Mastitis is an inflammatory breast disease that develops mostly during the first 12 weeks’ post-partum in women and it is the major reason for early cessation of breastfeeding. The mechanisms of human mastitis are mostly unknown, while many studies have been published in bovine, showing a chronic inflammatory response in mammary tissue with high and extended production of pro-inflammatory cytokines (IL-1B, IL-6, IL-8, TNF-), known to be mediated via increases in the activity of NF-B. During chronic inflammation these cytokines, reactive oxygen species (ROS), and phagocytes can cause considerable damage to the mammary tissue and hence, decrease in milk production and cessation of breastfeeding. Interestingly, numerous previous studies have reported the beneficial and protective anti-inflammatory properties mediated by the activation of the cannabinoid type 2 (CB2) receptor with endogenous cannabinoids, phytocannabinoids, and synthetic agonists in human models of chronic inflammatory diseases. Furthermore, CB2 expression is found upregulated in these tissues undergoing chronic inflammatory responses. Activation of CB2 with these agonists has been previously shown to causes a decrease in pro-inflammatory cytokines (IL-1B, IL-6, IL-8, TNF-) and ROS production also mediated by a downregulation of NF-B. Indeed, downregulation of oxidative enzymes (iNOS) and upregulation of anti-oxidant enzymes (SOD) is also observed. These are the same components and signaling pathways known to be chronically upregulated during mastitis inflammation. The overall goal of this project is to provide evidence of the potential role of CB2 as a therapeutic target in the modulation of mammary tissue inflammation during mastitis infection. The endogenous cannabinoid and CB2 agonist 2-AG has been detected in human milk during lactation, providing evidence this organ may be under control of the endocannabinoid system. Our main hypothesis is that CB2 expression in the mammary gland is elevated during mastitis, and its anti-inflammatory properties can be activated by endogenous and synthetic agonists in vitro. This hypothesis will be tested through three specific aims in bovine mammary tissue: 1) to characterize the gene and/or protein expression profile of the CB2 receptor, documented molecules related to CB2 immunomodulation (NF-K, COX2, iNOS), and inflammatory cytokines (IL-1, IL-6, IL-8, TNF-) in healthy and mastitis bovine mammary tissue (mixed cell types) during lactation, 2) to test the anti-inflammatory effect of CB2 receptor activation via endogenous and synthetic agonists (2-AG and JWH133) in vitro using bovine primary mammary epithelial cell culture by monitoring changes in expression of these aforementioned molecules, and 3) characterize host response to CB2 activation during infection with S. aureus compared to E. Coli by monitoring changes in expression of these aforementioned molecules These studies are expected to significantly advance current understanding ...