The Protective Effect Of Geniposide On LPS Induced Inflammatory Injury Of Mouse Mammary Gland

Geniposide is one of the active ingredients of traditional Chinese medicine named gardenia,the study found that it has significant anti-inflammatory and anti-microbial effect. In order tostudy the clinical medicinal value of geniposide, we select geniposide in the experiments on LPSinduced mouse mastitis model in depth study. In vivo, the experiments use lipopolysaccharide(LPS) stimulation to build mice mouse model, and then study the protective effect of geniposideon LPS-induced mouse mastitis. In vitro, separated and cultured mouse mammary epithelial cells,study the protective effect of geniposide on LPS-induced mouse mammary epithelial cellinflammatory injury, and explored its mechanism in NF-κB signaling pathway. This study providea theoretical basis and reference for the natural ingredient in traditional Chinese medicine used tocow mastitis prevention and treatment.Firstly, pour LPS into catheter of mice nipple to build the mice mouse model. Female micewhich farrowed five days were randomly divided into six groups (six each group): normal group;LPS group; geniposide low dose group (25mg/kg); geniposide dose group (50mg/kg); geniposidehigh dose group (100mg/kg); dexamethasone group (5mg/kg), then administered according to theexperimental design. We killed the mice, dissected mice mammary tissue and maked tissuehistopathology and immunohistochemistry sections, observed the MPO activity changes inorganization and other pathological changes. By detected the expression level of TNF-α, IL-6andIL-1β in mammary tissue of each group. The results show that mammary tissue have red, swollenand other inflammatory changes after LPS stimulated, we successfully builded the mice mousemodel. Histopathology and immunohistochemistry assay showed that geniposide significantlyreduce the LPS-induced mouse mastitis symptoms and with dose-dependent. ELISA test resultsshowed geniposide can significantly inhibit the expression level of TNF-α, IL-6and IL-1β inLPS-induced mammary tissue and with dose-dependent.Secondly, separated and cultured mouse mammary epithelial cells, identified bymorphological and biological methods to make sure the model of mammary epithelial cells ininflammatory injury are successful. Evaluated the toxic effects of geniposide add to mouse mammary epithelial cells by MTT assay. The cells cultured were randomly divided into5groups:normal group; LPS group; LPS+geniposide group (50mg/kg); LPS+geniposide group (100mg/kg);geniposide group (200mg/kg). LPS stimulated and given geniposide by the experimental design,we used the ELISA method to detect the expression level of cytokines as TNF-α, IL-6and IL-1βof mouse mammary epithelial cells, and used RT-PCR method to detect the mRNA expressionlevel of TNF-α, IL-6and IL-1β of mouse mammary epithelial cells. The results show wesuccessfully separated and cultured mouse mammary epithelial cells and cell purity meets the testrequirements. the concentration of geniposide and LPS we added in experiments have no toxiceffects on mouse mammary epithelial cells. Geniposide inhibited the expression level of TNF-α,IL-6and IL-1β in LPS-induced mouse mammary epithelial cells, it as same as the mRNAexpressed.Finally, The mouse mammary epithelial cells density of4×105cells per ml was seeded in6-well cell culture plate, and randomly divided into5groups: normal group; LPS group;LPS+geniposide group (50mg/kg); LPS+geniposide group (100mg/kg); geniposide group(200mg/kg), LPS stimulated and given geniposide by the experimental design, and then collectedprotein for western blot assay. The results of experiment show that: LPS stimulation of mousemammary epithelial cells can significantly lead to IκB and NF-κB p65phosphorylated;Geniposide at concentrations of50μg/mL,100μg/mL,200μg/mL can inhibit LPS inducedphosphorylation, Inhibited the activation of NF-κB signaling pathway with dose-dependent.The above study show geniposide play a excellent effect on protect LPS-induced injury inmice mastitis. The anti-inflammatory mechanism of geniposide may be through inhibitedactivation of NF-κB signaling pathway, then reduced the expression of TNF-α, IL-6and IL-1β andinflammatory injury in mammary tissue. The study suggested geniposide may as a potential drugon treatment of mastitis.