Fig 2: Hyaluronan preincubation attenuated inflammatory cytokine-induced CEMIP expression in OUMS-27 cells. OUMS-27 cells were incubated for 3 h with HA or medium alone and then treated with IL-1β for 12 h. (a) CEMIP mRNA expression was measured by qRT-PCR. (b) CEMIP protein expression was examined by Western blotting. Relative CEMIP expression change was measured by densitometric analysis. Values represent mean ± SD (a, mRNA, n = 6 per group; b, protein, n = 3 per group). ** p < 0.01 vs. control. ## p < 0.01 vs. HA preincubation group.

Fig 3: ERK inhibitor attenuated IL-1β-induced CEMIP expression in OUMS-27 cells. (a) ERK inhibitor (FR180204, 50 μM) was added 1 h prior to IL-1β stimulation and CEMIP mRNA expression was analyzed 12 h after treatment with IL-1β. (b) FR180204 attenuated IL-1β-induced CEMIP protein expression at 12 h in a dose-dependent manner. Values represent mean ± SD (n = 6 per group). ## p < 0.01 vs. IL-1β-treated group. ** p < 0.01 vs. IL-1β with low dose (5 μM) FR180204 treated group.

Fig 4: Localization of HYAL1, HYAL2, CEMIP and TMEM2 in the human skin. (A,B) Immunohistochemical and immunohistofluorescent analysis of IgG, (C,D) HYAL1, (E,F) HYAL2, (G,H) CEMIP and (I,J) TMEM2. The insets provide enlarged views of the regions in the gray boxes. The nuclei were stained with Höechst (blue). The asterisks indicate the fibroblasts, and the arrows indicate the keratinocytes. The scale bars at the bottom of the figures represent 50 µm. Abbreviations: hyaluronidase (HYAL), Transmembrane Protein type II (TMEM2), Cell Migration Inducing Hyaluronan Binding Protein (CEMIP).

Fig 5: Additive effect of the HYBID expression by TNF-α and IL-6 in OA chondrocytes. (a) and (b) Effects of TNF-α and IL-6 on the mRNA and protein expression of HYBID in OA chondrocytes, respectively. OA chondrocytes at P2 were treated with TNF-α (0 or 10 ng/ml) and/or IL-6 (0, 100 ng/ml) in the presence of sIL-6R (100 ng/ml) for 24 h and 48 h, and cell lysates were subjected to quantitative real-time PCR for HYBID expression using the ΔΔCt method (a) and immunoblotting with anti-HYBID antibody, followed by densitometric analysis (b). The average HYBID:GAPDH ratio in control OA chondrocytes treated with vehicle was set at 1. Values are expressed mean ± SD (n = 3). *, P < 0.05; **, P < 0.01. The uncropped full-length gels are presented in Supplementary Figure S5.