ADAMTS5 Antibody - #DF13268

Fig. 2 miR-17 inhibits cartilage destruction by targeting multiple pathological catabolic factors. a Luciferase activity of 293 T cells co-transfected with dual-luciferase reporter constructs containing wild-type or mutated 3′-UTRs, as well as negative control (mimic NC) or miR-17 mimic. The data presented are mean percentage changes over mimic NC ± s.e.m. n = 6 biologically independent samples (NC + wt UTR, miR-17+wt UTR); n = 4 biologically independent samples (NC + mut UTR, miR-17 + mut UTR). b Mouse articular chondrocytes were transfected with miR-17 mimic (50 nM) or mimic NC and treated with or without IL-1β (5 ng/mL) for 24 h. The protein levels of catabolic and anabolic factors were determined by western blot followed by densitometry analysis. Blots are representative of three independent experiments. c, d qRT-PCR analysis of catabolic genes in knee cartilage (c), and representative images of IHC staining and quantification of MMP13+, ADAMTS5+ and NOS2+ cells in joint sections (d) of mice subjected to sham or DMM surgery and intra-articular injections of agomir-NC or agomir-17 (1.5 nmol) for 4 weeks, beginning at 4 weeks after surgery. n = 5 biologically independent samples per group in c. For MMP13 and ADAMTS5 staining, n = 4 mice (sham); n = 5 mice (DMM, DMM + agomir-17). For NOS2 staining, n = 4 mice per group. e, f qRT-PCR analysis of catabolic genes (e) and representative images of MMP13, ADAMTS5, and NOS2 immunostaining, and quantification of positive cells (f) from knee joints of cKO mice subjected to DMM surgery and intra-articular injection of agomir-NC, or agomir-17 (2 nmol). The injections were performed at 1 week after surgery and samples were collected at 2.5 weeks after surgery. n = 4 biologically independent samples per group in (e); n = 4 mice per group in (f). All scale bars, 100 μm. Data are presented as mean ± s.e.m. or boxplots (center line, median; box limits, 25 to 75th percentiles; whiskers, min to max), and dots represent individual mice or biologically independent samples. NS, not significant, *P < 0.05, **P < 0.01, and ***P < 0.001 in (a, c–f). *P < 0.05 versus control group, °P < 0.05 versus IL-1β + mimic NC in (b). Two-sided Student’s t test for (a); one-way ANOVA with Bonferroni’s test for (b–f). Exact P values in (b): *P = 0.0011, °P = 0.0205 (MMP3); *P <0.0001, °P = 0.0039 (MMP13); *P = 0.0015, °P = 0.0040 (ADAMTS5); *P = 0.0001, °P = 0.0013 (NOS2); *P = 0.0013, P > 0.9999 (SOX9); *P = 0.0001, P > 0.9999 (COL2A1). Source data are provided as a Source Data file.

Fig. 2 miR-17 inhibits cartilage destruction by targeting multiple pathological catabolic factors. a Luciferase activity of 293 T cells co-transfected with dual-luciferase reporter constructs containing wild-type or mutated 3′-UTRs, as well as negative control (mimic NC) or miR-17 mimic. The data presented are mean percentage changes over mimic NC ± s.e.m. n = 6 biologically independent samples (NC + wt UTR, miR-17+wt UTR); n = 4 biologically independent samples (NC + mut UTR, miR-17 + mut UTR). b Mouse articular chondrocytes were transfected with miR-17 mimic (50 nM) or mimic NC and treated with or without IL-1β (5 ng/mL) for 24 h. The protein levels of catabolic and anabolic factors were determined by western blot followed by densitometry analysis. Blots are representative of three independent experiments. c, d qRT-PCR analysis of catabolic genes in knee cartilage (c), and representative images of IHC staining and quantification of MMP13+, ADAMTS5+ and NOS2+ cells in joint sections (d) of mice subjected to sham or DMM surgery and intra-articular injections of agomir-NC or agomir-17 (1.5 nmol) for 4 weeks, beginning at 4 weeks after surgery. n = 5 biologically independent samples per group in c. For MMP13 and ADAMTS5 staining, n = 4 mice (sham); n = 5 mice (DMM, DMM + agomir-17). For NOS2 staining, n = 4 mice per group. e, f qRT-PCR analysis of catabolic genes (e) and representative images of MMP13, ADAMTS5, and NOS2 immunostaining, and quantification of positive cells (f) from knee joints of cKO mice subjected to DMM surgery and intra-articular injection of agomir-NC, or agomir-17 (2 nmol). The injections were performed at 1 week after surgery and samples were collected at 2.5 weeks after surgery. n = 4 biologically independent samples per group in (e); n = 4 mice per group in (f). All scale bars, 100 μm. Data are presented as mean ± s.e.m. or boxplots (center line, median; box limits, 25 to 75th percentiles; whiskers, min to max), and dots represent individual mice or biologically independent samples. NS, not significant, *P < 0.05, **P < 0.01, and ***P < 0.001 in (a, c–f). *P < 0.05 versus control group, °P < 0.05 versus IL-1β + mimic NC in (b). Two-sided Student’s t test for (a); one-way ANOVA with Bonferroni’s test for (b–f). Exact P values in (b): *P = 0.0011, °P = 0.0205 (MMP3); *P <0.0001, °P = 0.0039 (MMP13); *P = 0.0015, °P = 0.0040 (ADAMTS5); *P = 0.0001, °P = 0.0013 (NOS2); *P = 0.0013, P > 0.9999 (SOX9); *P = 0.0001, P > 0.9999 (COL2A1). Source data are provided as a Source Data file.

Fig. 2 PTX3 accelerates synovitis, cartilage erosion, and exacerbates OA development in mice. A, B Safranin O/fast green staining and Osteoarthritis Research Society International (OARSI) grades of knee cartilage from DMM mice treated with vehicle, recombinant mouse PTX3 (rmPTX3), or PTX3 neutralizing antibody (PTX3-NAb) for 4 weeks and 8 weeks. Scale bar: 100 µm. C–F IF staining and quantification of ADATMS5 and MMP13 in knee cartilage of controls and DMM mice treated with vehicle, rmPTX3, or PTX3-NAb for 8 weeks. Scale bar: 50 µm. G IF staining and quantification of COL2 in knee cartilage of controls and DMM mice treated with rmPTX3 or PTX3-NAb for 8 weeks. Scale bar: 50 µm. H, I H&E and synovitis score of synovial tissues from DMM mice treated with vehicle, rmPTX3, or PTX3-NAb for 4 weeks and 8 weeks. Scale bar: 100 µm. *P 

Fig. 4. Knockdown of FOXO3a compromised the inhibitory effects of TMF on C/EBPβ-mediated ECM degradation. (A–B) IHC assays were used to detect the altered expression of FOXO3a. (C–D) The protein expression of FOXO3a in shRNA-FOXO3a-infected C28/I2 cells was detected by western blotting assays. (E–K) Western blotting assays were used to detect the protein expression of Aggrecan, ADAMTS5, C/EBPβ, p-C/EBPβ, caspase3, cleaved-caspase3, and FOXO3a in LV-sh-FOXO3a-infected C28/I2 cells treated with IL-1β. (L-M) The apoptosis rate was determined by a flow cytometer.

Figure 4. Exploration of the effects of two treatments on cartilage matrix in vivo. (A and B) The immunohistochemical staining was performed to clarify the expression of Collagen II, MMP13 and ADAMTS5 in articulate (The magnification in A is 100×, scale bar = 100 μm; in B is 400×, scale bar = 20 μm;). (C) Statistical analysis of the percentage of pixels in the image or selection in three groups. Data are expressed as mean ±SEM. Different number of asterisk (*) show significant differences between groups. (*= P