Journal: Frontiers in Immunology

Article Title: Thrombospondin-1 mitigates osteoarthritis progression by inhibiting mechanical stress-induced chondrocyte ferroptosis via the integrin/YAP pathway

doi: 10.3389/fimmu.2025.1577234

Figure Lengend Snippet: The interaction between THBS1 and IntegrinαVβ1 is enhanced under mechanical stress. (A) Interaction network of differential genes, each circle represented a gene, and the size of the circle represented the core level of the gene. The larger the circle, the higher the core level the gene had in the network. (B) Cartoon shows various binding domains of THBS1 and its receptors. (C) Representative immunostaining of THBS1 showing colocalization with integrins αv and β1 (white arrows) but not with integrin β3, CD47, or CD36 under mechanical stress (n = 6). Scale bars, 50 μm. (D) Quantification of colocalization of THBS1 with molecules shown in C using Image J software. (E) Immunoprecipitation (IP) with anti-THBS1 or control immunoglobulin G (IgG) followed by Western blotting for integrins αv and β1 (n = 3). IB, immunoblot. Data were presented as the mean ± SD. **P<0.01.

Article Snippet: IntegrinαVβ1 inhibitor αVβ1 integrin-IN- 1 was obtained from MCE (HY-145363). αVβ1 integrin-IN- 1(100ng/ml) ( ) was added to chondrocytes at the same time of mechanical stress stimulation with or without rhTHBS1, and incubated for 24 hours after the end of mechanical stress stimulation.

Techniques: Binding Assay, Immunostaining, Software, Immunoprecipitation, Control, Western Blot

Journal: Frontiers in Immunology

Article Title: Thrombospondin-1 mitigates osteoarthritis progression by inhibiting mechanical stress-induced chondrocyte ferroptosis via the integrin/YAP pathway

doi: 10.3389/fimmu.2025.1577234

Figure Lengend Snippet: THBS1 inhibits mechanical stress-induced ferroptosis in chondrocytes through integrinαVβ1. Human chondrocytes were stimulated at 1MPa mechanical stress for 2 hours with or without integrinαVβ1 inhibitor αVβ1 integrin-IN- 1(100ng/ml) and rhTHBS1(100ng/ml). Relevant tests were performed 24 hours after the end of mechanical stress stimulation. (A) The cell death ratio of chondrocytes was tested by cell death/live analysis. Scale bar = 50 μm. (B) The cell number of PI (red fluorescence)/calcein (green fluorescence) reflected the cell death ratio (n=3 for each group). (C) Mitochondrial membrane potential was detected by JC-1 assay. Scale bar = 10 μm. (D) The relative IOD ratio of red fluorescence to green fluorescence was used for quantitative analysis (n=3 for each group). (E) Representative TEM images of chondrocytes of indicated groups. (n=3 for each group). Green arrows show the normal mitochondria. Red arrows show the shrunken mitochondria. Scale bars, 1 μm (low field), 500 nm (high field). (F) Representative fluorescence images of mitochondria in chondrocytes. Scale bar = 50 μm. (G) Quantitative analysis of fluorescence intensity (n=3 for each group). (H) Representative images of ROS levels in chondrocytes. Scale bar = 50 μm. (I) Quantitative analysis of fluorescence intensity (n=3 for each group). (J) The expression of GSH in chondrocytes of indicated groups was detected by ELISA (n=3 for each group). (K) Western blot (WB) analysis of Col2, MMP-9 and GPX4. (L-N) Quantification of WB analysis (n=3 for each group). Data were presented as the mean ± SD. **P<0.01.

Article Snippet: IntegrinαVβ1 inhibitor αVβ1 integrin-IN- 1 was obtained from MCE (HY-145363). αVβ1 integrin-IN- 1(100ng/ml) ( ) was added to chondrocytes at the same time of mechanical stress stimulation with or without rhTHBS1, and incubated for 24 hours after the end of mechanical stress stimulation.

Techniques: Fluorescence, Membrane, Expressing, Enzyme-linked Immunosorbent Assay, Western Blot

Journal: Frontiers in Immunology

Article Title: Thrombospondin-1 mitigates osteoarthritis progression by inhibiting mechanical stress-induced chondrocyte ferroptosis via the integrin/YAP pathway

doi: 10.3389/fimmu.2025.1577234

Figure Lengend Snippet: THBS1 inhibits chondrocyte ferroptosis through the integrinαVβ1/YAP pathway. THBS1 knockout chondrocytes and normal chondrocytes were subjected to 1 MPa mechanical stress for 2 hours, with or without the addition of YAP1 inhibitor PROTAC YAP d1 (20μM) and rhTHBS1(100ng/ml). Immediately following stimulation, IF detection was performed. Additional analyses were conducted after chondrocytes were further incubated with or without YAP1 inhibitor PROTAC YAP d1 (20μM) and rhTHBS1(100ng/ml) for 24 hours post-stimulation. (A) Representative image of chondrocytes undergoing IF detection immediately after mechanical stress stimulation. YAP (red) and DAPI (blue) are also shown. Scale bar = 20 μm. (B) Quantification of YAP localization. In each experiment 150 to 200 cells were evaluated (n = 3). YAP localization in the nucleus (green) or cytoplasm (red) is shown. (C) Western blot (WB) analysis of YAP1. (D) Quantification of WB analysis (n=3 for each group). (E) Representative image of chondrocytes undergoing IF detection immediately after mechanical stress stimulation. YAP (red) and DAPI (blue) are also shown. Scale bar = 20 μm. (F) Quantification of YAP localization. In each experiment 150 to 200 cells were evaluated (n = 3). (G) Western blot (WB) analysis of YAP1. (H) Quantification of WB analysis (n=3 for each group). (I) Mitochondrial membrane potential was detected by JC-1 assay. Scale bar = 10 μm. (J) The relative IOD ratio of red fluorescence to green fluorescence was used for quantitative analysis (n=3 for each group). (K) Representative TEM images of chondrocytes of indicated groups. (n=3 for each group). Green arrows show the normal mitochondria. Red arrows show the shrunken mitochondria. Scale bars, 5.0 μm (low field), 500 nm (high field). (L) Representative fluorescence images of mitochondria in chondrocytes. Scale bar = 50 μm. (M) Quantitative analysis of fluorescence intensity (n=3 for each group). (N) The expression of GSH in chondrocytes of indicated groups was detected by ELISA (n=3 for each group). Data were presented as the mean ± SD. NS P>0.05 *P<0.05, **P<0.01.

Article Snippet: IntegrinαVβ1 inhibitor αVβ1 integrin-IN- 1 was obtained from MCE (HY-145363). αVβ1 integrin-IN- 1(100ng/ml) ( ) was added to chondrocytes at the same time of mechanical stress stimulation with or without rhTHBS1, and incubated for 24 hours after the end of mechanical stress stimulation.

Techniques: Knock-Out, Incubation, Western Blot, Membrane, Fluorescence, Expressing, Enzyme-linked Immunosorbent Assay

Journal: bioRxiv

Article Title: Overcoming Immune Checkpoint Inhibitor Resistance via Potent and Selective Dual αvβ6/8 Inhibitors Based on Engineered Lasso Peptides

doi: 10.1101/2025.01.28.635346

Figure Lengend Snippet:

Article Snippet: Human integrins αvβ1, αvβ3, αvβ5, αvβ6, αvβ8, α5β1, and αIIbβ3 were purchased from R&D Systems (Minneapolis, MN) or Acro Biosystems (Newark, DE, USA).

Techniques: Inhibition, Enzyme-linked Immunosorbent Assay, Positive Control