Cardamonin from Alpinia rafflesiana inhibits inflammatory responses in IFN-γ/LPS-stimulated BV2 microglia via NF-κB signalling pathway

The increasing prevalence of neurodegenerative diseases has prompted investigation into innovative therapeutics over the last two decades. Non-steroidal anti-inflammatory drugs (NSAIDs) are among the therapeutic choices to control and suppress the symptoms of neurodegenerative diseases. However, NSAIDs-associated gastropathy has hampered their long term usage despite their clinical advancement. On the natural end of the treatment spectrum, our group has shown that cardamonin (2′,4′-dihydroxy-6′-methoxychalcone) isolated from Alpinia rafflesiana exerts potential anti-inflammatory activity in activated macrophages. Therefore, we further explored the anti-inflammatory property of cardamonin as well as its underlying mechanism of action in IFN-γ/LPS-stimulated microglial cells. In this investigation, cardamonin shows promising anti-inflammatory activity in microglial cell line BV2 by inhibiting the secretion of pro-inflammatory mediators including nitric oxide (NO), prostaglandin E2 (PGE2), tumour necrosis factor-α (TNF-α), interleukin-1β (IL-1β) and interleukin-6 (IL-6). The inhibition of NO and PGE2 by cardamonin are resulted from the reduced expression of inducible nitric oxide synthase (iNOS) and cycloxygenase-2 (COX-2), respectively. Meanwhile the suppressive effects of cardamonin on TNF-α, IL-1β and IL-6 were demonstrated at both protein and mRNA levels, thus indicating the interference of upstream signal transduction pathway. Our results also validate that cardamonin interrupts nuclear factor-kappa B (NF-κB) signalling pathway via attenuation of NF-κB DNA binding activity. Interestingly, cardamonin also showed a consistent suppressive effect on the cell surface expression of CD14. Taken together, our experimental data provide mechanistic insights for the anti-inflammatory actions of cardamonin in BV2 and thus suggest a possible therapeutic application of cardamonin for targeting neuroinflammatory disorders.

Graphical abstractFigure above summarizes that cardamonin from Alpinia rafflesiana inhibits protein expression of iNOS and COX-2 as well as gene expression of TNF-α, IL-1β and IL-6 by inhibiting NF-κB DNA binding activity and CD14 cell surface expression in IFN-γ/LPS-stimulated BV2 cells. It also depicts the possible overall pathway in which cardamonin could have exhibited its anti-inflammatory effects.LPS, lipopolysaccharide; CD14, cluster of differentiation 14; TLR4, toll-like receptor 4; MD2, Lymphocyte antigen 96; MyD88, Myeloid differentiation primary response gene (88); IRAK, interleukin-1 receptor-associated kinase; TRAF-6, tumour necrosis factor receptor-associated factor 6; IĸB, I-kappa-B; P, phosphorylation; NF-κB, nuclear factor kappa-B, iNOS, inducible nitric oxide synthase; COX-2, cyclooxygenase; TNF-α, tumour necrosis factor-alpha, IL-1β, interleukin-beta; IL-6, interleukin-6.Download full-size imageHighlights► Cardamonin inhibits NO production by supressing iNOS enzyme expression in stimulated mouse microglial cells (BV2). ► Cardamonin inhibits PGE2 production by selectively suppressing COX-2 protein expression in stimulated BV2 cells. ► Cardamonin down-regulated cytokines (TNF-α, IL-1β and IL-6) gene expression in stimulated BV2 cells. ► Cardamonin inhibited NF-κB DNA binding. ► Cardamonin suppressed CD14 surface expression.