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. 2022 Feb 17;12(7):3073–3084. doi: 10.1016/j.apsb.2022.02.010

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SHP099 inhibited macrophage M1 polarization and inflammatory cytokines secretion during LPS-induced inflammation for 24 h. (A) Flow cytometry for detection and quantitative analysis of CD80⁺ cells after the treatment of lipopolysaccharide (LPS) with or without SHP099 for 24 h in RAW264.7 cells and bone marrow-derived macrophages (BMDMs) (n = 3). One-way ANOVA with Tukey's multiple comparison test; ∗∗P < 0.01. (B) qPCR analysis of mRNA of M1-related genes, interleukin-1β (Il1b), Il6, tumor necrosis factor α (Tnfa), inducible nitric oxide synthase (iNos) and CXC chemokineligand-10 (Cxcl10) in RAW264.7 cells after LPS stimulation with or without SHP099 treatment for 24 h (n = 3). One-way ANOVA with Tukey's multiple comparison test; ∗∗P < 0.01, ns represents no significance. (C) qPCR analysis of mRNA of M1-related genes, Il1b, Il6, Tnfa, iNos and Cxcl10 in BMDMs after LPS stimulation with or without SHP099 treatment for 24 h (n = 3). One-way ANOVA with Tukey's multiple comparison test; ∗∗P < 0.01, ns represents no significance. (D) Enzyme-linked immunosorbent assay (ELISA) results of TNF-α and IL-6 secretion of RAW264.7 cells after LPS stimulation with or without SHP099 treatment for 24 h (n = 3). One-way ANOVA with Tukey's multiple comparison test; ∗∗P < 0.01, ns represents no significance. (E) ELISA results of TNF-α and IL-6 secretion of BMDMs cells after LPS stimulation with or without SHP099 treatment for 24 h. One-way ANOVA with Tukey's multiple comparison test. ∗∗P < 0.01. (F) Western blot analysis of iNOS and cyclooxygenase 2 (COX2) in RAW264.7 after LPS stimulation with or without SHP099 treatment for 48 h. (G) Western blot analysis of iNOS and COX2 in BMDMs after LPS stimulation with or without SHP099 treatment for 48 h. (H) Western blot analysis of iNOS and COX2 in RAW264.7 after LPS stimulation with over-expression (OE) of SHP2 or not (NC) for 48 h. All data are shown as the mean ± SEM.