Journal: Journal of Cancer

Article Title: Development and Validation of a Serum Biomarker Panel for the Detection of Esophageal Squamous Cell Carcinoma through RNA Transcriptome Sequencing

doi: 10.7150/jca.19465

Figure Lengend Snippet: Serum levels of candidate biomarkers in the preliminary screening phase. Levels of serum ADAM12, CHI3L1, MMP13 and SPP1 were compared between 40 ESCC patients (ESCC) and 40 healthy controls (HC). The Mann-Whitney U test was performed for comparisons between groups. P < 0.05 was considered statistically significant.

Article Snippet: Immunohistochemistry was performed using standard protocols with antibodies against CHI3L1 (1:50; Abcam, UK), MMP13 (1:100; R&D systems, USA), and SPP1 (1:100; Abcam, UK).

Techniques: MANN-WHITNEY

Journal: Journal of Cancer

Article Title: Development and Validation of a Serum Biomarker Panel for the Detection of Esophageal Squamous Cell Carcinoma through RNA Transcriptome Sequencing

doi: 10.7150/jca.19465

Figure Lengend Snippet: Serum ADAM12, CHI3L1, MMP13 and SPP1 in the test cohort and the validation cohort. A: Levels of biomarkers were compared between 150 ESCC patients (ESCC), 140 controls (96 healthy controls and 44 patients with benign esophageal disease) and 71 early-stage ESCC patients. Statistical analyses were performed using the Mann-Whitney U test. B: ROC curves for biomarkers and their combination (Logit(p=ESCC)=-4.583+0.017×CHI3L1+0.018×SPP1+0.821×MMP13) for the discrimination of 150 patients with ESCC and 140 controls. C: ROC curves for biomarkers and their combination for the discrimination of 71 patients with early-stage ESCC and 140 controls. D: Levels of biomarkers were compared in 169 ESCC patients (ESCC), 154 controls (80 healthy controls and 74 patients with benign esophageal disease) and 84 early-stage ESCC patients. Statistical analyses were performed using the Mann-Whitney U test. E: ROC curves for biomarkers and their combination for the discrimination of 169 patients with ESCC and 154 controls. F: ROC curves for biomarkers and their combination (Logit(p=ESCC)=-4.583+0.017×CHI3L1+0.018×SPP1+0.821×MMP13) for the discrimination of 84 patients with early-stage ESCC and 154 controls. P < 0.05 was considered statistically significant.

Article Snippet: Immunohistochemistry was performed using standard protocols with antibodies against CHI3L1 (1:50; Abcam, UK), MMP13 (1:100; R&D systems, USA), and SPP1 (1:100; Abcam, UK).

Techniques: Biomarker Discovery, MANN-WHITNEY

Journal: Journal of Cancer

Article Title: Development and Validation of a Serum Biomarker Panel for the Detection of Esophageal Squamous Cell Carcinoma through RNA Transcriptome Sequencing

doi: 10.7150/jca.19465

Figure Lengend Snippet: The diagnostic performance of ADAM12, CHI3L1, MMP13, SPP1 and their combination (Logit(p=ESCC)=-4.583+0.017×CHI3L1+0.018×SPP1+0.821×MMP13) in discriminating ESCC, early stage ESCC and controls (healthy controls and patients with esophageal benign disease) in the test cohort and validation cohort.

Article Snippet: Immunohistochemistry was performed using standard protocols with antibodies against CHI3L1 (1:50; Abcam, UK), MMP13 (1:100; R&D systems, USA), and SPP1 (1:100; Abcam, UK).

Techniques: Diagnostic Assay, Biomarker Discovery

Journal: Journal of Cancer

Article Title: Development and Validation of a Serum Biomarker Panel for the Detection of Esophageal Squamous Cell Carcinoma through RNA Transcriptome Sequencing

doi: 10.7150/jca.19465

Figure Lengend Snippet: Expression of CHI3L1, MMP13 and SPP1 mRNA or protein in ESCC cell lines and tissues and their locations in tissue. The mRNA and protein expression levels in six pairs of matched ESCC and noncancerous tissues was analyzed by real-time PCR and Western blotting, respectively (A, B), and in an immortalized esophageal epithelial cell line (NE-3) and esophageal carcinoma cell lines by real-time PCR and ELISA, respectively (C, D). The expression level was normalized to the expression of GAPDH and α-tubulin, respectively. Error bars represent standard deviations (SD) calculated from three parallel experiments. The locations of CHI3L1, MMP13 and SPP1 were determined by immunohistochemistry (E). The normal esophageal epithelial tissue showed no expression of CHI3L1, MMP13 or SPP1, respectively (E a, e, i, 200 × and b, f, j, 400×). The ESCC tissues exhibited high expression levels of these three biomarkers (E c, f, k, 200 × and d, h, l, 400×).

Article Snippet: Immunohistochemistry was performed using standard protocols with antibodies against CHI3L1 (1:50; Abcam, UK), MMP13 (1:100; R&D systems, USA), and SPP1 (1:100; Abcam, UK).

Techniques: Expressing, Real-time Polymerase Chain Reaction, Western Blot, Enzyme-linked Immunosorbent Assay, Immunohistochemistry

Journal: Journal of Orthopaedic Translation

Article Title: Cyclooxygenase-2 regulates PTHrP transcription in human articular chondrocytes and is involved in the pathophysiology of osteoarthritis in rats

doi: 10.1016/j.jot.2021.06.003

Figure Lengend Snippet: Primer sequences used for real-time polymerase chain reaction.

Article Snippet: Sections were then blocked with 10% FBS for 1 ​h and incubated with primary antibodies against type II collagen (1:2000; mouse monoclonal antibody; Chemi-Con, Temecula, CA), type X collagen (rat polyclonal antibody) (1:200; Cosmo Bio, Tokyo, Japan), COX-2 (rabbit polyclonal antibody) (1:50; Santa Cruz, Europe), PTHrP (rabbit polyclonal antibody) (1:100; OriGene, MD, USA) and MMP13 (1:200; rabbit polyclonal antibody) (1:100; Bioss, MA, USA) at 4 ​°C overnight.

Techniques:

Journal: Journal of Orthopaedic Translation

Article Title: Cyclooxygenase-2 regulates PTHrP transcription in human articular chondrocytes and is involved in the pathophysiology of osteoarthritis in rats

doi: 10.1016/j.jot.2021.06.003

Figure Lengend Snippet: Celecoxib reduces type X collagen, MMP13 and PTHrP levels and inhibits type II collagen expression in osteoarthritic articular cartilage (A) Immunohistochemical analysis of type X collagen (scale bars: 100 ​μm) (B) MMP13 (scale bars: 200 ​μm) (C) type II collagen, and (D) PTHrP (scale bars: 100 ​μm) in articular cartilage of tibia sections in the OA ​+ ​Saline and OA ​+ ​Celec groups. The histogram illustrates the quantification of the positively stained type X collagen and the relative density of MMP13, type II collagen and PTHrP expression in articular cartilage. Each column represents the mean ​± ​SEM of six samples. Data from each group were compared with those of the OA contralateral control group and were evaluated by one-way ANOVA and Scheffe's test. ∗, P ​< ​0.05; ∗∗, P ​< ​0.01; the OA ​+ ​Celec groups compared to the OA ​+ ​Saline groups # , P ​< ​0.05; ## , P ​< ​0.01.

Article Snippet: Sections were then blocked with 10% FBS for 1 ​h and incubated with primary antibodies against type II collagen (1:2000; mouse monoclonal antibody; Chemi-Con, Temecula, CA), type X collagen (rat polyclonal antibody) (1:200; Cosmo Bio, Tokyo, Japan), COX-2 (rabbit polyclonal antibody) (1:50; Santa Cruz, Europe), PTHrP (rabbit polyclonal antibody) (1:100; OriGene, MD, USA) and MMP13 (1:200; rabbit polyclonal antibody) (1:100; Bioss, MA, USA) at 4 ​°C overnight.

Techniques: Expressing, Immunohistochemical staining, Staining

Journal: Journal of Orthopaedic Translation

Article Title: Cyclooxygenase-2 regulates PTHrP transcription in human articular chondrocytes and is involved in the pathophysiology of osteoarthritis in rats

doi: 10.1016/j.jot.2021.06.003

Figure Lengend Snippet: Overexpression of COX-2 increases Col2a1 , Col10a1 , PTHrP , MMP13 and Runx2 transcription (A) Col2a1 (B) Col10a1 (C) PTHrP (D) Ihh (E) MMP13, and (F) Runx2 mRNA expression was measured in vitro in cells transfected with pcDNA3.1-COX-2 and the control pcDNA3-empty vector for 1 and 3 days. GAPDH mRNA was used as the internal control. Each column represents the mean ​± ​SEM of four replicate cultures. Data from each group were compared with those of the control group and were evaluated by one-way ANOVA and Scheffe's test. ∗, P ​< ​0.05; ∗∗, P ​< ​0.01; ∗∗∗, P ​< ​0.001 compared to the control culture.

Article Snippet: Sections were then blocked with 10% FBS for 1 ​h and incubated with primary antibodies against type II collagen (1:2000; mouse monoclonal antibody; Chemi-Con, Temecula, CA), type X collagen (rat polyclonal antibody) (1:200; Cosmo Bio, Tokyo, Japan), COX-2 (rabbit polyclonal antibody) (1:50; Santa Cruz, Europe), PTHrP (rabbit polyclonal antibody) (1:100; OriGene, MD, USA) and MMP13 (1:200; rabbit polyclonal antibody) (1:100; Bioss, MA, USA) at 4 ​°C overnight.

Techniques: Over Expression, Expressing, In Vitro, Transfection, Plasmid Preparation

Journal: Experimental and Therapeutic Medicine

Article Title: The anti-inflammation effect of strontium ranelate on rat chondrocytes with or without IL-1β in vitro

doi: 10.3892/etm.2022.11131

Figure Lengend Snippet: Toluidine blue staining. (A) Hyp concentration and (B) relative gene expression. Relative gene expression of (C) aggrecan, (D) Col-II, (E) β-catenin, (F) MMP-9 (G) and MMP-13 of rat chondrocytes treated with different concentrations of SrR. The images of 2A were captured under x200 magnification. * P<0.05. Hyp, hydroxyproline; Col, collagen; SrR, strontium ranelate.

Article Snippet: Following blocking with 5% skimmed milk at room temperature for 1 h, the membrane was exposed to primary antibodies at 4˚C overnight, including Col-II (1:500; cat. no. 28459-1-AP; ProteinTech Group, Inc.), aggrecan (1:500; cat. no. 13880-1; ProteinTech Group, Inc.), β-catenin (1:1,000; cat. no. ab16051; Abcam), MMP-9 (1:500; cat. no. 10375-2; ProteinTech Group, Inc.), MMP-13 (1:500; cat. no. Nbp2-17310; Novus Biologicals) and β-actin (1:1,000; cat. no. 4970; Cell Signaling Technology, Inc.).

Techniques: Staining, Concentration Assay, Expressing

Journal: Experimental and Therapeutic Medicine

Article Title: The anti-inflammation effect of strontium ranelate on rat chondrocytes with or without IL-1β in vitro

doi: 10.3892/etm.2022.11131

Figure Lengend Snippet: Immunofluorescence staining assay of (A) β-catenin, (B) Col-II and (C) MMP-13. (D) Western blot assay of Col-II, aggrecan, β-catenin, MMP-13, MMP-9 and β-actin and (E) relative protein expression. The images were captured under x200 magnification, save for the right hand column of (A). Arrows indicate the position of β-catenin. * P<0.05 as indicated. Col, collagen.

Article Snippet: Following blocking with 5% skimmed milk at room temperature for 1 h, the membrane was exposed to primary antibodies at 4˚C overnight, including Col-II (1:500; cat. no. 28459-1-AP; ProteinTech Group, Inc.), aggrecan (1:500; cat. no. 13880-1; ProteinTech Group, Inc.), β-catenin (1:1,000; cat. no. ab16051; Abcam), MMP-9 (1:500; cat. no. 10375-2; ProteinTech Group, Inc.), MMP-13 (1:500; cat. no. Nbp2-17310; Novus Biologicals) and β-actin (1:1,000; cat. no. 4970; Cell Signaling Technology, Inc.).

Techniques: Immunofluorescence, Staining, Western Blot, Expressing

Journal: Experimental and Therapeutic Medicine

Article Title: The anti-inflammation effect of strontium ranelate on rat chondrocytes with or without IL-1β in vitro

doi: 10.3892/etm.2022.11131

Figure Lengend Snippet: Chondrocytes respond to IL-1β and SrR. (A) Toluidine blue staining and (B), relative gene expression of β-catenin. Relative gene expression of (C) Col-II, (D) MMP-9 and (E) MMP-13. (F) Western blotting analysis and (G) relative protein expression of chondrocytes treated with 10 ng/l IL-1β and 0 or 0.50 mmol/l SrR for three days. The images were captured under x200 magnification. * P<0.05. Col, collagen; SrR, strontium ranelate; d, day.

Article Snippet: Following blocking with 5% skimmed milk at room temperature for 1 h, the membrane was exposed to primary antibodies at 4˚C overnight, including Col-II (1:500; cat. no. 28459-1-AP; ProteinTech Group, Inc.), aggrecan (1:500; cat. no. 13880-1; ProteinTech Group, Inc.), β-catenin (1:1,000; cat. no. ab16051; Abcam), MMP-9 (1:500; cat. no. 10375-2; ProteinTech Group, Inc.), MMP-13 (1:500; cat. no. Nbp2-17310; Novus Biologicals) and β-actin (1:1,000; cat. no. 4970; Cell Signaling Technology, Inc.).

Techniques: Staining, Expressing, Western Blot

Journal: Experimental and Therapeutic Medicine

Article Title: The anti-inflammation effect of strontium ranelate on rat chondrocytes with or without IL-1β in vitro

doi: 10.3892/etm.2022.11131

Figure Lengend Snippet: Immunofluorescence staining assay of (A) β-catenin, (B) Col-II and (C) MMP-13 following three days of induction with IL-1β and SrR. Arrow showed the position of β-catenin. The images were captured under x200 magnification, save for the right hand column of (A). Arrows indicate the position of β-catenin. Col, collagen; SrR, strontium ranelate.

Article Snippet: Following blocking with 5% skimmed milk at room temperature for 1 h, the membrane was exposed to primary antibodies at 4˚C overnight, including Col-II (1:500; cat. no. 28459-1-AP; ProteinTech Group, Inc.), aggrecan (1:500; cat. no. 13880-1; ProteinTech Group, Inc.), β-catenin (1:1,000; cat. no. ab16051; Abcam), MMP-9 (1:500; cat. no. 10375-2; ProteinTech Group, Inc.), MMP-13 (1:500; cat. no. Nbp2-17310; Novus Biologicals) and β-actin (1:1,000; cat. no. 4970; Cell Signaling Technology, Inc.).

Techniques: Immunofluorescence, Staining

Journal: The journal of gene medicine

Article Title: Sp2 Transcription Factor Alleviates Chondrocyte Loss in Osteoarthritis by Repressing the DVL1-Dependent Wnt/β-Catenin Signaling Pathway.

doi: 10.1002/jgm.70021

Figure Lengend Snippet: FIGURE 3 | DVL1 knockdown ameliorates cartilage injury in mice. (A) Protein levels of COL10A1, MMP13, and SOX9 in mouse cartilage tissues determined using WB analysis. (B) Cartilage morphology in the mouse knee joints determined using Safranin O/fast green staining. (C) Positive TRAP staining in the mouse knee joints. (D) Apoptosis in the extracted chondrocytes determined using TUNEL assay. (E) Protein levels of pro- cleaved-caspase-3 in the extracted chondrocytes determined using WB analysis. (F) Protein levels of COL10A1, MMP13, and SOX9 in the mouse chondrocytes determined using WB analysis. For animal experiments, each group contained five mice. For cell experiments, three biological repli- cates were performed. Differences were compared by the unpaired t-test (D–F) or ANOVA (A–C).

Article Snippet: The membranes were blocked at room temperature for 1 h in Tris- buffered saline (T5912, Merck) and incubated with antibodies against DVL1 (1:1000, 13- 706, ProSci), COL10A1 (collagen type X alpha 1 chain) (1:1000, A11645, ABclonal Technology Co. Ltd., Wuhan, Hubei, China), MMP13 (matrix metallopeptidase 13) (1:4000, ab39012, Abcam), SOX9 (SRY- box transcription factor 9) (1:1000, orb1258160, Biorbyt LLC, San Francisco, California, United States), cleavedcaspase- 3 (1:1000, PA5- 114687, Thermo Fisher Scientific), βcatenin (1:5000, ab32572, Abcam), COL2A1 (collagen type II alpha 1 chain) (1:1000, orb1259434, Biorbyt), SP2 (1:1000, PA5- 103254, Thermo Fisher Scientific), and GAPDH (1:2500, ab9485, Abcam) overnight at 4°C.

Techniques: Knockdown, Staining, TUNEL Assay

Journal: The journal of gene medicine

Article Title: Sp2 Transcription Factor Alleviates Chondrocyte Loss in Osteoarthritis by Repressing the DVL1-Dependent Wnt/β-Catenin Signaling Pathway.

doi: 10.1002/jgm.70021

Figure Lengend Snippet: FIGURE 4 | CHIR-99021 restores cartilage injury mitigated by DVL1 silencing. Mice stably administered sh-DVL1 were further treated with the Wnt/β-catenin agonist CHIR-99021 via intra-articular injection. (A) Protein levels of β-catenin in cells determined using WB analysis. (B) Cartilage morphology in the mouse knee joints determined using Safranin O/fast green staining. (C) Protein levels of COL10A1 and COL2A1 in mouse chon- drocytes determined using WB analysis. (D) Positive TRAP staining in the mouse knee joints; in vitro, chondrocytes with stable DVL1 knockdown were treated with 5 μM CHIR-99021 for 24 h. (E) Apoptosis in the extracted chondrocytes determined using TUNEL assay. (F) Protein levels of pro- cleaved-caspase-3 in the extracted chondrocytes determined using WB analysis. (G) Protein levels of expression of MMP13 and SOX9 in the mouse chondrocytes determined using WB analysis. For animal experiments, each group contained five mice. For cell experiments, three biological repli- cates were performed. Differences were compared by the unpaired t-test (A–G).

Article Snippet: The membranes were blocked at room temperature for 1 h in Tris- buffered saline (T5912, Merck) and incubated with antibodies against DVL1 (1:1000, 13- 706, ProSci), COL10A1 (collagen type X alpha 1 chain) (1:1000, A11645, ABclonal Technology Co. Ltd., Wuhan, Hubei, China), MMP13 (matrix metallopeptidase 13) (1:4000, ab39012, Abcam), SOX9 (SRY- box transcription factor 9) (1:1000, orb1258160, Biorbyt LLC, San Francisco, California, United States), cleavedcaspase- 3 (1:1000, PA5- 114687, Thermo Fisher Scientific), βcatenin (1:5000, ab32572, Abcam), COL2A1 (collagen type II alpha 1 chain) (1:1000, orb1259434, Biorbyt), SP2 (1:1000, PA5- 103254, Thermo Fisher Scientific), and GAPDH (1:2500, ab9485, Abcam) overnight at 4°C.

Techniques: Stable Transfection, Injection, Staining, In Vitro, Knockdown, TUNEL Assay, Expressing

Journal: The journal of gene medicine

Article Title: Sp2 Transcription Factor Alleviates Chondrocyte Loss in Osteoarthritis by Repressing the DVL1-Dependent Wnt/β-Catenin Signaling Pathway.

doi: 10.1002/jgm.70021

Figure Lengend Snippet: FIGURE 6 | DVL1 upregulation aggravates cartilage injury in mice alleviated by SP2. DMM-challenged mice were introduced with adenoviral vectors carrying OE-NC/OE-SP2 alone or with the additional OE-NC/OE-DVL1. (A) mRNA expression of SP2 and DVL1 in the joint cartilage deter- mined using RT-qPCR. (B) Positive staining of SP2 and DVL1 in the joint cartilage determined using IHC. (C) Protein levels of β-catenin in the joint cartilage determined using WB analysis. (D) Cartilage morphology in the mouse knee joints determined using Safranin O/fast green staining. (E) Positive TRAP staining of the mouse knee joints. (F) Protein levels of COL10A1 and MMP13 in the mouse cartilage determined using WB analysis. Each group contained five mice. Differences were compared by ANOVA (A–F).

Article Snippet: The membranes were blocked at room temperature for 1 h in Tris- buffered saline (T5912, Merck) and incubated with antibodies against DVL1 (1:1000, 13- 706, ProSci), COL10A1 (collagen type X alpha 1 chain) (1:1000, A11645, ABclonal Technology Co. Ltd., Wuhan, Hubei, China), MMP13 (matrix metallopeptidase 13) (1:4000, ab39012, Abcam), SOX9 (SRY- box transcription factor 9) (1:1000, orb1258160, Biorbyt LLC, San Francisco, California, United States), cleavedcaspase- 3 (1:1000, PA5- 114687, Thermo Fisher Scientific), βcatenin (1:5000, ab32572, Abcam), COL2A1 (collagen type II alpha 1 chain) (1:1000, orb1259434, Biorbyt), SP2 (1:1000, PA5- 103254, Thermo Fisher Scientific), and GAPDH (1:2500, ab9485, Abcam) overnight at 4°C.

Techniques: Expressing, Quantitative RT-PCR, Staining