Effect Of Osthole On Activation Of Microglial Cells Induced By Lipopolysaccharides

Objective:In this study, we investigated the inhibitory effects of osthole (OST) on activation in BV-2 microglial cells induced by lipopolysaccharides (LPS), and explored the possible mechanisms.Methods:BV-2 microglia cells were stimulated by LPS as the inflammation modle. Nittic oxide assay kit and typing nitric oxide synthase assay kit were performed to establish the effects of osthole on production of NO and iNOS activity in LPS-stimulated BV-2 microglia cells. ELISA was performed to establish the effects of osthole on IL-1 and PGE2 production. The inhibitory effects of osthole on the expression of iNOS, COX-2 and TLR4 were detected by Western blot in LPS-stimulated BV-2 microglia cells, and miR-155 level was detected by quantitative PCR (qPCR).Results:1. LPS (0.02-2.5μg/mL) showed no significant decrease in cell viability compared to control group. NO, IL-1βand PGE2 levels increase in a time-dependent manner in 0.lμg/mL LPS-stimulated BV-2 microglia cells as well as iNOS, COX-2, TLR4 protein and miRNA155 expression.2. MTT assay was performed to determine the viability of BV-2 microglia cells after treatment with various osthole concentrations for 24h. Osthole (0.3-27μg/mL, 1.7-135μM) showed no significant decrease in cell viability compared to control group.3. Osthole (1-9μg/mL,5-45μM) significantly suppressed the production of NO, iNOS activity and iNOS protein expression in LPS-stimulated BV-2 microglia cells in a dose dependent manner.4. Osthole (1-9μg/mL,5-45μM) significantly decrease the production of PGE2 and COX-2 protein expression in LPS-stimulated BV-2 microglia cells in a dose dependent manner. 5. Osthole (1-9μg/mL,5-45μM) significantly suppressed the IL-1βexpression in LPS-stimulated BV-2 microglia cells in a dose dependent manner.6. Osthole (1-9μg/mL,5-45μM) significantly decrease TLR4 protein expression in LPS-stimulated BV-2 microglia cells in a dose dependent manner.7. Osthole (1-9μg/mL,5-45μM) significantly decrease miR-155 expression in LPS-stimulated BV-2 microglia cells in a dose dependent manner.Conclusion:Osthole (1-9μg/mL,5-45μM) significantly suppressed the production of inflammatory mediatoes such as NO, PGE2, IL-1βin LPS-stimulated BV-2 microglia cells in a dose dependent manner. We hypothesize that this is achieved by suppressing the level of iNOS, COX-2, TLR4 and miR-155 in LPS-stimulated BV-2 microglia cells.