Wu‐Mei‐Wan Reduces Insulin Resistance via Inhibition of NLRP3 Inflammasome Activation in HepG2 Cells

Figures

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  Figure 1. Effects of 0.25 mM palmitate (PA) on glucose consumption rate of HepG2 cells at different treatment times. Control group is HepG2 cells in the normal state without PA treatment, whereas PA 12 h, PA 24 h, and PA 48 h groups are HepG2 cells treated with 0.25 mM palmitate for 12, 24, and 48 h, respectively. Compared to the control group, the glucose consumption rate in palmitate-treated HepG2 cells for 12, 24, and 48 h was 81.3, 63, and 62.3%, respectively. ∗P < 0.05; ∗∗∗P < 0.001.

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  Figure 2. Effects of the medicated serum of WMW on cell proliferation in palmitate-treated HepG2 cells. HepG2 cells were treated with 0.25 mM palmitate for 24 h, and normal serum without WMW (N) or medicated serum containing low (L), middle (M), or high (H) dose of WMW was then added for 24 h. The number of viable cells was examined by the MTT assay.

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  Figure 3. Effects of WMW on insulin signaling pathway in HepG2 cells. HepG2 cells were treated with 10% serum without WMW (N) or medicated serum containing high (H) dose of WMW for 24 h. Then, cells were stimulated with or without 100 nM insulin for 15 min, and cell extracts were collected. The results of western blot analysis of p-IR (a), p-IRS-1 (Ser307) (b), PI3K (p85) (c), p-AKT (d), and GLUT4 (e) are presented by a bar graph. ∗P < 0.05; ∗∗P < 0.01.

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  Figure 4. Effects of WMW on the secretion of proinflammatory cytokines in HepG2 cells. HepG2 cells were treated with serum without WMW (N) or medicated serum containing low (L), middle (M), or high (H) dose of WMW for 24 h. Then, cells were stimulated with 5 μM nigericin for 45 min, and the supernatants were collected and analyzed by ELISA for measurement of IL-1β (a), MCP-1α (b), and TNF-α (c) concentrations. ∗P < 0.05, ∗∗P < 0.01, and ∗∗∗P < 0.001.

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  Figure 5. Effects of WMW on mRNA expression of NLRP3 inflammasome in HepG2 cells. Control group is HepG2 cells in the normal state with no palmitate (PA) treatment. HepG2 cells were treated with serum without WMW (PA + N) or medicated serum containing low (PA + L), middle (PA + M), or high (PA + H) dose of WMW for 24 h. RT-qPCR results showed that the medicated serum of WMW increased the mRNA expression of IL-1β (a), NLRP3 (b), and caspase-1 (c). ∗P < 0.05, ∗∗P < 0.01, and ∗∗∗P < 0.001.

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  Figure 6. Effects of WMW on the expression of proteins related to NLRP3 inflammasome in HepG2 cells. HepG2 cells were treated with serum without WMW (N) or medicated serum containing low (L), middle (M), or high (H) dose of WMW for 24 h. Then, cells were stimulated with 5 μM nigericin for 45 min, and cell extracts were collected and analyzed by western blotting for measurement of NLRP3 (a), ASC (b), caspase-1 (p20) (c), and IL-1β (d) expression. Immunofluorescence staining for caspase-1 (p20) (red) and IL-1β (red) in HepG2 cells from the control group, disease group (PA), and group cotreated with serum containing high dose of WMW (PA + WMW) is shown in ((e) and (f)). The nuclei were counterstained with DAPI (blue). Signals for caspase-1 (P20) and IL-1β are presented in a bar graph ((g) and (h)). Scale, 100 μm. ∗P < 0.05, ∗∗P < 0.01, and ∗∗∗P < 0.001.