Abstract
Microglial cells are the resident macrophages and intrinsic arm of the central nervous system innate immune defense. Microglial cells become activated in response to injury, infection, environmental toxins, and other stimuli that threaten neuronal survival. Therefore, regulating microglial activation may have therapeutic benefits that lead to alleviating the progression of inflammatory-mediated neurodegeneration. In the present study, we investigated the effect of glaucocalyxin A (GLA) isolated from Rabdosia japonica on the production of pro-inflammatory mediators in lipopolysaccharide (LPS)-stimulated primary microglia and BV-2 cells. GLA significantly inhibited LPS-induced production of nitric oxide and reversed the morphological changes in primary microglia. Further, GLA suppressed expression of inducible nitric oxide synthase and cyclooxygenase-2 dose-dependently at the mRNA and protein levels. The production of proinflammatory cytokines such as tumor necrosis factor-α, interleukin-1β (IL)-1β, and IL-6 were inhibited by suppressing their transcriptional activity. Furthermore, GLA suppressed nuclear factor-κB activation by blocking degradation of IκB-α and inhibited the induction of lipocalin-2 expression in LPS-stimulated BV-2 cells. Mechanistic study revealed that the inhibitory effects of GLA were accompanied by blocking the p38 mitogen activated protein kinase signaling pathway in activated microglia. In conclusion, given that microglial activation contributes to the pathogenesis of neurodegenerative diseases, GLA could be developed as a potential therapeutic agent for treating microglia-mediated neuroinflammatory diseases.
Highlights
Dysregulation of the neuroimmune system has been suggested to play a decisive role in the pathogenesis of several neurodegenerative disorders
Nuclear extract kit was purchased from Thermo Scientific (Rockford, IL, USA), prostaglandin E2 (PGE2) enzyme-linked immunoassay (EIA) kit was obtained from Cayman Chemical Co. (Ann Arbor, MI, USA), and nitric oxide synthase (NOS) 2 and inducible NOS 2 EIA kits were purchased from Uscn Life Science Inc. (Wuhan, China)
LPS-induced NO production decreased prominently when glaucocalyxin A (GLA) was co-treated with pyrrolidine dithiocarbamate (PDTC) (p,0.001, Fig. 7D). These results indicate that GLA suppressed the production of NO, Inducible nitric oxide synthase (iNOS), COX-2 and pro-inflammatory cytokines (TNF-a, IL-1b, and IL-6) and this inhibition might be through regulation of nuclear factor (NF)-kB activation and degradation of IkB-a in LPS-stimulated BV-2 cells
Summary
Dysregulation of the neuroimmune system has been suggested to play a decisive role in the pathogenesis of several neurodegenerative disorders. The resident immune cells of the central nervous system, become activated and induce significant and highly detrimental neurotoxic effects by excessively producing a large array of cytotoxic and proinflammatory factors [1,2,3]. Lipopolysaccharide (LPS) is a common toxin used to investigate the impact of inflammation on neuronal death, and microglial cells are necessary for LPS-induced neurotoxicity [6]. BV-2 microglial cells respond to the LPS endotoxin by synthesizing inflammatory factors such as nitric oxide (NO), cyclooxygenase-2 (COX-2), tumor necrosis factoralpha (TNF-a), interleukin (IL)-1b, and IL-6. These microglial products are thought to be responsible for neuroglia-mediated neurotoxicity [7]
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