Nuciferine Inhibits Proinflammatory Cytokines via the PPARs in LPS-Induced RAW264.7 Cells

Chao Zhang, Jianjun Deng, Dan Liu, Xingxia Tuo, Yan Yu, Haixia Yang, Nanping Wang, Chao Zhang, Jianjun Deng, Dan Liu, Xingxia Tuo, Yan Yu, Haixia Yang, Nanping Wang

Abstract

Inflammation is important and has been found to be an underlying cause in many acute and chronic human diseases. Nuciferine, a natural alkaloid containing an aromatic ring, is found in the nelumbo nucifera leaves. It has been shown to have potential anti-inflammatory activities, but the molecular mechanism has remained unclear. In this study, we found that nuciferine (10 μM) significantly inhibited the lipopolysaccharide (LPS)-induced inflammatory cytokine IL-6 and TNF-α production in RAW 264.7 cells. In addition, the luciferase reporter assay results of different subtypes of the peroxisome proliferator-activated receptor (PPAR) showed that nuciferine dose-dependently activated all the PPAR activities. Specific inhibitors of PPARα and PPARγ significantly abolished the production of inflammatory cytokines as well as IκBα degradation. However, PPARδ inhibitor did not show this effect. Our results suggested a potential molecular mechanism of the anti-inflammatory effects of nuciferine in LPS-induced inflammation, at least in part, by activating PPARα and PPARγ in RAW 264.7 cells.

Keywords: IL-6; PPARs; TNF-α; inflammation; nuciferine.

Conflict of interest statement

The authors declare no conflict of interest.

Figures

Figure 1
Figure 1
Cytotoxicity of Nuciferine on macrophage RAW264.7 cells. Cell viabilities of RAW264.7 cells treated with Nuciferine (0, 1, 3, 10 or 50 μM), for 24 h, were measured by MTT assay.
Figure 2
Figure 2
Nuciferine inhibits the LPS-induced TNFα and IL-6 production in RAW264.7 cells. RAW 264.7 cells were pretreated with nuciferine (0, 1, 10 or 50 μM for 24 h) and then stimulated with the LPS (500 ng/mL for 12 h), with a nuciferine withdrawal. (A,B) TNFα and IL-6 releases and (C,D) mRNA level of TNFα and IL-6, respectively. * p < 0.05 ** p < 0.01 vs. control, #p < 0. 05 ##p < 0.01 vs. LPS treatment.
Figure 3
Figure 3
Effect of Nuciferine on PPARs transcription activities. (A–C) Luciferase reporter assay in RAW264.7 cells. (D) The relative mRNA expression of PPARs target genes. * p < 0.05 ** p < 0.01 vs. control.
Figure 4
Figure 4
Antagonist of PPARα and PPARγ abolished the effects of nuciferine on the LPS-induced TNFα and IL-6 production. RAW264.7 cells were pretreated with GW6417/GSK0660/GW9662, for 12 h, followed by the nuciferine incubation, for 24 h, and then stimulated with LPS for 12 h. (A,B) production of the IL-6 and the TNFα in a cell medium supernatant. (C,D) mRNA expression of IL-6 and TNFα. * p < 0.05 ** p < 0.01 vs. control. #p < 0.05, ##p < 0.01 vs. LPS treatment. $p < 0.05 vs. antagonist pretreatment followed by the LPS stimulation.
Figure 5
Figure 5
Anti-inflammatory effects of nuciferine on the LPS-induced inflammatory response, are PPARs dependent. RAW264.7 cells were pretreated with GW6417/GSK0660/GW9662, for 12 h, followed by the Nuciferine incubation and then stimulated by the LPS with a nuciferine withdrawal. Levels of the expression of IκBα were detected by Western blotting. * p < 0.05 vs. control group; #p < 0.05 vs. LPS group.

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