| In this study, the LPS induced mouse model of mastitis was established and evaluated. Invivo, the protective effect and mechanism of emodin on LPS induced mouse mastitis was studied.And in vitro, the mouse mammary epithelial cell was cultured. The protective effect andmechanism of emodin on LPS induced mouse mammary epithelial cell was also studied, and theresults were in concert with which in vivo. All the results will provide the reference andexperimental basis for prevention and treatment of bovine mastitis by Chinese medicine inveterinary clinical practice.Firstly, the LPS induced mouse mastitis model was established. The lactating mice of5-7dwere random divided into two groups, including black control group and LPS group. Each groupcontains6mice. The fourth pair mammary gland of the mice in LPS group were infusion withLPS (0.2mg/ml,50μL) through the mammary duct. All the mice were killed24h after LPSinduction. The pathological and histopathological changes of mice mammary gland were observed,the distribution and activity of MPO were detected, the level of TNF-α was measured by ELISA.The results suggested that, compared with the black control group, the redness, swelling, andcongestion could be seen in the LPS induce mammary gland, the thickening and edema of thealveolus walls and inflammatory cells infiltration could be seen in the the histopathological sectionof the LPS induced mice mammary, and the level of TNF-α was significantly increased. Theseresults illustrated that the mouse mastitis model induced by LPS was successfully established.Secondly, the protective effect and mechanism of emodin on LPS induced mouse mastitiswas evaluated. The lactating mice were random divided into6groups, including black controlgroup, LPS group, LPS+emodin groups with the dose1,2, and4mg/kg respectively, LPS+DEXgroup with dose of5mg/kg DEX. Each group contained12mice. In the LPS+emodin andLPS+DEX groups, emodin and DEX were administer through intraperitoneal injection1h beforeand12h after LPS instillation respectively. All the mice were kill24h after infusion of LPS. Thehistopathological changes and the distribution and activity of MPO were observed, the levels ofTNF-α, IL-1β, and IL-6were measured by ELISA, the activation of NF-κB and MAPKs signal pathways were analyzed by western blot. The result suggested that emodin could significantlyreduce the histopathological changes and the distribution and activity of MPO, decline the proteinand mRNA expression of TNF-α, IL-1β, and IL-6, inhibit the activation of NF-κB and MAPKssignal pathways. This study confirmed the protective effect and mechanism of emodin on LPSinduced mouse mastitis in vivo.Finally, to further explain the protective effect of emodin on mastitis, the mouse mammaryepithelial cell was cultured by the digestion method of CollagenaseⅠ, CollagenaseⅡ andtrypsogen. The immunofluorescence staining of keratinose-18was used to identify the purity ofmouse mammary epithelial cell. The protective effect and mechanism of emodin on LPS inducedmouse mammary epithelial cell were studied. The cytotoxicity of emodin on mouse mammaryepithelial cell were measured by MTT, the level of TNF-α, IL-1β, and IL-6were measured byELISA, and the activation of NF-κB and MAPKs signal pathways were analyzed by western blot.The results showed that there is no cytotoxicity of emodin with the dose of10,20,40μg/ml, andemodin could decrease the level of TNF-α and IL-6, the result of western blot suggest that emodincould exert the anti-inflammatory effect on LPS induced mouse mammary epithelial cell byinhibiting the activation of NF-κB and MAPKs signal pathways.In this paper, all the results show that emodin may protect the mammary gland throughinhibiting the activation of NF-κB and MAPKs signal pathways. Thereby, the level ofpro-inflammatory cytokines, such as TNF-α and IL-6were decreased, the inflammatory cell werealso declined in the mammary gland. All the results revealed that emodin may be used as apotential candidate in treatment of mastitis. |
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