Human MMP-13 Antibody

Detection of MMP-13 by Western Blot

MiR-23b-3p inhibits HS6ST2 expression by targeting HS6ST2 mRNA and effects specific gene expression in chondrocytes. A, b Dual luciferase reporter assay to validate target relationship between HS6ST2 and miR-23b-3p. SW1353 cells were transfected with mimic miR-23b-3p (a) or anti-miR-23b-3p (b) by control vector, pmirGLO-HS6ST2 wild-type 3′UTR vector or pmirGLO-HS6ST2 mutant 3′UTR vector, respectively, for 48 h. c, d Stem-loop RT-qPCR and western blotting results in SW1353 cells (c) or C28/I2 cells (d) transfected with 10 nM mimic miR-23b-3p (left panel) or 50 nM anti-miR-23b-3p sequence (right panel). e, f RT-qPCR (e) and western blotting (f) results of cartilage-specific gene expression in SW1353 cells transfected with mimic miR-23b-3p or negative control. g, h RT-qPCR (g) and western blotting (h) results of cartilage-specific gene expression in SW1353 cells transfected with anti-miR-23b-3p or negative control. RNA was harvested at 24 h, while protein was isolated at 48 h after transfection. U6 snRNA was used as internal controls in miRNA stem-loop RT-qPCR detection and GAPDH was used as internal controls in mRNA RT-qPCR and western blotting detection. Bars represent standard error of the mean (SEM) from three independent experiments. Mann–Whitney U test was used to identify statistical differences between two groups. *P value < 0.05 Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/29899528), licensed under a CC-BY license. Not internally tested by R&D Systems.

Detection of MMP-13 by Western Blot

MiR-23b-3p could enhance matrix degradation by means of regulating activity of p38 MAPK in human chondrocytes under TNF-alpha treatment. A, b SW1353 cells were transfected by mimic miR-23b-3p (a) or anti-miR-23b-3p sequence (b) with or without TNF-alpha for 24 h, and the protein expression of phosphorylation form of p38 MAPK (p-p38), total p38 MAPK, and MMP13 was detected by western blotting. Asterisk (*): compared with the mimic NC (a) or anti-NC group (b). c SW1353 cells were treated with mimic miR-23b-3p with or without p38 MAPK inhibitor SB203580 (10 μM) under stimulation of TNF-alpha. The protein expression of MMP13 was determined by western blotting. Asterisk (*): compared with the mimic NC group. d Interfering efficiency of siRNAs against p38 MAPK was determined by western blotting under 50 nM p38 siRNA, 50 nM negative control (NC), and Mock (transfection regent only) transfection for 48 h. e Under stimulation with TNF-alpha, SW1353 cells were transfected by mimic miR-23b-3p under treatment with si-p38 mixture (containing three siRNA target sequences), and p-p38, total p38, and MMP13 were determined by western blotting. Asterisk (*): compared with mimic NC or si-NC. Each relative expression of phosphorylation form was normalized by the total form. GAPDH was used as internal controls in western blotting detection. Mann–Whitney U test was used to identify statistical differences between two groups. Asterisk (*) or hash (#) stands for P value <0.05. NS stands for not significant Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/29899528), licensed under a CC-BY license. Not internally tested by R&D Systems.

Detection of MMP-13 by Western Blot

MiR-23b-3p could enhance matrix degradation by means of regulating activity of p38 MAPK in human chondrocytes under TNF-alpha treatment. A, b SW1353 cells were transfected by mimic miR-23b-3p (a) or anti-miR-23b-3p sequence (b) with or without TNF-alpha for 24 h, and the protein expression of phosphorylation form of p38 MAPK (p-p38), total p38 MAPK, and MMP13 was detected by western blotting. Asterisk (*): compared with the mimic NC (a) or anti-NC group (b). c SW1353 cells were treated with mimic miR-23b-3p with or without p38 MAPK inhibitor SB203580 (10 μM) under stimulation of TNF-alpha. The protein expression of MMP13 was determined by western blotting. Asterisk (*): compared with the mimic NC group. d Interfering efficiency of siRNAs against p38 MAPK was determined by western blotting under 50 nM p38 siRNA, 50 nM negative control (NC), and Mock (transfection regent only) transfection for 48 h. e Under stimulation with TNF-alpha, SW1353 cells were transfected by mimic miR-23b-3p under treatment with si-p38 mixture (containing three siRNA target sequences), and p-p38, total p38, and MMP13 were determined by western blotting. Asterisk (*): compared with mimic NC or si-NC. Each relative expression of phosphorylation form was normalized by the total form. GAPDH was used as internal controls in western blotting detection. Mann–Whitney U test was used to identify statistical differences between two groups. Asterisk (*) or hash (#) stands for P value <0.05. NS stands for not significant Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/29899528), licensed under a CC-BY license. Not internally tested by R&D Systems.

Detection of MMP-13 by Western Blot

MiR-23b-3p could enhance matrix degradation by means of regulating activity of p38 MAPK in human chondrocytes under TNF-alpha treatment. A, b SW1353 cells were transfected by mimic miR-23b-3p (a) or anti-miR-23b-3p sequence (b) with or without TNF-alpha for 24 h, and the protein expression of phosphorylation form of p38 MAPK (p-p38), total p38 MAPK, and MMP13 was detected by western blotting. Asterisk (*): compared with the mimic NC (a) or anti-NC group (b). c SW1353 cells were treated with mimic miR-23b-3p with or without p38 MAPK inhibitor SB203580 (10 μM) under stimulation of TNF-alpha. The protein expression of MMP13 was determined by western blotting. Asterisk (*): compared with the mimic NC group. d Interfering efficiency of siRNAs against p38 MAPK was determined by western blotting under 50 nM p38 siRNA, 50 nM negative control (NC), and Mock (transfection regent only) transfection for 48 h. e Under stimulation with TNF-alpha, SW1353 cells were transfected by mimic miR-23b-3p under treatment with si-p38 mixture (containing three siRNA target sequences), and p-p38, total p38, and MMP13 were determined by western blotting. Asterisk (*): compared with mimic NC or si-NC. Each relative expression of phosphorylation form was normalized by the total form. GAPDH was used as internal controls in western blotting detection. Mann–Whitney U test was used to identify statistical differences between two groups. Asterisk (*) or hash (#) stands for P value <0.05. NS stands for not significant Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/29899528), licensed under a CC-BY license. Not internally tested by R&D Systems.

Detection of MMP-13 by Western Blot

TNF-alpha -treated chondrocytes increases matrix degradation regulated by miR-23b-3p.a Stem-loop RT-qPCR result of miR-23b-3p (left panel) and protein levels of HS6ST2 and MMP13 (right panel) were assayed in SW1353 cells under stimulation by 10 ng/ml TNF-alpha for 24 h. b, c Western blotting result of HS6ST2 and MMP13 protein in SW1353 cells transfected with 10 nM mimic miR-23b-3p (b) or 50 nM anti-miR-23b-3p sequence (c) and stimulated by 10 ng/ml TNF-alpha for 24 h. d, e Toluidine blue staining results of SW1353 cells transfected with 10 nM mimic miR-23b-3p (d) or 50 nM anti-miR-23b-3p sequence (e) and stimulated by 10 ng/ml TNF-alpha for 24 h. Scale bar, 100 μm. Lower panel, statistical analysis of average optical density of matrix staining of toluidine blue. Asterisk (*): compared with mimic NC or anti-NC group. U6 snRNA and GAPDH were used as internal controls in RT-qPCR for miRNA and western blotting detection, respectively. Bars represent standard error of the mean (SEM) from three independent experiments. One representative result and quantitative data from three independent western blotting and toluidine blue staining. Mann–Whitney U test was used to identify statistical differences between two groups. Asterisk (*) or hash (#) stands for P value <0.05 Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/29899528), licensed under a CC-BY license. Not internally tested by R&D Systems.

Detection of MMP-13 by Western Blot

MiR-23b-3p could enhance matrix degradation in human chondrocytes via regulating HS6ST2.a, b SW1353 cells transfected with 50 nM HS6ST2 siRNA mixture (containing three target sequences, si-HS6ST2) or negative control (si-NC) were stimulated by 10 ng/ml TNF-alpha for 24 h. Protein level of HS6ST2 and MMP13 were assayed by western blotting (a) and matrix content of chondrocytes was determined by toluidine blue staining (b). Asterisk (*): compared with the si-NC group. c, d SW1353 cells transfected with empty vector (FLAG-Ctrl) or pcDNA3.1-FLAG-HS6ST2 vector (FLAG-HS6ST2) for 24 h were stimulated by TNF-alpha for another 24 h. The protein expression of MMP13 was assayed by western blotting (c) and matrix content of chondrocytes was determined by toluidine blue staining (d). Asterisk (*): compared with the FLAG-Ctrl group. e, f Under stimulation with TNF-alpha, SW1353 cells were treated with mimic NC or mimic miR-23b-3p for 24 h and then transfected with empty vector (FLAG-Ctrl) or pcDNA3.1-FLAG-HS6ST2 vector (FLAG-HS6ST2) for another 24 h to observe the rescuing effect of mimic miR-23b-3p in MMP13 expression (e) and matrix content (f). Asterisk (*): compared with the mimic NC group. GAPDH was used as internal controls in western blotting detection. In toluidine blue staining results, scale bar, 100 μm. Lower panel, statistical analysis of average optical density of matrix staining of toluidine blue. Bars represent standard error of the mean (SEM) from three independent experiments. Mann–Whitney U test was used to identify statistical differences between two groups. Asterisk (*) or hash (#) stands for P value <0.05. NS stands for not significant Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/29899528), licensed under a CC-BY license. Not internally tested by R&D Systems.

Detection of MMP-13 by Western Blot

TNF-alpha -treated chondrocytes increases matrix degradation regulated by miR-23b-3p.a Stem-loop RT-qPCR result of miR-23b-3p (left panel) and protein levels of HS6ST2 and MMP13 (right panel) were assayed in SW1353 cells under stimulation by 10 ng/ml TNF-alpha for 24 h. b, c Western blotting result of HS6ST2 and MMP13 protein in SW1353 cells transfected with 10 nM mimic miR-23b-3p (b) or 50 nM anti-miR-23b-3p sequence (c) and stimulated by 10 ng/ml TNF-alpha for 24 h. d, e Toluidine blue staining results of SW1353 cells transfected with 10 nM mimic miR-23b-3p (d) or 50 nM anti-miR-23b-3p sequence (e) and stimulated by 10 ng/ml TNF-alpha for 24 h. Scale bar, 100 μm. Lower panel, statistical analysis of average optical density of matrix staining of toluidine blue. Asterisk (*): compared with mimic NC or anti-NC group. U6 snRNA and GAPDH were used as internal controls in RT-qPCR for miRNA and western blotting detection, respectively. Bars represent standard error of the mean (SEM) from three independent experiments. One representative result and quantitative data from three independent western blotting and toluidine blue staining. Mann–Whitney U test was used to identify statistical differences between two groups. Asterisk (*) or hash (#) stands for P value <0.05 Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/29899528), licensed under a CC-BY license. Not internally tested by R&D Systems.

Detection of MMP-13 by Western Blot

TNF-alpha -treated chondrocytes increases matrix degradation regulated by miR-23b-3p.a Stem-loop RT-qPCR result of miR-23b-3p (left panel) and protein levels of HS6ST2 and MMP13 (right panel) were assayed in SW1353 cells under stimulation by 10 ng/ml TNF-alpha for 24 h. b, c Western blotting result of HS6ST2 and MMP13 protein in SW1353 cells transfected with 10 nM mimic miR-23b-3p (b) or 50 nM anti-miR-23b-3p sequence (c) and stimulated by 10 ng/ml TNF-alpha for 24 h. d, e Toluidine blue staining results of SW1353 cells transfected with 10 nM mimic miR-23b-3p (d) or 50 nM anti-miR-23b-3p sequence (e) and stimulated by 10 ng/ml TNF-alpha for 24 h. Scale bar, 100 μm. Lower panel, statistical analysis of average optical density of matrix staining of toluidine blue. Asterisk (*): compared with mimic NC or anti-NC group. U6 snRNA and GAPDH were used as internal controls in RT-qPCR for miRNA and western blotting detection, respectively. Bars represent standard error of the mean (SEM) from three independent experiments. One representative result and quantitative data from three independent western blotting and toluidine blue staining. Mann–Whitney U test was used to identify statistical differences between two groups. Asterisk (*) or hash (#) stands for P value <0.05 Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/29899528), licensed under a CC-BY license. Not internally tested by R&D Systems.