Journal: International Journal of Molecular Sciences

Article Title: ER Stress in ERp57 Knockout Knee Joint Chondrocytes Induces Osteoarthritic Cartilage Degradation and Osteophyte Formation

doi: 10.3390/ijms23010182

Figure Lengend Snippet: ERp57 cKO mice show an accelerated development of osteoarthritis during aging. ( A ) Histological analysis and OARSI scoring of toluidine blue-stained knee sections of 18-month-old mice reveals an increase in cartilage degradation, the occurrence of hypertrophic chondrocytes (*), and osteophyte formation (scattered box) in ERp57 cKO mice, but not in WT animals ( p = 0.0039). No significant difference was detected between younger WT and ERp57 cKO mice at the age of 9 months ( p = 0.7924) and 12 months ( p = 0.2503). Scale Bars = 100 µm. ( B ) TUNEL staining indicates a higher percentage of apoptotic cells in the articular cartilage of ERp57 cKO mice at 9 ( p = 0.0271), 12 ( p = 0.0490) and 18 months ( p = 0,000000028) compared to WT animals. Scale bars = 50 µm ( C ) Immunofluorescence of 18-month-old ERp57 cKO cartilage reveals increased staining intensities of matrix degrading proteases MMP3 ( p = 0.0004), MMP9 ( p = 0.0097), ADAMTS4 ( p = 0.0428), and ADAMTS5 ( p = 0.0288) compared to WT controls. M = months. n ≥ 4. *** p < 0.005, ** p < 0.01, * p < 0.05.

Article Snippet: Then the Vectastain ® Elite ABC Universal Kit (VEC-PK-7200, BIOZOL, Eching, Germany) and the Vector ® DAB Peroxidase Substrate (VEC-SK-4100, BIOZOL, Eching, Germany) were used according to the manufacturer’s instructions with polyclonal antibodies (anti-ADAMTS4: bs-4191R, Bioss Antibodies Inc., Woburn, MA, USA, 1:200 and ADAMTS5: bs-3573R, Bioss Antibodies Inc, Woburn, MA, USA, 1:50), which were incubated on the slides overnight at 4 °C.

Techniques: Staining, TUNEL Assay, Immunofluorescence

Journal: International Journal of Molecular Sciences

Article Title: ER Stress in ERp57 Knockout Knee Joint Chondrocytes Induces Osteoarthritic Cartilage Degradation and Osteophyte Formation

doi: 10.3390/ijms23010182

Figure Lengend Snippet: ER stress-induced osteoarthritic changes in the knee joint. Knee joints of aged ERp57 cKO animals display severe OA cartilage degeneration with enhanced MMP3, MMP9, ADAMTS4, and ADAMTS5 expression and osteophyte formation. These changes are preceded by reduced compressive stiffness of the cartilage ECM and increased apoptotic cell death. Both effects are likely induced by ER stress that cannot be adequately compensated by the UPR.

Article Snippet: Then the Vectastain ® Elite ABC Universal Kit (VEC-PK-7200, BIOZOL, Eching, Germany) and the Vector ® DAB Peroxidase Substrate (VEC-SK-4100, BIOZOL, Eching, Germany) were used according to the manufacturer’s instructions with polyclonal antibodies (anti-ADAMTS4: bs-4191R, Bioss Antibodies Inc., Woburn, MA, USA, 1:200 and ADAMTS5: bs-3573R, Bioss Antibodies Inc, Woburn, MA, USA, 1:50), which were incubated on the slides overnight at 4 °C.

Techniques: Expressing

Journal: Nature Communications

Article Title: An injectable liposome-anchored teriparatide incorporated gallic acid-grafted gelatin hydrogel for osteoarthritis treatment

doi: 10.1038/s41467-023-38597-0

Figure Lengend Snippet: a , d Live/dead stained fluorescent images and statistical analysis for cells’ cytotoxicity, green stand for the live cells and red is the dead cells. b , e EdU/DAPI stained fluorescent images to evaluate the cell proliferation (red represents the newly proliferated cells (EdU staining), blue represents all cells (DAPI staining)). c , f Flow cytometry (FCM) results and G2/M rate calculated from FCM results. g – l Immunofluorescent images for Ki-67, c-Fos, and PTH1R expression for the ATDC5 cells cultured with hydrogels and the statistical stained positively area for the expression of such genes. m , n Western Blot results for the expression of proliferation proteins (PTH1R, SOX9, MMP13, ADAMTS5) of ATDC5 cells when cultured with hydrogels and the corresponding statistical analysis results. Data are presented as means ± SD of at least three replicate experiments. Unpaired two-tailed Student’s t tests was used to calculate significant difference, * p < 0.05, ** p < 0.01. GGA gallic acid-grafted gelatin, GLP GGA@Lipo@PTH (1–34).

Article Snippet: The immunohistochemical stainings including PTH1R (Cat#BS2710, 1:400, Bioword Technology, US), SOX9 (Cat#ab185966, 1:1000, abcam, UK), ACAN (Cat#13880-1-AP, 1:500, Proteintech, China), COLIIA1 (Cat#28459-1-AP, 1:500, Proteintech, China), MMP13 (Cat#18165-1-AP, 1:500, Proteintech, China); ADAMTS5 (Cat#bs-3573R, 1:200, Bioss, China), Bcl-2 (Cat#68103-1-Ig, 1:500, Proteintech, China), and BAX (Cat#AF0057, 1:200, Beyotime Biotechnology, China). were performed with 4 μm in thick sections.

Techniques: Staining, Flow Cytometry, Expressing, Cell Culture, Western Blot, Two Tailed Test

Journal: Nature Communications

Article Title: An injectable liposome-anchored teriparatide incorporated gallic acid-grafted gelatin hydrogel for osteoarthritis treatment

doi: 10.1038/s41467-023-38597-0

Figure Lengend Snippet: a , b The FCM results and statistical analysis results for the percentage of apoptosis. c – h Immunofluorescent images for SOX9, Bcl-2, and BAX protein expression as well as the statistical stained positively area from the images. Sox9 is the characteristic gene to indicate the ECM expression, Bcl-2 and Bax genes are the anti-apoptosis and the apoptosis-promoting genes of the cells. i – n The RT-qPCR results of the mRNA expression levels for Sox9, Col2a1, Acan, Adamts5, iNOS, and COX2. o , p The ELISA results for inflammatory mediators, IL-6 and TNF-α. q , r The Western Blot results for the expression of proteins (p-PI3K, PI3K, p-AKT/AKT, ADAMTS5) of IL-1β-induced ATDC5 cells cultured with hydrogels and the corresponding statistical analysis results. Data are presented as means ± SD of at least three replicate experiments. Unpaired two-tailed Student’s t tests was used to calculate significant difference, * p < 0.05, ** p < 0.01. GGA gallic acid-grafted gelatin, GLP GGA@Lipo@PTH (1–34).

Article Snippet: The immunohistochemical stainings including PTH1R (Cat#BS2710, 1:400, Bioword Technology, US), SOX9 (Cat#ab185966, 1:1000, abcam, UK), ACAN (Cat#13880-1-AP, 1:500, Proteintech, China), COLIIA1 (Cat#28459-1-AP, 1:500, Proteintech, China), MMP13 (Cat#18165-1-AP, 1:500, Proteintech, China); ADAMTS5 (Cat#bs-3573R, 1:200, Bioss, China), Bcl-2 (Cat#68103-1-Ig, 1:500, Proteintech, China), and BAX (Cat#AF0057, 1:200, Beyotime Biotechnology, China). were performed with 4 μm in thick sections.

Techniques: Expressing, Staining, Quantitative RT-PCR, Enzyme-linked Immunosorbent Assay, Western Blot, Cell Culture, Two Tailed Test

Journal: Nature Communications

Article Title: An injectable liposome-anchored teriparatide incorporated gallic acid-grafted gelatin hydrogel for osteoarthritis treatment

doi: 10.1038/s41467-023-38597-0

Figure Lengend Snippet: The GLP hydrogel would promote ATDC5 cells proliferation by up-regulating the expression of cell proliferation and anti-apoptosis genes (Ki-67, c-Fos), and this hydrogel would protect IL-1β-induced ATDC5 cells from further progression by up-regulating the expression of key anabolic genes (Sox9, Bcl-2, Col2a1, Acan) and down-regulating the expression of key catabolic genes (Bax, Adamts5), which potentially suggested regulating the PI3K/AKT signaling pathway. GLP GGA@Lipo@PTH (1–34).

Article Snippet: The immunohistochemical stainings including PTH1R (Cat#BS2710, 1:400, Bioword Technology, US), SOX9 (Cat#ab185966, 1:1000, abcam, UK), ACAN (Cat#13880-1-AP, 1:500, Proteintech, China), COLIIA1 (Cat#28459-1-AP, 1:500, Proteintech, China), MMP13 (Cat#18165-1-AP, 1:500, Proteintech, China); ADAMTS5 (Cat#bs-3573R, 1:200, Bioss, China), Bcl-2 (Cat#68103-1-Ig, 1:500, Proteintech, China), and BAX (Cat#AF0057, 1:200, Beyotime Biotechnology, China). were performed with 4 μm in thick sections.

Techniques: Expressing

Journal: International Journal of Molecular Sciences

Article Title: Inhibitory Effect of a Tankyrase Inhibitor on Mechanical Stress-Induced Protease Expression in Human Articular Chondrocytes

doi: 10.3390/ijms25031443

Figure Lengend Snippet: ( A ) The effect of mechanical stress and XAV939 on the expression of ADAMTS-5 and MMP-13 in normal human chondrocytes. Up-regulation of ADAMTS-5 and MMP-13 was localized to the cytoplasm (DAPI: blue signal, ADAMTS-5 and MMP-13: green signal). Scale bar = 20 μm. ( B ) The percentages of chondrocytes positive for ADAMTS-5 and MMP-13. Cell numbers were counted in four fields, at 10× magnification, and the mean was calculated (* p < 0.05, ** p < 0.01, *** p < 0.0001). ( C ) The effect of XAV939 on the protein expression of ADAMTS-5 and MMP-13 by Western blot and quantification analysis.

Article Snippet: The membranes of the culture chambers were then removed and incubated with anti-ADAMTS-5 antibody (1:100, ab45042, Abcam, Cambridge, MA, USA), anti-MMP-13 antibody (1:100, ab39012, Abcam), anti-β-catenin antibody (1:3200, 15B8, Cell Signaling Technology, Danvers, MA, USA), or anti-NF-kB p65 antibody (1:400, D14E12, Cell Signaling Technology) for 120 min at room temperature.

Techniques: Expressing, Western Blot