We aimed to analyze transcriptional changes of mammary epithelialcells (MEC) isolated from bovine mammary glands after intramammarychallenge with lipopolysaccharide (LPS). Ten multiparous cowswere used in the study. Five treatment (T) and 5 control (C) cows werepaired based on days in milk, milk yield and parity. For T cows, bothmammary glands on one side of the udder were infused with LPS (50μg in 10 mL saline); these glands were designated (TL). The contralateralglands received 10 mL saline and were designated (TS). Likewise,for C cows 2 ipsilateral glands received saline (CS) and the contralateralglands remained uninfused (CU). Mammary tissues were biopsiedbefore (0 h) and at 3 and 12 h post-infusion and processed for laser capturemicrodissection (LCM). MEC were collected using LCM, total RNAwas isolated and subjected to low-put RNA sequencing. Among variouscomparisons, we found 3167 (TL3 vs. TL0), 670 (TL12 vs. TL0), 2555(TL3 vs. TS3), and 3823 (TL3 vs. CS3) differentially expressed genes[DEGs; FDR < 0.05, Log2 (fold change) ≥ 1]. The major local responsesof MEC in TL glands at 3h included upregulation of ribosome biogenesis,innate immunity and KEGG pathways of TNF, NOD-like receptorand NFKB signaling. Ingenuity pathway analysis showed activation ofTNFR2, PI3/AKT, iNOS and acute phase protein response. Upstream regulatorsof these pathways predicted invasion of cell, chemotaxis and cellmigration, and showed activated HIF1A network. Downregulated genesincluded network of carbohydrate metabolism, PPAR fatty acid biosynthesis,and several ionic transporters. Major systemic responses of MECin TS glands showed weak cell mediated immune response, lymphocyteactivation, and cytokine production responses. Ingenuity pathway analysisof systemic response genes at 3 h showed p53 senescence pathwaywith activated upstream regulators as NFKB and TNFA. These results ofcomprehensive transcriptome profiling of MEC may explain gene regulationof local and systemic responses of MEC during E. coli mastitis.
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