Abstract
Prolonged inflammatory responses can lead to the development of several chronic diseases, such as autoimmune disorders and the development of natural therapeutic agents is required. A murine model was used to assess the anti-inflammatory effects of the megastigmane glucoside, icariside B2 (ICSB), and the assessment was carried out in vitro, and in vivo. The in vitro anti-inflammatory effects of ICSB were tested using LPS-stimulated BV2 cells, and the protein expression levels of inflammatory genes and cytokines were assessed. Mice were subcutaneously injected with 1% carrageenan (CA) to induce acute phase inflammation in the paw. Inflammation was assessed by measuring paw volumes hourly; subsequently, the mice were euthanized and the right hind paw skin was expunged and processed for reverse transcription-polymerase chain reaction (RT-PCR) and Western blot analyses. ICSB inhibits LPS-stimulated nitric oxide (NO) and prostaglandin E2 (PGE2) generation by reducing the expression of inducible NO synthase (iNOS) and cyclooxygenase 2 (COX-2). ICSB also inhibits the COX-2 enzyme with an IC50 value of 7.80 ± 0.26 µM. Molecular docking analysis revealed that ICSB had a strong binding affinity with both murine and human COX-2 proteins with binding energies of −8 kcal/mol and −7.4 kcal/mol, respectively. ICSB also reduces the manifestation of pro-inflammatory cytokines, such as TNF-α, IL-6, and IL-1β, at their transcriptional and translational level. ICSB hinders inhibitory protein κBα (IκBα) phosphorylation, thereby terminating the nuclear factor kappa-light-chain-enhancer of activated B cell (NF-κB) nuclear translocation. ICSB also represses the mitogen-activated protein kinases (MAPKs) signaling pathways. ICSB (50 mg/kg) showed an anti-edema effect in CA-induced mice and suppressed the CA-induced increases in iNOS and COX-2 protein levels. ICSB attenuated inflammatory responses by downregulating NF-κB expression through interference with extracellular signal-regulated kinase (ERK) and p38 phosphorylation, and by modulating the expression levels of iNOS, COX-2, TNF-α, IL-1β, and IL-6.
Highlights
Microglia are specialized macrophages of the central nervous system (CNS) that play a vital role in both CNS homeostasis and immune defense
FiguFriegu2rAe 2sAhsohwows sththaatttthhee eexxppeerirmimenetnaltadlosdeossoefsICoSfBIC(2S5,B50(,2a5n,d5100,0aμnMd)1d0i0d nμoMt h)adveidannyottoxhiacve any toxiceeffffecetcot nonBVB2Vc2elclse,lwls,hewrehaesrIeCaSsBICatS2B00atμ2M00shμoMwesdhcoewllueldarcteolxliuclitayr. tAosxischiotyw.nAins Fshigouwren2Bin, LFPigSure 2B, LPS ttrreeaattmmeennttssigignnifiificacnatnlytlyinicnrecareseadsetdhethceellcuellalrulNarONgOenegreantieornat(icoonlu(mconlu3)mcnom3)pcaoremdpwairtehduwntirtehatuendtreated cells, whereas icariside B2 (ICSB) treatment significantly reduced nitric oxide (NO) generation in a dose-dependent manner
These results demonstrate that ICSB might trigger inducible NO synthase (iNOS) and cyclooxygenase 2 (COX-2) inhibition, resulting in a decrease in NO and prostaglandin E2 (PGE2) production in LPS-stimulated microglial cells
Summary
Microglia are specialized macrophages of the central nervous system (CNS) that play a vital role in both CNS homeostasis and immune defense. The development of paw edema was maximal at approximately 4–5 h post-CA injection and is viewed as an immediate inflammatory response, which is decreased by the action of inhibitors within the inflammatory cascade. This model has and will continue to have a vital role in drug development. The mitogen-activated protein kinases (MAPKs), such as extracellular signal-regulated kinase (ERK), the p38 mitogen-activated protein kinase (p38 MAPK), and the c-Jun NH2-terminal kinase (JNK), are a group of signaling proteins—their phosphorylation is acknowledged as a critical component for the generation of pro-inflammatory cytokines in activated microglia, and they play a vital role in neuroinflammation [6,7]. We investigate the anti-inflammatory effects and molecular mechanisms of ICSB in LPS-induced microglial cells and CA-induced inflammatory mice that influence MAPK signaling pathways
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