Glutamine pretreatment protects bovine mammary epithelial cells from inflammation and oxidative stress induced by γ-d-glutamyl-meso-diaminopimelic acid (iE-DAP).

Abstract

Glutamine (GLN) has many types of biological activity in rats, including anti-inflammatory, antioxidative stress, and anti-apoptosis effects. However, little is known about the effects of GLN on bovine mammary epithelial cells (BMEC). γ-d-Glutamyl-meso-diaminopimelic acid (iE-DAP) is a cell wall peptidoglycan component of gram-negative bacteria that can be recognized by the intracellular receptor nucleotide-binding oligomerization domain-containing protein 1 (NOD1) and can cause bovine mastitis. The goal of the present study was to investigate whether GLN protects BMEC from iE-DAP-induced inflammation, oxidative stress, and apoptosis. We cultured BMEC in a GLN-free medium for 24 h and then separated them into 4 groups: cells treated with 1× PBS for 26 or 32 h (control); cells stimulated by 10 μg/mL iE-DAP for 2 or 8 h (2- or 8-h iE-DAP); cells pretreated with 8 or 4 mM GLN for 24 h followed by 2 or 8 h of 1× PBS treatment (8 or 4 mM GLN); and cells pretreated with 8 or 4 mM GLN for 24 h followed by 2 or 8 h of iE-DAP treatment (DG). In the 2-h iE-DAP group, when levels of inflammation peaked, iE-DAP treatment increased both the mRNA and protein expression of NOD1, inhibitor of nuclear factor-κB (NFKBIA, IκB), and nuclear factor-κB subunit p65 (RELA, NF-κB p65), as well as the mRNA expression of IL6 and IL8 and levels of IL-6 and tumor necrosis factor-α in cell culture supernatants. In contrast, 8 mM GLN pretreatment inhibited the mRNA and protein expression of inflammatory-related factors by suppressing the NOD1/NF-κB pathway. In the 8-h iE-DAP group, iE-DAP treatment decreased the mRNA and protein expression of extracellular regulated kinase (Erk, ERK) and nuclear factor erythroid 2-associated factor2 (NFE2L2, Nrf2), as well as the mRNA expression of superoxide dismutase 1 (SOD1), catalase (CAT), coenzyme II oxidoreductase 1 (NQO1), and heme oxygenase 1 (HMOX1, HO1). In addition, iE-DAP treatment increased the expression of malondialdehyde in BMEC when oxidative stress levels peaked. Interestingly, 4 mM GLN pretreatment induced the mRNA and protein expression of antioxidative stress-related factors and inhibited the expression of reactive oxygen species in BMEC by promoting the ERK/Nrf2 pathway. Moreover, GLN reduced apoptosis caused by inflammation and oxidative stress in BMEC. This is the first report showing that GLN protects against iE-DAP-induced inflammation and oxidative stress via the NOD1/NF-κB and ERK/Nrf2 pathways in BMEC.

• Authors: Xiaoye Cheng, Zain-Ui Aabdin, Yan Wang, Nana Ma, Hongyu Dai, Xiaoli Shi, Xiangzhen Shen
• Journal: Journal of dairy science (J Dairy Sci) Vol. 104 Issue 2 Pg. 2123-2139 (Feb 2021) ISSN: 1525-3198 [Electronic] United States
• PMID: 33358155 (Publication Type: Journal Article)
• Copyright: Copyright © 2021 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

Chemical References

• NF-kappa B
• Reactive Oxygen Species
• Transcription Factor RelA
• Glutamine
• Diaminopimelic Acid
• N(2)-(gamma-D-glutamyl)-meso-2,2'-diaminopimelic acid
• Heme Oxygenase-1

Topics

• Animals
• Apoptosis (drug effects)
• Cattle
• Cattle Diseases (prevention & control)
• Cells, Cultured
• Diaminopimelic Acid (analogs & derivatives, antagonists & inhibitors)
• Epithelial Cells (metabolism)
• Female
• Glutamine (therapeutic use)
• Heme Oxygenase-1 (metabolism)
• Inflammation (chemically induced, prevention & control, veterinary)
• Mammary Glands, Animal (cytology, drug effects)
• NF-kappa B (metabolism)
• Oxidative Stress (drug effects)
• Reactive Oxygen Species (metabolism)
• Transcription Factor RelA (metabolism, pharmacology)