Orai calcium release-activated calcium modulator 1 (ORAI1) plays a role in endoplasmic reticulum stress in bovine mammary epithelial cells challenged with physiological levels of ketone bodies

Abstract IGF-binding protein (IGFBP)-3 is a multifunctional protein that can exert IGF-independent effects on apoptosis. Anisomycin (ANS) is a potent inducer of IGFBP-3 production in bovine mammary epithelial cells (MECs), and knockdown of IGFBP-3 attenuates ANS-induced apoptosis. IGFBP-3 is present in the nucleus and the conditioned media in response to ANS. The goal of this study was to determine whether ribotoxic stress induced by ANS or a second ribotoxin, deoxynivalenol (DON), specifically regulates transport of IGFBP-3 to the nucleus and to determine the pathway by which it traffics. In ribotoxin-treated cells, both endogenous IGFBP-3 and transfected IGFBP-3 translocated to the nucleus. Inhibition of the nuclear transport protein importin-β with importazole reduced ribotoxin-induced nuclear IGFBP-3. Immunoprecipitation studies showed that ANS induced the association of IGFBP-3 and importin-β, indicating that ribotoxins specifically induce nuclear translocation via an importin-β‒dependent mechanism. To determine whether secretion of IGFBP-3 is required for nuclear localization, cells were treated with Pitstop 2 or brefeldin A to inhibit clathrin-mediated endocytosis or overall protein secretion, respectively. Neither inhibitor affected nuclear localization of IGFBP-3. Although the IGFBP-3 present in both the nucleus and conditioned media was glycosylated, secreted IGFBP-3 exhibited a higher molecular weight. Deglycosylation experiments with endoglycosidase Hf and PNGase indicated that secreted IGFBP-3 completed transit through the Golgi apparatus, whereas intracellular IGFBP-3 exited from the endoplasmic reticulum before transit through the Golgi. In summary, ANS and DON specifically induced nuclear localization of nonsecreted IGFBP-3 via an importin-β‒mediated event, which may play a role in their ability to induce apoptosis in MECs.

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IGF-1 induces IRE1-XBP1–dependent endoplasmic reticulum biogenesis in bovine mammary epithelial cells

Journal of Dairy Science ◽

10.3168/jds.2021-20268 ◽

2021 ◽

Author(s):

Mst Mamuna Sharmin ◽

Satoko Hayashi ◽

Makoto Miyaji ◽

Hiroshi Ishizaki ◽

Hiroki Matsuyama ◽

...

Keyword(s):

Endoplasmic Reticulum ◽

Epithelial Cells ◽

Mammary Epithelial Cells ◽

Mammary Epithelial ◽

Bovine Mammary Epithelial Cells

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1,25-hydroxyvitamin D3 decreases endoplasmic reticulum stress-induced inflammatory response in mammary epithelial cells

PLoS ONE ◽

10.1371/journal.pone.0228945 ◽

2020 ◽

Vol 15 (2) ◽

pp. e0228945

Author(s):

Gaiping Wen ◽

Klaus Eder ◽

Robert Ringseis

Keyword(s):

Endoplasmic Reticulum ◽

Epithelial Cells ◽

Inflammatory Response ◽

Endoplasmic Reticulum Stress ◽

Mammary Epithelial Cells ◽

Mammary Epithelial

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Polydatin Protects Bovine Mammary Epithelial Cells against Zearalenone-Induced Apoptosis by Inhibiting Oxidative Responses and Endoplasmic Reticulum Stress

Toxins ◽

10.3390/toxins13020121 ◽

2021 ◽

Vol 13 (2) ◽

pp. 121

Author(s):

Yurong Fu ◽

Yongcheng Jin ◽

Anshan Shan ◽

Jing Zhang ◽

Hongyu Tang ◽

...

Keyword(s):

Endoplasmic Reticulum ◽

Treatment Group ◽

Epithelial Cells ◽

Er Stress ◽

Cell Apoptosis ◽

Mammary Epithelial Cells ◽

Mammary Epithelial ◽

Bovine Mammary Epithelial Cells ◽

Stress Related Genes ◽

Induced Apoptosis

Zearalenone (ZEA) is a mycotoxin of the Fusarium genus that can cause endoplasmic reticulum (ER) stress and Apoptosis in bovine mammary epithelial cells (MAC-T). Polydatin (PD), a glycoside purified from Polygonum cuspidatum, has antioxidant properties. This study aimed to explore whether PD can alleviate ZEA-induced damage on bovine mammary epithelial cells (MAC-T). We found that incasing the concentration of ZEA (0, 7.5, 15, 30, 60, 90, 120, and 240 μM) gradually decreased the cell viability. PD treatment alone at 5, 10, and 20 μM did not affect cell viability. Follow-up studies then applied 30 μM of ZEA and 5 μM of PD to treat cells; the results showed that the ZEA + PD treatment group effectively reduced cell oxidative damage compared with the ZEA treatment group. The qPCR analysis showed that ZEA treatment significantly up-regulated the expression of ER stress-related genes, relative to the control. However, adding PD significantly down-regulated the expression of ER stress-related genes. The cell apoptosis detection results showed that, compared with the ZEA treatment group, the ZEA + PD treatment group down-regulated the Bax gene and up-regulated the Bcl-2 gene expressions, which reduced the cell apoptosis rate and Caspase-3 activity. Taken together, these results indicate that PD reduces ZEA-induced apoptosis by inhibiting oxidative damage and ER stress.

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5-ALA Attenuates the Palmitic Acid-Induced ER Stress and Apoptosis in Bovine Mammary Epithelial Cells

Molecules ◽

10.3390/molecules26041183 ◽

2021 ◽

Vol 26 (4) ◽

pp. 1183

Author(s):

Mst Mamuna Sharmin ◽

Md Aminul Islam ◽

Itsuki Yamamoto ◽

Shin Taniguchi ◽

Shinichi Yonekura

Keyword(s):

T Cells ◽

Epithelial Cells ◽

Palmitic Acid ◽

Er Stress ◽

Aminolevulinic Acid ◽

Mammary Epithelial Cells ◽

Mammary Epithelial ◽

Protective Effects ◽

Bovine Mammary Epithelial Cells ◽

Induced Apoptosis

The conservation of mammary gland physiology by maintaining the maximum number of mammary epithelial cells (MECs) is of the utmost importance for the optimum amount of milk production. In a state of negative energy balance, palmitic acid (PA) reduces the number of bovine MECs. However, there is no effective strategy against PA-induced apoptosis of MECs. In the present study, 5-aminolevulinic acid (5-ALA) was established as a remedial agent against PA-induced apoptosis of MAC-T cells (an established line of bovine MECs). In PA-treated cells, the apoptosis-related genes BCL2 and BAX were down- and upregulated, respectively. The elevated expression of major genes of the unfolded protein response (UPR), such as CHOP, a proapoptotic marker (C/EBP homologous protein), reduced the viability of PA-treated MAC-T cells. In contrast, 5-ALA pretreatment increased and decreased BCL2 and BAX expression, respectively. Moreover, cleaved caspase-3 protein expression was significantly reduced in the 5-ALA-pretreated group in comparison with the PA group. The downregulation of major UPR-related genes, including CHOP, extended the viability of MAC-T cells pretreated with 5-ALA and also reduced the enhanced intensity of the PA-induced expression of phospho-protein kinase R-like ER kinase. Moreover, the enhanced expression of HO-1 (antioxidant gene heme oxygenase) by 5-ALA reduced PA-induced oxidative stress (OxS). HO-1 is not only protective against OxS but also effective against ER stress. Collectively, these findings offer new insights into the protective effects of 5-ALA against PA-induced apoptosis of bovine MECs.

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

Journal of Dairy Research ◽

10.1017/s0022029919000232 ◽

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.

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