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. 2019 Sep 18;10:2243. doi: 10.3389/fimmu.2019.02243

Figure 6.

Figure 6

RAGE-mediated p38/MAPK and TLR4-mediated NF-κB are responsible for the chemotaxis and activation of MDSCs resulted by S100A9. (A) Western blot analysis of p-p38, p-ERK, p-JNK, p-AKT, and p-p65 expression in LoVo-induced MDSCs treated by GST-S100A9 (20 μg/ml) or GST (20 μg/ml) for 0, 15, 30, and 60 min. (B) Western blot analysis of p-p38 and p-p65 expression in GST-S100A9-treated MDSCs with inhibitors FPS-ZM1 (100 nM) and TAK-242 (100 nM) for 60 min. (C) Chemotaxis assay for LoVo-induced MDSCs pretreated with and without inhibitor SB203580 (50 nM) for 30 min followed by stimulation with GST-S100A9 (20 μg/ml) and GST (20 μg/ml) proteins for 24 h. (D–F) Real-time PCR analysis for mRNA expression of immunosuppressive molecules Arg-1 (D), iNOS (E), and IL-10 (F) in LoVo-induced MDSCs pretreated with and without inhibitor BAY11-7082 (50 nM) for 30 min followed by stimulation with GST-S100A9 (20 μg/ml) and GST (20 μg/ml) proteins for 24 h. (G) Fluorescence intensity analysis for ROS production in LoVo-induced MDSCs pretreated with and without inhibitor BAY11-7082 (50 nM) for 30 min followed by stimulation with GST-S100A9 (20 μg/ml) and GST (20 μg/ml) proteins for 24 h. Data represent three independent experiments and are represented as Mean±SD. *p < 0.05, **p < 0.01, ***p < 0.001.