primary antibody against adamts5  (Boster Bio)

Journal: Aging (Albany NY)

Article Title: Lei’s formula attenuates osteoarthritis mediated by suppression of chondrocyte senescence via the mTOR axis: in vitro and in vivo experiments

doi: 10.18632/aging.205582

Figure Lengend Snippet: LSF regulates chondrocyte anabolic-catabolic homeostasis and inhibits chondrocyte senescence. ( A – D ) Western blotting assessed the expression of Col2, Adamts5 and P16 proteins in chondrocytes. ( E – J ) qRT-PCR was performed to detect the expression of Col2, Sox9, MMP13, Adamts5, P16 and P21 in chondrocytes. ( K ) Cellular senescence was assessed by SA-β-gal staining. All data represent mean ± SD. Compared to the control group, **P < 0.01, ***P < 0.001, ****P < 0.0001; compared to the H 2 O 2 -stimulated group, *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001; compared to the same concentration of blank serum group, *P < 0.05, **P < 0.01, ****P < 0.0001.

Article Snippet: Primary antibodies against CDKN2A/p16INK4a (1:100) were purchased from Zenbio (Chengdu, China), and the primary antibody against Adamts5 (1:50) was acquired from HUABIO (Hangzhou, China).

Techniques: Western Blot, Expressing, Quantitative RT-PCR, Staining, Concentration Assay

Journal: Aging (Albany NY)

Article Title: Lei’s formula attenuates osteoarthritis mediated by suppression of chondrocyte senescence via the mTOR axis: in vitro and in vivo experiments

doi: 10.18632/aging.205582

Figure Lengend Snippet: Activation of mTOR exacerbates H 2 O 2 -stimulated chondrocyte senescence and anabolic catabolic disorders that can be reversed by LSF. ( A ) Western blotting for protein expression of Col2, Adamts5, P16 and mTOR in chondrocytes processed with H 2 O 2 (200 μmol/L) and MHY1485 (10 μmol/L). ( B – E ) Quantitative results of Western blotting of the 5 groups of proteins. ( F – M ) Expression of Col2, Sox9, MMP13, Adamts5, P16, P21, mTOR, and S6 in chondrocytes processed with H 2 O 2 (200 μmol/L) and MHY1485 (10 μmol/L) was detected by qRT-PCR. ( N ) Cellular senescence was assessed by SA-β-gal staining. Compared to the control group, *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001; compared to the H 2 O 2 -stimulated group, *P <0.05, *P < 0.01, ***P < 0.001, ****P<0.0001; compared to the H 2 O 2 -stimulated combined MHY1485 group, *P < 0.05, **P < 0.01, ***P <0.001, ****P < 0.0001.

Article Snippet: Primary antibodies against CDKN2A/p16INK4a (1:100) were purchased from Zenbio (Chengdu, China), and the primary antibody against Adamts5 (1:50) was acquired from HUABIO (Hangzhou, China).

Techniques: Activation Assay, Western Blot, Expressing, Quantitative RT-PCR, Staining

Journal: Aging (Albany NY)

Article Title: Lei’s formula attenuates osteoarthritis mediated by suppression of chondrocyte senescence via the mTOR axis: in vitro and in vivo experiments

doi: 10.18632/aging.205582

Figure Lengend Snippet: mTOR-SiRNA contributes to LSF suppression of H 2 O 2 -stimulated chondrocyte senescence and promotion of anabolism. ( A ) Westen blotting for protein expression of Col2, Adamts5, P16 and mTOR in lentivirus-transfected chondrocytes stimulated by H2O2 (200 μmol/L). ( B – E ) Quantitative results of Western blotting of the 4 groups of proteins. ( F – M ) Expression of Col2, Sox9, MMP13, Adamts5, P16, P21, mTOR, and S6 in lentivirus-transfected chondrocytes stimulated with H2O2 (200 μmol/L) was detected by qRT-PCR. ( N ) Cellular senescence was assessed by SA-β-gal staining. Compared to Con-SiRNA control group, ****P < 0.0001; compared to the H 2 O 2 -stimulated group, *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001; compared to the mTOR-SiRNA group, *P < 0.05, **P < 0.01, ***P<0.001, ****P < 0.0001.

Article Snippet: Primary antibodies against CDKN2A/p16INK4a (1:100) were purchased from Zenbio (Chengdu, China), and the primary antibody against Adamts5 (1:50) was acquired from HUABIO (Hangzhou, China).

Techniques: Expressing, Transfection, Western Blot, Quantitative RT-PCR, Staining

Journal: Aging (Albany NY)

Article Title: Lei’s formula attenuates osteoarthritis mediated by suppression of chondrocyte senescence via the mTOR axis: in vitro and in vivo experiments

doi: 10.18632/aging.205582

Figure Lengend Snippet: LSF ameliorates the progression of OA and inhibits mTOR expression in the mice DMM model. ( A ) Immunohisto- chemical typical images of pathological sections of mice knee joints from different experimental groups (SHAM, DMM, DMM+LSF-L (0.08 g/kg), DMM+LSF-M (0.17 g/kg), and DMM+LSF-H (0.35 g/kg)), which were examined for the detection of Col2, ACAN, Adamts5, MMP13, Col10, P16, P21 and mTOR expression in mice cartilage. (Scale bar. 50 μm). ( B – I ) Quantitative results of immunohistochemical staining in 5 groups of mice. All data represent mean ± SD. compared to the SHAM group; ****P < 0.0001; compared to the DMM group, *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001.

Article Snippet: Primary antibodies against CDKN2A/p16INK4a (1:100) were purchased from Zenbio (Chengdu, China), and the primary antibody against Adamts5 (1:50) was acquired from HUABIO (Hangzhou, China).

Techniques: Expressing, Immunohistochemical staining, Staining

Journal: Yonsei Medical Journal

Article Title: LncRNA MALAT1/MiR-145 Adjusts IL-1β-Induced Chondrocytes Viability and Cartilage Matrix Degradation by Regulating ADAMTS5 in Human Osteoarthritis

doi: 10.3349/ymj.2019.60.11.1081

Figure Lengend Snippet: MALAT1 modulates IL-1β-induced chondrocytes cell viability and cartilage ECM degradation via miR-145 in IL-1β-induced chondrocytes. (A and B) Chondrocytes were treated with different concentrations of IL-1β for 24 h, and expression of MALAT1 and miR-145 was detected using qRT-PCR. (C and D) Cell viability was measured by MTT assay in IL-1β-induced chondrocytes when transfected with si-MALAT1 #1 and MALAT1 vector only or combined with miR-145 mimics. (E) Expression of ECM-related proteins was examined by Western blotting in chondrocytes transfected with si-MALAT1 #1 and MALAT1 vector. (F and G) Quantitative analysis of ADAMTS5, COL2A1, ACAN, and COMP expressions using Image J. (H and I) Expression of ADAMTS5, COL2A1, ACAN, and COMP in IL-1β-induced chondrocytes transfected with MALAT1 vector only or combined with miR-145 mimics. All data are presented as means±SD. * p <0.05. ECM, extracellular matrix.

Article Snippet: Then, membranes were incubated with primary antibody against ADAMTS5 (1:200 dilution; Sigma-Aldrich), COL2A1 (1:2500 dilution, Abcam, Cambridge, UK), ACAN (1:200 dilution, Abcam), COMP (1:200 dilution, Abcam), GAPDH (1:5000; Cell Signaling Technology, Danvers, MA, USA) overnight at 4℃.

Techniques: Expressing, Quantitative RT-PCR, MTT Assay, Transfection, Plasmid Preparation, Western Blot

Journal: Yonsei Medical Journal

Article Title: LncRNA MALAT1/MiR-145 Adjusts IL-1β-Induced Chondrocytes Viability and Cartilage Matrix Degradation by Regulating ADAMTS5 in Human Osteoarthritis

doi: 10.3349/ymj.2019.60.11.1081

Figure Lengend Snippet: Correlation analysis between expression levels of ADAMTS5, MALAT1, and miR-145. Spearman rank analysis revealed (A) a negative correlation between ADAMTS5 and miR-145 expression and (B) a positive correlation between ADAMTS5 and MALAT1 expression.

Article Snippet: Then, membranes were incubated with primary antibody against ADAMTS5 (1:200 dilution; Sigma-Aldrich), COL2A1 (1:2500 dilution, Abcam, Cambridge, UK), ACAN (1:200 dilution, Abcam), COMP (1:200 dilution, Abcam), GAPDH (1:5000; Cell Signaling Technology, Danvers, MA, USA) overnight at 4℃.

Techniques: Expressing

Journal: Yonsei Medical Journal

Article Title: LncRNA MALAT1/MiR-145 Adjusts IL-1β-Induced Chondrocytes Viability and Cartilage Matrix Degradation by Regulating ADAMTS5 in Human Osteoarthritis

doi: 10.3349/ymj.2019.60.11.1081

Figure Lengend Snippet: miR-145 might sequester ADAMTS5 by target binding. (A) The databases TargetScan ( http://www.targetscan.org/ ) and miRcode ( http://www.mircode.org/ ) showed two conserved binding sites of miR-145 in the 3′ UTR of ADAMTS5. The wild-type of ADAMTS5 (ADAMTS5-WT) was mutated as MALAT1-MUT. (B, C) Relative luciferase activity of ADAMTS5-WT/MUT was determined by luciferase reporter assay in chondrocytes when co-transfected with miR-145 mimics or inhibitors. (D, E) Western blotting detected ADAMTS5 protein expression level after chondrocytes transfected with miR-145 mimics or inhibitors. (F, G) qRT-PCR detected ADAMTS5 mRNA expression level after chondrocytes transfected with miR-145 mimics or inhibitors. Data are presented as means±SD. * p <0.05.

Article Snippet: Then, membranes were incubated with primary antibody against ADAMTS5 (1:200 dilution; Sigma-Aldrich), COL2A1 (1:2500 dilution, Abcam, Cambridge, UK), ACAN (1:200 dilution, Abcam), COMP (1:200 dilution, Abcam), GAPDH (1:5000; Cell Signaling Technology, Danvers, MA, USA) overnight at 4℃.

Techniques: Binding Assay, Luciferase, Activity Assay, Reporter Assay, Transfection, Western Blot, Expressing, Quantitative RT-PCR

Journal: Yonsei Medical Journal

Article Title: LncRNA MALAT1/MiR-145 Adjusts IL-1β-Induced Chondrocytes Viability and Cartilage Matrix Degradation by Regulating ADAMTS5 in Human Osteoarthritis

doi: 10.3349/ymj.2019.60.11.1081

Figure Lengend Snippet: ADAMTS5 modulates IL-1β-induced chondrocyte viability and cartilage degradation mediated by MALAT1/miR-145. Chondrocytes were transfected with MALAT1 only or together with miR-145 mimics and miR-145 mimics only or combined with ADAMTS5 vector. After 10 ng/mL of IL-1β treatment, (A) cell viability was detected by MTT assay at 12 h, 24 h, 48 h, and 72 h, and (B) ECM-related protein expression was measured by Western blotting at 24 h. (C and D) Quantitative analysis of ADAMTS5, COL2A1, ACAN, and COMP expression was conducted by Image J. All data are presented as means±SD. * p <0.05.

Article Snippet: Then, membranes were incubated with primary antibody against ADAMTS5 (1:200 dilution; Sigma-Aldrich), COL2A1 (1:2500 dilution, Abcam, Cambridge, UK), ACAN (1:200 dilution, Abcam), COMP (1:200 dilution, Abcam), GAPDH (1:5000; Cell Signaling Technology, Danvers, MA, USA) overnight at 4℃.

Techniques: Transfection, Plasmid Preparation, MTT Assay, Expressing, Western Blot