Anti-inflammatory Activity And Regulation Mechanisms Of Taraxasterol On Mammitis Induced By LPS

Bovine mastitis is one of the most important diseases of dairy cows that oftencauses severe economic lost. The incidence of mastitis in dairy cows was20%-70%,or even higher incidence in individual herds. Induced mastitis in dairy cows occur fora variety of causes, Staphylococcus aureus, Escherichia coli and Streptococcus. Themain mechanism of pathopoiesia of E.coli showed that the main causes of its cell walllipopolysaccharide component constituting the surface.Generally, Antibiotics play an important role in the process of prevention and treatment of severe mastitis. However, due to long time use of antibiotic drug, it alwayswas often accompanied by problems, lead to human drug tolerance, and harmful to human health. Recently, in view of people’s understanding to connotation of health andpursuit of organic milk, the application of antibiotics in mastitis gradually being restricted or disabled. Therefore, it forced people to look for safer alternatives to antibioticsto control mastitis in dairy cows. As we all know, Chinese medicine was used for preventing and curing diseases for thousands of years. Among many Chinese medicines for breast disease, Mongolian Dandelion is often used as herbal prescriptions involvedin mastitis treatment. Taraxasterol, one of the important active ingredients, has been shown to have anti-inflammatory, anti-bacterial, and anti-tumor effect. It draws increasing attention of researchers.Taraxasterol, a pentacyclic-triterpene isolated from Taraxacum officinale, hasbeen used in this study. Firstly, we detected the anti-inflammatory effects oftaraxasterol on LPS-induced acute lung injury in mice. Secondly, we investigated theanti-inflammatory effects of taraxasterol on LPS-induced mastitis. The results showedthat taraxasterol exhibited anti-inflammatory effects on LPS-induced mastitis throughinhibiting mammary tissue MPO acitivity, the production of inflammatory cytokines,and TLR4-mediated NF-κB signaling pathway. To further investigate theanti-inflammatory effects of taraxasterol, LPS-stimulated mice mammary epithelialcells were used in this study. We found that taraxasterol inhibited LPS-inducedinflammatory cytokines, as well as NF-κB and MAPKs signaling pathway. Finally, weinvestigated the anti-inflammatory effects of taraxasterol on LPS-stimulated bovine mammary epithelial cells. Our results showed that taraxasterol exhibitedanti-inflammatory effects by inhibiting NF-κB activation and inflammatory cytokinesexpression.In this study, we firstly detected the anti-inflammatory effects of taraxasterol inLPS-induced acute lung injury. Mice were randomly divided into seven groups:control group, taraxasterol (10mg/kg) group, LPS group, taraxasterol (2.5mg/kg,5mg/kg, and10mg/kg)+LPS group, and dexamethasone (DEX)+LPS group. MaleBALB/c mice were pretreated with taraxasterol1h before intranasal instillation ofLPS.7h after LPS administration, the myeloperoxidase (MPO) in lung tissues, lungwet/dry ratio and inflammatory cells in the bronchoalveolar lavage fluid (BALF) weredetected. The levels of tumor necrosis factor-α (TNF-α), interleukin-6(IL-6),interleukin-1β (IL-1β) in the BALF were measured by ELISA. The results showedthat taraxasterol attenuated the infiltration of inflammatory cells, the activity ofmyeloperoxidase (MPO), lung wet/dry ratio, and the expression of tumor necrosisfactor-α (TNF-α), interleukin-6(IL-6) and interleukin-1β (IL-1β). These resultssuggested that taraxasterol had an anti-inflammatory effect on LPS-induced ALI.Secondly, we detected the anti-inflammatory effects of taraxasterol onLPS-induced mastitis in mice. Mice were randomly divided into six groups: controlgroup, taraxasterol (10mg/kg) group, LPS group, taraxasterol (2.5mg/kg,5mg/kg,and10mg/kg)+LPS group, and dexamethasone (DEX)+LPS group. Mice werepretreated with taraxasterol or DEX1h before instillation of LPS. The quantity anddistribution of MPO were detected by immunohistochemical examination. Theproduction of inflammatory cytokines TNF-α, IL-1β, and IL-6were detected byELISA. The activation of NF-κB was detected by western blotting. Mammary tissuesshowed inflammatory cells infiltration, mammary gland alveolus congestion andmarked thickening of the alveolus walls after LPS instillation. Taraxasterol couldattenuate the injury and MPO activity of mammary tissues induced by LPS.Meanwhile, taraxasterol inhibited LPS-induced NF-κB activation and inflammatorycytokines TNF-α, IL-1β, and IL-6production. These results showed that taraxasterolinhibited LPS-induced inflammatory response through inhibiting NF-κB activation.Thirdly, we detected the anti-inflammatory effects of taraxasterol onLPS-stimulated mice mammary epithelial cells. The effect of taraxasterol on cellviability was evaluated using the MTT assay. The effects of taraxasterol onLPS-induced inflammatory cytokines TNF-α, and IL-6production were detected by ELISA. The effects of taraxasterol on LPS-induced NF-κB and MAPKs activationwere detected by western blotting. We found that taraxasterol (2.5μg/mL,5μg/mL,10μg/mL) inhibited LPS-induced TNF-α, and IL-6production. Meanwhile,taraxasterol suppressed LPS-induced NF-κB and MAPKs activation in adose-dependent manner.In the last, the anti-inflammatory effects and mechanism of taraxasterol weredetected in LPS-stimulated bovine mammary epithelial cells. The effect oftaraxasterol on the viability of bovine mammary epithelial cell was evaluated usingthe MTT assay. The effects of taraxasterol on NF-κB activation was detected byWestern blotting. The effects of taraxasterol on LPS-induced inflammatory cytokinesexpression were detected by qRT-PCR. The results showed that the cell viability wasnot affected by taraxasterol at the dose of2.5μg/mL,5μg/mL,10μg/mL. Meanwhile,taraxasterol inhibited LPS-induced TNF-α, IL-1β, and IL-6expression. Western blotanalysis showed that taraxasterol suppressed LPS-induced NF-κB activation in adose-dependent manner.In this study, we firstly established LPS-induced acute lung injury model in miceby intranasal instillation of LPS. Secondly, we established LPS-induced mastitis inmice. Thirdly, LPS-stimulated mouse mammary epithelial cells were used toinvestigate the anti-inflammatory effects and mechanism of taraxasterol. In the last,the anti-inflammatory effects and mechanism of taraxasterol were investigated inLPS-stimulated bovine mammary epithelial cells. Our results showed that taraxasterolinhibited LPS-induced NF-κB and MAPKs activation in mice mammary gland tissues.Meanwhile,taraxasterol suppressed LPS-induced NF-κB、p38、ERK、JNK activationin mouse mammary epithelial cells. Furthermore, taraxasterol inhibited LPS-indcuedNF-κB activation and inflammatory cytokines TNF-α、IL-1β, and IL-6expression.This paper investigates the anti-inflammatory mechanism of taraxasterol ininflammatory signaling pathways. It provides a basis for reference to the study of drugprevention and treatment of mastitis in dairy cows.