Isobutyrylshikonin inhibits lipopolysaccharide-induced nitric oxide and prostaglandin E2 production in BV2 microglial cells by suppressing the PI3K/Akt-mediated nuclear transcription factor-κB pathway

Microglia are important macrophages to defend against pathogens in the central nervous system (CNS); however, persistent or acute inflammation of microglia lead to CNS disorders via neuronal cell death. Therefore, we theorized that a good strategy for the treatment of CNS disorders would be to target inflammatory mediators from microglia in disease. Consequently, we investigated whether isobutyrylshikonin (IBS) attenuates the production of proinflammatory mediators, such as nitric oxide (NO) and prostaglandin E-2, in lipopolysaccharide (LPS)-stimulated BV2 microglial cells. Treatment with IBS inhibited the secretion of NO and prostaglandin E-2 (as well as the expression of their key regulatory genes), inducible NO synthase (iNOS), and cyclooxygenase-2 (COX-2). Isobutyrylshikonin also suppressed LPS-induced DNA-binding activity of nuclear transcription factor-kappa B (NF-kappa B), by inhibiting the nuclear translocation of p50 and p65 in addition to blocking the phosphorylation and degradation of I kappa B alpha. Pretreatment with pyrrolidine dithiocarbamate, a specific NF-kappa B inhibitor, showed the down-regulation of LPS-induced iNOS and COX-2 messenger RNA by suppressing NF-kappa B activity. This indirectly suggests that IBS-mediated NF-kappa B inhibition is the main signaling pathway involved in the inhibition of iNOS and COX-2 expression. In addition, IBS attenuated LPS-induced phosphorylation of PI3K and Akt, which are upstream molecules of NF-kappa B, in LPS-stimulated BV2 microglial cells. The functional aspects of the PI3K/Akt signaling pathway were analyzed with LY294002, which is a specific PI3K/Akt inhibitor that attenuated LPS-induced iNOS and COX-2 expression by suppressing NF-kappa B activity. These data suggest that an IBS-mediated anti-inflammatory effect may be involved in suppressing the PI3K/Akt-mediated NF-kappa B signaling pathway. (C) 2014 Elsevier Inc. All rights reserved.