Abstract
BackgroundMorus alba L. bark has been widely used in traditional medicine for treating several inflammatory diseases, such as hypertension, diabetes mellitus and coughing; however, the molecular mechanisms underlying its anti-inflammatory effects are not well understood.MethodsWe examined the effects of an extract of Morus alba L. bark (MabE) on Toll-like receptor (TLR) ligand-induced activation of RAW264.7 macrophages using a luciferase reporter assay and immunoassays. For the in vivo experiment, we used an imiquimod-induced ear edema model to examine the anti-inflammatory effects of MabE.ResultsMabE inhibited the TLR ligand-induced activation of NF-κB in RAW264.7 cells without affecting their viability. Consistent with the inhibition of NF-κB activation, MabE also inhibited the production of IL-6 and IL-1β from TLR ligand-treated RAW264.7 cells. In vivo MabE treatment inhibited the ear swelling of IMQ-treated mice, in addition to the mRNA expression of IL-17A, IL-1β and COX-2. The increases in splenic γδT cells in IMQ-treated mice and the production of IL-17A from splenocytes were significantly inhibited by MabE treatment.ConclusionOur study suggests that the anti-inflammatory effects of MabE on the activation of the macrophage cell line RAW246.7 by TLRs and IMQ-induced ear edema are through the inhibition of NF-κB activation and IL-17A-producing γδT cells, respectively.
Highlights
Morus alba L. bark has been widely used in traditional medicine for treating several inflammatory diseases, such as hypertension, diabetes mellitus and coughing; the molecular mechanisms underlying its anti-inflammatory effects are not well understood
The cell types and location of Toll-like receptors (TLRs) expression may vary depending on the receptor type [12, 13], there are many TLRs expressed on immune cells, such as macrophages, dendritic cells and neutrophils, that play important roles in the initiation of innate immunity [11]
Using the dose and time of each TLR ligand yielding similar degrees of Nuclear factor-kappa B (NF-κB) activity, we examined the effects of MabE on TLR ligand stimulation in RAW264.7 cells
Summary
We examined the effects of an extract of Morus alba L. bark (MabE) on Toll-like receptor (TLR) ligandinduced activation of RAW264.7 macrophages using a luciferase reporter assay and immunoassays. Experimental materials The extract of Morus alba L. bark (Sohakuhi) was prepared as described previously [21]. Co., Ltd., Osaka, Japan) was refluxed in MeOH (5 L, 90 min, × 2) to obtain the MeOH extract, and subsequently suspended in water (500 mL) and defatted by Hexane (500 mL, × 3) to separate the aqueous layer. The aqueous layer was further extracted with EtOAc (500 mL, × 3) to obtain EtOAc layer. This EtOAc layer was used as an extract of Morus alba L. bark (MabE). The extract was further diluted in 50% EtOH
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