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Fig. 5 | Arthritis Research & Therapy

Fig. 5

From: Soluble Siglec-9 suppresses arthritis in a collagen-induced arthritis mouse model and inhibits M1 activation of RAW264.7 macrophages

Fig. 5

a Potential mechanism of action of soluble sialic acid-binding immunoglobulin-type lectin (sSiglec)-9 in murine macrophages. Sialidase was used to assess the effects of eliminating sialic acid from the cell surface on sSiglec-9 efficacy. sSiglec-9 treatment suppressed the messenger RNA (mRNA) expression of M1 markers. However, pretreatment with sialidase significantly blunted the suppressive effect of sSiglec-9, suggesting that sSiglec-9 required sialic acid to bind to its receptor. Pretreatment of cells with RS504393, a specific inhibitor for C-C chemokine receptor type 2 (CCR2), did not affect the suppressive effect of sSiglec-9, suggesting that CCR2 was not the primary receptor for sSiglec-9. *p < 0.05, **p < 0.01 (Bonferroni post hoc test). b Effects of sSiglec-9 on mRNA expression of M1 macrophage markers in RAW264.7 cells and peritoneal macrophages (pMACs). mRNA expression of M1 macrophage markers (tumor necrosis factor [TNF]-α, interleukin [IL]-6, and inducible nitric oxide synthase [iNOS]) was determined by real-time polymerase chain reaction (RT-PCR). Cells were stimulated with interferon (IFN)-γ in the presence or absence of sSiglec-9 (0–20 ng/ml) for 12 h. Expression of glyceraldehyde 3-phosphate dehydrogenase (GAPDH) mRNA was used as a control. mRNA expression of all M1 markers was significantly suppressed by sSiglec-9 treatment in both cell types. *p < 0.05, **p < 0.01 versus samples stimulated with IFN-γ without sSiglec-9 treatment (Bonferroni post hoc test). c Effects of sSiglec-9 on mRNA expression of M2 macrophage markers in RAW264.7 cells and pMACs. mRNA expression of M2 macrophage markers (CD206, Arginase-1, and IL-10) was determined by RT-PCR under the same conditions as described above for M1 markers. mRNA expression of all M2 markers did not significantly change with sSiglec-9 treatment in both cell types (Bonferroni post hoc test). n.s. not significant

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