Pro-inflammatory cytokine TNF-alpha is a key inhibitory factor for lactose synthesis pathway in lactating mammary epithelial cells

Lactose is a milk-specific carbohydrate synthesized by mammary epithelial cells (MECs) in mammary glands during lactation. Lactose synthesis is downregulated under conditions causing inflammation such as mastitis, in which MECs are exposed to high concentrations of inflammatory cytokines. In this study, we investigated whether inflammatory cytokines (TNF-alpha, IL-1 beta, and IL-6) directly influence the lactose synthesis pathway by using two types of murine MEC culture models: the monolayer culture of MECs to induce lactogenesis; and the three-dimensional culture of MECs surrounded by Matrigel to induce reconstitution of the alveolar structure in vitro. TNF-alpha caused severe down-regulation of lactose synthesis related genes concurrently with the degradation of glucose transporter 1 (GLUTI) from the basolateral membranes in MECs. IL-1 beta also caused degradation of GLUTI along with a decrease in the expression level of beta-1,4-galactosylransferase 3. IL-6 caused both up-regulation and down-regulation of the expression levels of lactose synthesis-related genes in MECs. These results indicate that TNF-alpha, IL-1 beta, and IL-6 have different effects on the lactose synthesis pathway in MECs. Furthermore, TNF-alpha triggered activation of NFKB and inactivation of STAT5, suggesting that NF kappa B and STAT5 signaling pathways are involved in the multiple adverse effects of TNF-alpha on the lactose synthesis pathway. (C) 2015 Elsevier Inc. All rights reserved

Prolactin and glucocorticoid signaling induces lactation-specific tight junctions concurrent with beta-casein expression in mammary epithelial cells

Alveolar mammary epithelial cells (MECs) in mammary glands are highly specialized cells that produce milk for suckling infants. Alveolar MECs also form less permeable tight junctions (TJs) to prevent the leakage of milk components after parturition. In the formation process of less permeable TJs, MECs show a selective downregulation of Cldn4 and a localization change of Cldn3. To investigate what induces less permeable TJs through these compositional changes in Cldns, we focused on two lactogenesis-related hormones: prolactin (Prl) and glucocorticoids. Prl caused a downregulation of Cldn3 and Cldn4 with the formation of leaky TJs in MECs in vitro. Prl-treated MECs also showed low beta-casein expression with the activation of STAT5 signaling. By contrast, dexamethasone (Dex), a glucocorticoid analogue, upregulated Cldn3 and Cldn4, concurrent with the formation of less permeable TJs and the activation of glucocorticoid signaling without the expression of beta-casein. Cotreatment with Prl and Dex induced the selective downregulation of Cldn4 and the concentration of Cldn3 in the region of Tjs concurrent with less permeable TJ formation and high beta-casein expression. The inhibition of Prl secretion by bromocriptine in lactating mice induced the upregulation of Cldn3 and Cldn4 concurrent with the downregulation of milk production. These results indicate that the coactivation of Prl and glucocorticoid signaling induces lactation-specific less permeable TJs concurrent with lactogenesis

Underlying mechanisms involved in the decrease of milk secretion during Escherichia coli endotoxin induced mastitis in lactating mice

Mastitis, the inflammation of mammary glands resulting from bacterial infection, disrupts milk production in lactating mammary glands. In this study, we injected lipopolysaccharide (LPS), one of the endotoxins from Escherichia coli into mouse mammary glands to disrupt milk production, and we investigated the influence of LPS on nutrient uptake, synthesis, and secretion processes for milk component production in alveolar epithelial cells (AEC). The expression of genes relevant to the three-staged milk component production process (nutrient uptake, synthesis, and secretion of milk components) were down-regulated within 12 h after LPS injection in AEC. The internalization of glucose transporter 1 (GLUT-1) from the basolateral membrane to the cytoplasm occurred in accordance with the down-regulation of gene expression 3 h after LPS injection. The abnormal localization of adipophilin and beta-casein was also observed in the LPS-injected mammary glands. SLC7A1, an amino acid transporter, was up-regulated 3 and 6 h after LPS injection. Furthermore, the inactivation of signal transducer and activator of transcription 5 (STAT5) and the activation of STAT3 and nuclear factor-kappa B (NFkappaB) occurred 3 h after LPS injection. These results indicate that the nutrient uptake, synthesis, and secretion of milk components in AEC are rapidly shut down in the lactating mammary glands after LPS injection