The relationship between milk, mammary adipocytes and ECM in regulating murine mammary gland involution
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This thesis investigated the role of milk, extracellular matrix and mammary adipocytes in regulating mammary gland function during involution in mice and explored the use of an in vitro culture model, the mammosphere model system to study the same.Keywords:
Involution (esoterism)
Mammary tissue
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microRNAs(miRNAs)play a key role in cell proliferation,differentiation and apoptosis.miRNA involved in the regulation of mammary gland development in cell proliferation and differentiation.In this experiment,we used mice as an animal model to study the role of miR-138 in mouse mammary gland development and lactation.Maternal tail vein injection of miR-138 inhibitors,pups weighing method applied to detect the changes of milk yield;electron microscopy techniques was used to observe ultrastructure changes in mammary gland tissue.The results showed that after intravenous injection of miR-138 inhibitor,female mouse increased maternal mammary epithelial cells,increasing the amount of milk secretion,ultrastructure of cells found in the metabolic activity,collect female mouse milk after miR-138 inhibitor,detect lactose and casein of the milk were all increased.The results suggested that the inhibition of miR-138 could stimulate mammary epithelial cell proliferation,increase the secretion of milk,and regulation of the ingredient of the milk.
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Milk protein
Bovine milk
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The development and lactation of mammary gland is a complex process regulated by various growth factors and one of which is leptin. Besides in circulation, in many species, leptin is also produced and secreted by the mammary fat pad and the mammary epithelial cells. Leptin can influence the branches of ducts and the proliferation of the mammary epithelial cell in pregnancy. In lactation, it influences the expression of milk protein gene and delays the involution after the removing of the pups. Leptin act all those function through JAK/STAT or JAK/MAPK pathway. However, it is not very clear about these pathways in mammary epithelial cell and it remains to be tested. (The Journal of American Science. 2005;1(1):63-67).
Involution (esoterism)
Mammary tissue
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Organ culture
Polyamine
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Regulation of milk synthesis and secretion is controlled mostly through local (intramammary) mechanisms. To gain insight into the molecular pathways comprising this response, an analysis of mammary gene expression was conducted in 12 lactating cows shifted from twice daily to once daily milking. Tissues were sampled by biopsy from adjacent mammary quarters of these animals during the two milking frequencies, allowing changes in gene expression to be assessed within each animal. Using bovine-specific, oligonucleotide arrays representing 21,495 unique transcripts, a range of differentially expressed genes were found as a result of less frequent milk removal, constituting transcripts and pathways related to apoptotic signaling (NF-kappaB, JUN, ATF3, IGFBP5, TNFSF12A) mechanical stress and epithelial tight junction synthesis (CYR61, CTGF, THBS1, CLDN4, CLDN8), and downregulated milk synthesis (LALBA, B4GALT1, UGP2, CSN2, GPAM, LPL). Quantitative real-time PCR was used to assess the expression of 13 genes in the study, and all 13 of these were correlated (P < 0.05) with values derived from array analysis. It can be concluded that the physiological changes that occur in the bovine mammary gland as a result of reduced milk removal frequency likely comprise the earliest stages of the involution response and that mechano-signal transduction cascades associated with udder distension may play a role in triggering these events.
Udder
Milking
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In contrast to the exstensive research done on milk synthesis in animal models, there are only few studies on human milk synthesis. Using human mammary epithelial cells (HMEC) from mammoplasty specimens, we first studied different aspects of growth and differentiation with the aim of creating a cell culture model for studies on human milk synthesis. The cells, which are commercially available (Mammary Pack®, Clonetics Corp.), were cultured in serum-free media on different substrates. By culturing the cells on top of a reconstituted basal membrane matrix (Matrigel®, Beckton Dickinson Labware) a morphological differentiation was achieved with development of alveolar and duct-like structures. We have also studied lipid synthesis using a radioactive fatty acid precursor (3H-acetate) while stimulating the cells with different lactogenic hormones. Acetate was incorporated into lipids at a rate of 0,8-1,3 nmoles/million cells/hour in media without epidermal growth factor (EGF), insulin (I), hydrocortison (HC) and prolactin-rich bovine pituitary extract (BPE). This incorporation rate is similar to the rate reported in a previous study on HMEC from human breast milk. The incorporation rate was highest during the first 2 hours of incubation. Addition of EGF, I, HC and BPE caused an increase in the acetate incorporation rate to 2,4-3,4 nmoles/million cells/hour. Most of this increase was caused by EGF and I, while HC and BPE seemed to have minor effects, In some cases even inhibitory. The composition of the synthesised lipids was studied by thin layer chromatography and subsequent radiomaging, showing that there was a mixture dominated by triglycerides and phospholipids (preliminary results). Gas chromatography of lipid extracts from the cells showed that the chain lenghts of the main fatty acids were 18 and 16 carbon atoms. We conclude that this system could be a useful cell culture model for studying human milk fat synthesis.
Matrigel
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