Journal: Materials Today Bio

Article Title: Cuprorivaite microspheres inhibit cuproptosis and oxidative stress in osteoarthritis via Wnt/β-catenin pathway

doi: 10.1016/j.mtbio.2024.101300

Figure Lengend Snippet: Anti-inflammatory role of Cuprorivaite (CaCuSi 4 O 10 ) microspheres in IL-1β-stimulated chondrocytes. (A) Cell counting kit-8 test for cell proliferation. (B) Fluorescein diacetate for cell viability detection. (C) Cell apoptosis by flow cytometry. (D) ELISA detection for TNF-α, IL-6 and MMP13 in cell supernatant. (E) Western blot detection for TNF-α, IL-6 and MMP13 in cell lysates. (F) RT-qPCR quantification for in TNF-α, IL-6 and MMP13. ∗ P < 0.05, ∗∗ P < 0.01, ∗∗∗ P < 0.001.

Article Snippet: Then, levels of TNF-α, IL-6, and MMP13 in cell supernatant and serum samples were detected by the multi-functional microplate detector, with mouse TNF-α (CSB-E04741m, CUSABIO, Wuhan, China), mouse IL-6 (CSB-E04639m, CUSABIO, Wuhan, China) and mouse MMP13 (CSB-E07413m, CUSABIO, Wuhan, China) kit.

Techniques: Cell Counting, Flow Cytometry, Enzyme-linked Immunosorbent Assay, Western Blot, Quantitative RT-PCR

Journal: Materials Today Bio

Article Title: Cuprorivaite microspheres inhibit cuproptosis and oxidative stress in osteoarthritis via Wnt/β-catenin pathway

doi: 10.1016/j.mtbio.2024.101300

Figure Lengend Snippet: Cuprorivaite (CaCuSi 4 O 10 ) microspheres improved IL-1β-induced injury in chondrocytes via inhibiting Wnt/β-catenin pathway. (A) Cell counting kit-8 test for cell viability. (B) Fluorescein diacetate for cell viability detection. (C) Cell apoptosis by flow cytometry. (D) ELISA detection for TNF-α, IL-6 and MMP13 in cell supernatant. (E) Western blot detection for TNF-α, IL-6 and MMP13 in cell lysates. (F) RT-qPCR quantification for extracellular matrix components including collagen II and SOX9. (G) Intracellular copper content. (H) RT-qPCR quantification for cuproptosis biomarkers including ATP7B and FDX1. (I) Western blot detection for ATP7B and FDX1. (J) Oxidative stress detection including MDA, SOD and GSH. (K) RT-qPCR quantification for Wnt1, GSK3β and β-catenin. ∗ P < 0.05, ∗∗ P < 0.01, ∗∗∗ P < 0.001.

Article Snippet: Then, levels of TNF-α, IL-6, and MMP13 in cell supernatant and serum samples were detected by the multi-functional microplate detector, with mouse TNF-α (CSB-E04741m, CUSABIO, Wuhan, China), mouse IL-6 (CSB-E04639m, CUSABIO, Wuhan, China) and mouse MMP13 (CSB-E07413m, CUSABIO, Wuhan, China) kit.

Techniques: Cell Counting, Flow Cytometry, Enzyme-linked Immunosorbent Assay, Western Blot, Quantitative RT-PCR

Journal: Materials Today Bio

Article Title: Cuprorivaite microspheres inhibit cuproptosis and oxidative stress in osteoarthritis via Wnt/β-catenin pathway

doi: 10.1016/j.mtbio.2024.101300

Figure Lengend Snippet: Validation of mechanism of Cuprorivaite (CaCuSi 4 O 10 ) microspheres in OA. (A) Hematoxylin and eosin staining. (B) Safranin-O staining/fast green staining. (C) OARSI score. (D) ELISA detection for TNF-α, IL-6 and MMP13 in the serum. (E) Western blot detection for TNF-α, IL-6 and MMP13 in cartilage tissue. (F) RT-qPCR quantification for extracellular matrix components including collagen II and SOX9 in cartilage tissue. (G) Copper content in cartilage tissue. (H) RT-qPCR quantification for cuproptosis biomarkers including ATP7B and FDX1 in cartilage tissue. (I) Western blot detection for ATP7B and FDX1 in cartilage tissue. (J) Representative images of FDX1 expression in cartilage tissue detected by immunohistochemistry. (K) Oxidative stress detection including MDA, SOD and GSH in cartilage tissue. (L) RT-qPCR quantification for Wnt1, GSK3β and β-catenin in cartilage tissue. ∗∗∗ P < 0.001.

Article Snippet: Then, levels of TNF-α, IL-6, and MMP13 in cell supernatant and serum samples were detected by the multi-functional microplate detector, with mouse TNF-α (CSB-E04741m, CUSABIO, Wuhan, China), mouse IL-6 (CSB-E04639m, CUSABIO, Wuhan, China) and mouse MMP13 (CSB-E07413m, CUSABIO, Wuhan, China) kit.

Techniques: Biomarker Discovery, Staining, Enzyme-linked Immunosorbent Assay, Western Blot, Quantitative RT-PCR, Expressing, Immunohistochemistry

Journal: Antioxidants

Article Title: Matrix Metalloproteinase 13 Is Associated with Age-Related Choroidal Neovascularization

doi: 10.3390/antiox12040884

Figure Lengend Snippet: MMP13 localization and overexpression in ARPE-19 cells after oxidative stress induction. ( A ) Immunofluorescence of precursor and active MMP13 under basal conditions with 1% saline and under oxidative stress conditions (600 µM H 2 O 2 for 2 h). The corresponding orthogonal view of the obtained z-stack is shown next to it. A significant increase in MMP13 expression is observed when subjected to oxidative stress. Mann–Whitney U; * p < 0.05 (n = 3). Scale bar: 10 µm. Nuclei are labeled with DAPI (blue). ( B ) WB analysis of MMP13 in ARPE-19 cells exposed to H 2 O 2 600 μM at different times (6, 24, 30, and 48 h) vs. saline control 6 h. Ponceau S red is shown below the WB image and confirms the equal loading for each sample. The semi-quantitative histograms of the WB analyses show that MMP13 is expressed under basal conditions over time and that H 2 O 2 exposition increases its expression. One-way ANOVA followed by Bonferroni. * p < 0.05, *** p < 0.001, and **** p < 0.0001. Bars represent percentage vs. saline 6 h ± S.D. (n = 3). CNV = choroidal neovascularization. MMP13 = matrix metalloproteinase 13. Western blot = WB. MW= molecular weight.

Article Snippet: They were then exposed for 3 days at RT to mouse monoclonal anti-MMP13 (for precursor and active forms; 1:100, MAB3321, Merck), biotinylated isolectin (1:240, B-1205; Vector Labs, Burlingame, CA, USA), and a goat collagen IV antibody (1:250, 1340-01, Southern Biotech, CA, USA).

Techniques: Over Expression, Immunofluorescence, Expressing, MANN-WHITNEY, Labeling, Western Blot, Molecular Weight

Journal: Antioxidants

Article Title: Matrix Metalloproteinase 13 Is Associated with Age-Related Choroidal Neovascularization

doi: 10.3390/antiox12040884

Figure Lengend Snippet: MMP13 characterization in mice posterior pole using immunofluorescence. While no MMP13 expression was observed in the RPE, choroid, or sclera (upper row), the retinal blood vessels on the INL (deep capillary plexus, middle row) and the GCL (superficial capillary plexus, bottom row) showed MMP13 expression, and some lectin-negative cells expressed MMP13. The corresponding orthogonal view of the obtained z-stack is shown on the upper and right-hand side. n = 8. MMP13, white; lectin, green; and DAPI, blue. GCL = ganglion cell layer. INL = inner nuclear layer. MMP13 = matrix metalloproteinase 13. RPE = retinal pigment epithelium. Scale bars: 20 µm.

Article Snippet: They were then exposed for 3 days at RT to mouse monoclonal anti-MMP13 (for precursor and active forms; 1:100, MAB3321, Merck), biotinylated isolectin (1:240, B-1205; Vector Labs, Burlingame, CA, USA), and a goat collagen IV antibody (1:250, 1340-01, Southern Biotech, CA, USA).

Techniques: Immunofluorescence, Expressing

Journal: Antioxidants

Article Title: Matrix Metalloproteinase 13 Is Associated with Age-Related Choroidal Neovascularization

doi: 10.3390/antiox12040884

Figure Lengend Snippet: MMP13 expression in laser-induced CNV. WT mouse retina subjected to laser-induced CNV showing MMP13 expression in retinal vessels and diffuse overexpression in retinal parenchyma above the lesions. ( A ) Whole flatmount micrograph; the outlined region represents the magnification shown in ( B ). ( B ) The dotted square represents the magnified area shown in ( C – E ), and the retina overlying the CNV area is located in the center (dotted circle). ( C ) MMP13-positive retinal vessels and a cluster of nuclei (blue) above the CNV (dotted circle). ( D ) The cluster of cells shown in ( C ) showed MMP13 expression. ( E ) Retinal vessels positive for lectin (green). n = 8. Hoechst 33342-labeled nuclei, blue; lectin, green; and MMP13, white. EC = endothelial cells. CNV = choroidal neovascularization. MMP13 = matrix metalloproteinase 13. ON = optic nerve. WT = wild type. Scale bars: 500 µm ( A ), 100 µm ( C – E ).

Article Snippet: They were then exposed for 3 days at RT to mouse monoclonal anti-MMP13 (for precursor and active forms; 1:100, MAB3321, Merck), biotinylated isolectin (1:240, B-1205; Vector Labs, Burlingame, CA, USA), and a goat collagen IV antibody (1:250, 1340-01, Southern Biotech, CA, USA).

Techniques: Expressing, Over Expression, Labeling

Journal: Antioxidants

Article Title: Matrix Metalloproteinase 13 Is Associated with Age-Related Choroidal Neovascularization

doi: 10.3390/antiox12040884

Figure Lengend Snippet: MMP13 is overexpressed in CNV, showing expression in RPE cells and endothelial cells. Representative laser-induced CNV lesions on choroidal flatmounts from WT mice stained with MMP13 (white), lectin (green), and Hoechst 33342 (blue). ( A ) Whole flatmount micrograph with successfully induced CNV. The outlined region represents the CNV shown in ( B – J ). ( B ) Magnification of CNV area showing increased MMP13 expression. ( C ) The neovascular membrane is clearly visible and stained with lectin (green). ( D ) Merged image of the MMP13 and lectin staining. ( E ) Orthogonal view of the obtained z-stack at RPE level. ( F ) The RPE showed MMP13 expression, especially in the center of the CNV. ( G ) No lectin-positive cells were observed at this level. ( H ) Orthogonal view of the obtained z-stack at the level of the CNV. ( I ) ECs were MMP13-positive. ( J ) Endothelial cells were lectin-positive (green). n = 8. Hoechst 33342, blue; lectin, green; and MMP13, white. EC = endothelial cells. CNV = choroidal neovascularization. MMP13 = Matrix metalloproteinase 13. ON = optic nerve. RPE = retinal pigment epithelium. WT = wild type. Scale bars: 20 µm ( B – J ), 200 µm ( A ).

Article Snippet: They were then exposed for 3 days at RT to mouse monoclonal anti-MMP13 (for precursor and active forms; 1:100, MAB3321, Merck), biotinylated isolectin (1:240, B-1205; Vector Labs, Burlingame, CA, USA), and a goat collagen IV antibody (1:250, 1340-01, Southern Biotech, CA, USA).

Techniques: Expressing, Staining

Journal: Antioxidants

Article Title: Matrix Metalloproteinase 13 Is Associated with Age-Related Choroidal Neovascularization

doi: 10.3390/antiox12040884

Figure Lengend Snippet: MMP13 is overexpressed in the RPE underlying the CNV. Representative laser-induced CNV lesions on choroidal flatmounts from WT mice stained with MMP13 (white), lectin (green), and Hoechst 33342 (blue). RPE cells underlying the laser-induced CNV mice were positive for MMP13 staining, while the RPE far from this area did not express MMP13. ( A ) Whole flatmount micrograph with three successfully laser-induced CNV lesions; the outlined region represents the magnification in ( B , C ). ( B ) MMP13 staining in the CNV area. ( C ) Lectin staining in the CNV area. The outlined region represents the magnification in ( D , E ). Panels ( D , E ) show that lectin-negative cells, corresponding to RPE cells, expressed MMP13 close to the area of CNV but not outside the lesion. n = 8. Hoechst 33342, blue; lectin, green; and MMP13, white. CNV = choroidal neovascularization. MMP13 = matrix metalloproteinase 13. ON = optic nerve. RPE = retinal pigment epithelium. Scale bars: 200 µm ( A ), 20 µm ( B – E ).

Article Snippet: They were then exposed for 3 days at RT to mouse monoclonal anti-MMP13 (for precursor and active forms; 1:100, MAB3321, Merck), biotinylated isolectin (1:240, B-1205; Vector Labs, Burlingame, CA, USA), and a goat collagen IV antibody (1:250, 1340-01, Southern Biotech, CA, USA).

Techniques: Staining

Journal: Antioxidants

Article Title: Matrix Metalloproteinase 13 Is Associated with Age-Related Choroidal Neovascularization

doi: 10.3390/antiox12040884

Figure Lengend Snippet: MMP13 immunofluorescence characterization in the human posterior pole. ( A ) The RPE and ( B ) the choriocapillaris did not show MMP13 expression. ( C ) Some cells located in the inner part of the sclera expressed MMP13. ( D ) Retinal vessels showed MMP13 expression, as observed in the z-stack cross-section of the vessel. ( E ) Lectin-positive endothelial cells colocalized with MMP13. ( F ) Some retinal parenchymal cells, predominantly in inner retinal layers, showed a marked MMP13 signal. Some of these cells showed a microglial-like activated morphology. ( G ) MMP13-positive cells were also observed in the lumen of the vessels. n = 3. MMP13, white; lectin, green; collagen IV, red; and DAPI, blue. MMP13 = matrix metalloproteinase 13. ON = optic nerve. RPE = retinal pigment epithelium. Scale bars: 20 µm.

Article Snippet: They were then exposed for 3 days at RT to mouse monoclonal anti-MMP13 (for precursor and active forms; 1:100, MAB3321, Merck), biotinylated isolectin (1:240, B-1205; Vector Labs, Burlingame, CA, USA), and a goat collagen IV antibody (1:250, 1340-01, Southern Biotech, CA, USA).

Techniques: Immunofluorescence, Expressing

Journal: Antioxidants

Article Title: Matrix Metalloproteinase 13 Is Associated with Age-Related Choroidal Neovascularization

doi: 10.3390/antiox12040884

Figure Lengend Snippet: Comparison of mean total MMP13 plasma levels in the age- and sex-matched case–control cohort. A statistically significant difference was found in patients with wet AMD presenting lower plasma levels than controls. AMD = age-related macular degeneration, MMP13 = matrix metalloproteinase 13. ** p < 0.01. Error bars correspond to the SEM. Control group, n = 52; AMD group, n = 52. Mann–Whitney U test was used.

Article Snippet: They were then exposed for 3 days at RT to mouse monoclonal anti-MMP13 (for precursor and active forms; 1:100, MAB3321, Merck), biotinylated isolectin (1:240, B-1205; Vector Labs, Burlingame, CA, USA), and a goat collagen IV antibody (1:250, 1340-01, Southern Biotech, CA, USA).

Techniques: MANN-WHITNEY