title: 14-3-3γ Regulates Lipopolysaccharide-Induced Inflammatory Responses and Lactation in Dairy Cow Mammary Epithelial Cells by Inhibiting NF-κB and MAPKs and Up-Regulating mTOR Signaling.
abstract: As a protective factor for lipopolysaccharide (LPS)-induced injury, 14-3-3γ has been the subject of recent research. Nevertheless, whether 14-3-3γ can regulate lactation in dairy cow mammary epithelial cells (DCMECs) induced by LPS remains unknown. Here, the anti-inflammatory effect and lactation regulating ability of 14-3-3γ in LPS-induced DCMECs are investigated for the first time, and the molecular mechanisms responsible for their effects are explored. The results of qRT-PCR showed that 14-3-3γ overexpression significantly inhibited the mRNA expression of tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), interleukin-1β (IL-1β) and inducible nitric oxide synthase (iNOS). Enzyme-linked immunosorbent assay (ELISA) analysis revealed that 14-3-3γ overexpression also suppressed the production of TNF-α and IL-6 in cell culture supernatants. Meanwhile, CASY-TT Analyser System showed that 14-3-3γ overexpression clearly increased the viability and proliferation of cells. The results of kit methods and western blot analysis showed that 14-3-3γ overexpression promoted the secretion of triglycerides and lactose and the synthesis of β-casein. Furthermore, the expression of genes relevant to nuclear factor-κB (NF-κB) and mitogen-activated protein kinase (MAPKs) and lactation-associated proteins were assessed by western blot, and the results suggested that 14-3-3γ overexpression inactivated the NF-κB and MAPK signaling pathways by down-regulating extracellular signal regulated protein kinase (ERK), p38 mitogen-activated protein kinase (p38MAPK) and inhibitor of NF-κB (IκB) phosphorylation levels, as well as by inhibiting NF-κB translocation. Meanwhile, 14-3-3γ overexpression enhanced the expression levels of β-casein, mammalian target of rapamycin (mTOR), ribosomal protein S6 kinase 1 (S6K1), serine/threonine protein kinase Akt 1 (AKT1), sterol regulatory element binding protein 1 (SREBP1) and peroxisome proliferator-activated receptor gamma (PPARγ). These results suggest that 14-3-3γ was able to attenuate the LPS-induced inflammatory responses and promote proliferation and lactation in LPS-induced DCMECs by inhibiting the activation of the NF-κB and MAPK signaling pathways and up-regulating mTOR signaling pathways to protect against LPS-induced injury.
authors.affiliation: Key Laboratory of Dairy Science of Education Ministry, Northeast Agricultural University, Harbin 150030, China. liulixin_2003@163.com.
authors.forename: Lixin
authors.initials: L
authors.lastname: Liu
authors.affiliation: Key Laboratory of Dairy Science of Education Ministry, Northeast Agricultural University, Harbin 150030, China. linlu516@163.com.
authors.forename: Ye
authors.initials: Y
authors.lastname: Lin
authors.affiliation: Key Laboratory of Dairy Science of Education Ministry, Northeast Agricultural University, Harbin 150030, China. liulili-liulili@163.com.
authors.forename: Lili
authors.initials: L
authors.lastname: Liu
authors.affiliation: Key Laboratory of Dairy Science of Education Ministry, Northeast Agricultural University, Harbin 150030, China. bianjie.033@163.com.
authors.forename: Yanjie
authors.initials: Y
authors.lastname: Bian
authors.affiliation: Key Laboratory of Dairy Science of Education Ministry, Northeast Agricultural University, Harbin 150030, China. zhangli_zl18800@163.com.
authors.forename: Li
authors.initials: L
authors.lastname: Zhang
authors.affiliation: Key Laboratory of Dairy Science of Education Ministry, Northeast Agricultural University, Harbin 150030, China. gxjgxj112233@163.com.
authors.forename: Xuejun
authors.initials: X
authors.lastname: Gao
authors.affiliation: Key Laboratory of Dairy Science of Education Ministry, Northeast Agricultural University, Harbin 150030, China. qzli@neau.edu.cn.
authors.forename: Qingzhang
authors.initials: Q
authors.lastname: Li
mesh_terms: D048948:14-3-3 Proteins / Q000235:genetics / Q000378:metabolism*; D000818:Animals; D002364:Caseins / Q000235:genetics / Q000378:metabolism*; D002417:Cattle; D002478:Cells, Cultured; D004847:Epithelial Cells / Q000378:metabolism*; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism; D005260:Female; D006801:Humans; D007378:Interleukins / Q000235:genetics / Q000378:metabolism; D007785:Lactose / Q000378:metabolism; D008070:Lipopolysaccharides / Q000494:pharmacology; D020935:MAP Kinase Signaling System; D008321:Mammary Glands, Animal / Q000166:cytology / Q000378:metabolism*; D016328:NF-kappa B / Q000378:metabolism*; D052247:Nitric Oxide Synthase Type II / Q000235:genetics / Q000378:metabolism; D058570:TOR Serine-Threonine Kinases / Q000378:metabolism*; D014409:Tumor Necrosis Factor-alpha / Q000235:genetics / Q000378:metabolism; D048051:p38 Mitogen-Activated Protein Kinases / Q000378:metabolism
chemical_list: D048948:14-3-3 Proteins; D002364:Caseins; D007378:Interleukins; D008070:Lipopolysaccharides; D016328:NF-kappa B; D014409:Tumor Necrosis Factor-alpha; D052247:Nitric Oxide Synthase Type II; D058570:TOR Serine-Threonine Kinases; D048049:Extracellular Signal-Regulated MAP Kinases; D048051:p38 Mitogen-Activated Protein Kinases; D007785:Lactose
keywords: 14-3-3γ; MAPKs; NF-κB; dairy cow mammary epithelial cells; inflammatory responses; mTOR signaling pathway