integrin αvβ3 antagonist cilengitide 268  (MedChemExpress)

Journal: Nature communications

Article Title: CCN1 interacts with integrins to regulate intestinal stem cell proliferation and differentiation.

doi: 10.1038/s41467-022-30851-1

Figure Lengend Snippet: Fig. 5 CCN1 regulates ISC differentiation through integrins αvβ3/αvβ5-mediated NF-κB activation. a AP staining and immunofluorescence staining of markers (Lyz1, Muc2, Lgr5 + GFP, Ki67) for the analysis of differentiation and proliferation of the Ccn1ΔLgr5 organoids. DAPI is counterstaining. Organoids were treated with vehicle or 4-OHT (1 µM) for 5 days. For add-back experiments, CCN1 (WT) or CCN1-D125A proteins (4 µg per ml each) were treated to organoids 1 day after 4-OHT treatment began. Small-molecule inhibitors for integrin αv (SB273005, 1 µM), FAK inhibitor (GSK2256098, 500 nM), and IκB kinase inhibitor (Bay11-7082, 5 µM) were pretreated 30 min before CCN1 add-back. Images are representative of three independent replicates. Bars: 100 µm. b qPCR analysis of lineage markers in organoids treated as in a. Data represent the mean ± SEM of four biological replicates. One-sided t-test. **p < 0.01; ***p < 0.001; ns not significant. c, d Organoids were treated as described in a. Inhibitors used are SB273005 (1 µM), Bay11-7082 (5 µM), and DAPT (10 µM). c Immunoblot analysis of p-p65, total p65, Jag1, and N1-ICD. Representative images are shown from three replicates. d qPCR analysis of Jag1 and Hes1 expression in organoids treated as in c. Data represent the mean ± SEM of four biological replicates. One-sided t-test. ***p < 0.001; ns not significant. e, f Ccn1ΔLgr5 organoids with 4-OHT (1 µM, e) or Ccn1D125A/D125A organoids (f) were treated with CCN1 (4 µg per ml) or Jag1 (2 µg per ml) and examined for differentiation by AP staining and immunofluorescence staining of Lyz1 and Muc2. Bar: 100 µm. Representative images are shown from three independent replicates. Source data and the exact p-values are provided in a Source Data file.

Article Snippet: Integrin αvβ3/αvβ5 inhibitor (SB273005; S7540), NFκB inhibitor (Bay11-7082; S2913), γ-secretase inhibitor (DAPT; S2215), YAPTEAD association inhibitors (Super-TDU; S8554, Verterporfin; S1786), FAK inhibitors (GSK2256098; S8523), and GSK3β inhibitor (CHIR99021; CT99021) were purchased from Selleckchem.

Techniques: Activation Assay, Staining, Western Blot, Expressing

Journal: Nature communications

Article Title: CCN1 interacts with integrins to regulate intestinal stem cell proliferation and differentiation.

doi: 10.1038/s41467-022-30851-1

Figure Lengend Snippet: Fig. 6 CCN1 regulates Wnt activity through integrins αvβ3/αvβ5-Src-YAP axis. a AP staining and immunofluorescence staining of markers (Lyz1, Muc2, Lgr5 + GFP, and Ki67) for the analysis of differentiation and proliferation of the Ccn1ΔLgr5 organoids. Ccn1ΔLgr5 organoids were treated with vehicle or 4-OHT (1 µM) for 5 days. CCN1 (4 µg per ml) was added to organoids 1 day after 4-OHT. Inhibitors (Dasatinib, 10 nM; Super-TDU, 50 nM) were pretreated 30 min before CCN1. Representative images are shown from three independent replicates. Bars: 100 µm. b qPCR analysis of lineage markers in organoids treated as in a. Data represent the mean ± SEM of four biological replicates. One-sided t-test. *p < 0.05; **p < 0.01; *** p < 0.001; ns not significant. c Immunoblot analysis of indicated proteins from Ccn1ΔLgr5 organoids treated as described in a. Representative images are shown from three replicates. d qPCR analysis of Wnt ligands (Wnt3a, Wnt5a, Wnt5b, and Wnt9b) and Wnt antagonists (Dkk1, Dkk2, and sFRP5) in Ccn1ΔLgr5 organoids treated with vehicle or 4-OHT (1 µM, d5). Data represent the mean ± SEM of four biological replicates. One-sided t-test. ***p < 0.001; ns not significant. e qPCR analysis of Dkk1 expression in Ccn1ΔLgr5 organoids treated as indicated (CCN1, 4 µg per ml; Super-TDU, 50 nM). Data represent the mean ± SEM of four biological replicates. One-sided t-test. **p < 0.01; ns not significant. f Immunoblot analysis of active and total β-catenin in Ccn1ΔLgr5 organoids treated as indicated (CCN1, 4 µg per ml; Super-TDU, 10 and 50 nM). β-actin as a loading control. Representative images are shown from three replicates. g qPCR analysis of Wnt target genes (Axin2, Cd44, and Ephb2) in Ccn1ΔLgr5 organoids treated as indicated. Data represent the mean ± SEM of four biological replicates. One- sided t-test. *p < 0.05; **p < 0.01; ***p < 0.001. h Immunofluorescence staining of Lgr5 + GFP and Ki67 in Ccn1ΔLgr5 organoids treated as indicated. A neutralizing anti-Dkk1 antibody (5 µg per ml) or isotype control IgG was treated in place of inhibitors as in a. Representative images are shown from three replicates. Bars: 100 µm. Source data and the exact p-values are provided in a Source Data file.

Article Snippet: Integrin αvβ3/αvβ5 inhibitor (SB273005; S7540), NFκB inhibitor (Bay11-7082; S2913), γ-secretase inhibitor (DAPT; S2215), YAPTEAD association inhibitors (Super-TDU; S8554, Verterporfin; S1786), FAK inhibitors (GSK2256098; S8523), and GSK3β inhibitor (CHIR99021; CT99021) were purchased from Selleckchem.

Techniques: Activity Assay, Staining, Western Blot, Expressing, Control

Journal: Frontiers in Cell and Developmental Biology

Article Title: Role of Integrin αvβ3 in Doxycycline-Induced Anti-Proliferation in Breast Cancer Cells

doi: 10.3389/fcell.2022.829788

Figure Lengend Snippet: Primer sequences for the qPCR.

Article Snippet: The integrin αvβ3 inhibitor (HSDVHK-NH 2 ) was purchased from Calbiochem (San Diego, CA, United States).

Techniques:

Journal: Frontiers in Cell and Developmental Biology

Article Title: Role of Integrin αvβ3 in Doxycycline-Induced Anti-Proliferation in Breast Cancer Cells

doi: 10.3389/fcell.2022.829788

Figure Lengend Snippet: Blockage of integrin αvβ3 activity inhibits doxycycline-induced signal transduction in breast cancer MDA-MB-231 cells. (A) Cells were pre-treated with HSDVHK-NH 2 (10 µM) for 24 h, then treated with doxycycline (60 µM) for 24 h. Cells were harvested and protein was extracted for Western blotting analysis of pFAK (Y397), pFAK (Y925), FAK, pERK1/2, and total ERK1/2. The HSDVHK-NH 2 compound reduced doxycycline-induced phosphorylation of Y397 (B) , Y925 (C) , and pERK1/2 (D) . Numbers of independent studies ( n ) = 3. ANOVA was used to assess statistical significance. * p < 0.05, ** p < 0.01, *** p < 0.001, compared to the untreated control. ### p < 0.001, compared to doxycycline alone.

Article Snippet: The integrin αvβ3 inhibitor (HSDVHK-NH 2 ) was purchased from Calbiochem (San Diego, CA, United States).

Techniques: Activity Assay, Transduction, Western Blot, Phospho-proteomics, Control

Journal: Frontiers in Cell and Developmental Biology

Article Title: Role of Integrin αvβ3 in Doxycycline-Induced Anti-Proliferation in Breast Cancer Cells

doi: 10.3389/fcell.2022.829788

Figure Lengend Snippet: Blockage of integrin αvβ3 activity inhibits doxycycline-induced gene expression in breast cancer MDA-MB-231 cells. MDA-MB-231 cells were pre-treated with HSDVHK-NH 2 (10 µM) for 24 h, then treated with doxycycline (60 µM) for 24 h. Cells were harvested. RNA was extracted for qPCR. Numbers of independent studies ( n ) = 3. ANOVA was used to assess statistical significance. * p < 0.05, ** p < 0.01, *** p < 0.001, compared to the untreated control. ## p < 0.01, ### p < 0.001, compared to doxycycline alone.

Article Snippet: The integrin αvβ3 inhibitor (HSDVHK-NH 2 ) was purchased from Calbiochem (San Diego, CA, United States).

Techniques: Activity Assay, Gene Expression, Control

Journal: Frontiers in Cell and Developmental Biology

Article Title: Role of Integrin αvβ3 in Doxycycline-Induced Anti-Proliferation in Breast Cancer Cells

doi: 10.3389/fcell.2022.829788

Figure Lengend Snippet: Integrin αvβ 3 and ERK1/2 collaborate to thyroxine- or doxycycline-induced PD-L1 expression in breast cancer MDA-MB-231 cells. MDA-MB-231 cells were pre-treated with (A) HSDVHK-NH 2 (10 µM) or (B) PD98059 (25 µM) for 24 h, then treated with T 4 (100 nM) or doxycycline (60 µM) for 24 h. Cells were harvested. RNA was extracted for qPCR. Numbers of independent studies ( n ) = 3. ANOVA was used to assess statistical significance. ** p < 0.01, *** p < 0.001, compared to the untreated control. # p < 0.05, ### p < 0.001, compared to doxycycline or T 4 treatment alone. (C) MDA-MB-231 cells were pre-treated with HSDVHK-NH 2 (10 µM) for 24 h, then treated with doxycycline (60 µM) for 24 h. Cells were harvested and protein was extracted for PD-L1 expression analysis. Numbers of independent studies ( n ) = 3. ANOVA was used to assess statistical significance. # p < 0.05, compared to doxycycline alone.

Article Snippet: The integrin αvβ3 inhibitor (HSDVHK-NH 2 ) was purchased from Calbiochem (San Diego, CA, United States).

Techniques: Expressing, Control