMEC in response to the GH, DIP and GH+DIP treatments. Expression was greater for GH and GH+DIP than for just DIP. Expression of GHR mRNA was increased by DIP treatment, while the mRNA expression was little changed by GH treatment. We conclude that GH acts on BMEC and induces the expression of both the α-casein gene and protein. GHR gene expression was shown to be regulated by DIP and GHR. GHR may be involved in a synergic effect between GH and DIP on casein secretion. These results suggest that GH, in addition to its widely accepted homeorhetic role in vivo, also can act on the mammary parenchyma, and that the effects of GH on mammary epithelial cells could partly account for the clear galactopoietic effect of recombinant bovine GH seen in lactating dairy cows.

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Lactoferrin decreases primary bovine mammary epithelial cell viability and casein expression

Journal of Dairy Research ◽

10.1017/s0022029907002920 ◽

2008 ◽

Vol 75 (2) ◽

pp. 135-141 ◽

Cited By ~ 11

Author(s):

Lisa G Riley ◽

Peter Williamson ◽

Peter C Wynn ◽

Paul A Sheehy

Keyword(s):

Gene Expression ◽

Mrna Expression ◽

Milk Protein ◽

Mammary Epithelial Cells ◽

Mammary Epithelial ◽

Regulatory Role ◽

Bovine Mammary Epithelial Cells ◽

Lactogenic Hormones ◽

Interfering Rna

The concentration of lactoferrin (LTF) in milk varies during lactation, rising sharply during involution. We proposed that LTF might have a regulatory role in involution and investigated its effects in vitro on the viability of bovine mammary epithelial cells (BMEC) and on casein expression in bovine mammospheres. Mammospheres capable of milk protein expression were formed by culturing primary BMEC on extracellular matrix in the presence of lactogenic hormones. Exogenously added LTF decreased β-casein and κ-casein mRNA expression in mammospheres while transfection of a short interfering RNA (siRNA) to suppress LTF expression resulted in increased casein mRNA expression. We believe that LTF exerts its effect on casein gene expression by up-regulating interleukin-1β (IL-1β) as IL-Iβ gene expression was elevated in mammospheres treated with LTF. LTF also decreased viability of BMEC grown as monolayers and as mammospheres. Interestingly, LTF was only effective in reducing casein mRNA expression and viability in mammospheres when added at concentrations found during early involution but was inactive when used at concentrations found in milk. We suggest that LTF has a regulatory role during early involution, decreasing casein expression and reducing BMEC viability.

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Silibinin Inhibits TGF-β-induced MMP-2 and MMP-9 Through Smad Signaling Pathway in Colorectal Cancer HT-29 Cells

Basic & Clinical Cancer Research ◽

10.18502/bccr.v12i2.5752 ◽

2021 ◽

Author(s):

Zahra Zare ◽

Tina Nayerpour dizaj ◽

Armaghan Lohrasbi ◽

Zakieh Sadat Sheikhalishahi ◽

Amirhooman Asadi ◽

...

Keyword(s):

Colorectal Cancer ◽

Protein Expression ◽

Cell Viability ◽

Cancer Cells ◽

Cell Line ◽

Dependent Manner ◽

Smad2 Phosphorylation ◽

Mrna And Protein Expression ◽

Dose Dependent ◽

Ht 29

Background: Metastasis of cancer cells is the primary responsible for death in patients with colorectal cancer (CRC). Transforming growth factor-β (TGF-β)-induced matrix metalloproteinases (MMPs) are essential for the metastasis process. Silibinin is a natural compound extracted from the Silybum marianum that exhibits anti-neoplastic activity in cancer cell lines. In this study, we evaluated the effects of silibinin on MMP-2 and MMP-9 induced by TGF-β in human HT-29 CRC cell line and the potential mechanism underlying the effects. Methods: The present in vitro study was done on the HT-29 cell line. The HT-29 cell line was cultured in RPMI1640 and exposed to TGF- β (5 ng/ml) in the absence and presence of different concentrations of silibinin (10, 25, 50, and 100 μM). The effect of silibinin on HT-29 cell viability was measured with the MTT assay. A real-time polymerase chain reaction (Real-Time PCR) determined the relative mRNA expression of MMP-2 and MMP-9. Western blotting was employed to examine MMP-2 and MMP 9 protein expression and Smad2 phosphorylation. Results: Silibinin inhibits cell viability of HT-29 cell line at 24 hours in a dose-dependent manner. TGF-β increased the mRNA and protein expression of MMP-2, MMP-9, and phosphorylated Smad2 compared to controls. Pharmacological inhibition with silibinin markedly blocked TGF-β–induced MMP-2 and MMP-9 mRNA and protein expression and Smad2 phosphorylation. Conclusion: Silibinin decreased the cell viability of HT-29 cancer cells in a dose-dependent manner. Silibinin also inhibited TGF-β-stimulated MMP-2 and MMP-9 expression in HT-29 cells, possibly mediated with the Smad2 signaling pathway.

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