Journal: European journal of pharmacology

Article Title: PlGF signaling and macrophage repolarization contribute to the anti-neoplastic effect of metformin.

doi: 10.1016/j.ejphar.2019.172696

Figure Lengend Snippet: Fig. 5. VEGFR-1 signaling mediated autocrine and exogenous PlGF-induced proliferation. (A and B) In vitro proliferative curves of (A) 4T1 and MCF-7 cells, treated with PBS (Con), recombinant PlGF, or in combination of siRNA against VEGFR-1 or VEGFR-2 (n = 6). (C) Immunoblotting for VEGFR-2 of endothelial cell (EC), 4T1 and MCF-7 breast cancer cells. EC is used as the control. (D) Immunostaining for VEGFR-1 (Left) and VEGFR-2 (Right) in 4T1 cells from untreated mouse. (E and F) In vitro proliferation curves of (E) MCF-7 and (F) 4T1 breast cells, treated with PBS (Con), recombinant PlGF or in combination with GDC-0994 (n = 6). Measurement data are expressed as mean ± S.D. **P < 0.01; ***P < 0.001; ns indicates not statistically significant (P > 0.05).

Article Snippet: Recombinant human and mouse PlGF proteins were obtained from R&D SYSTEMs.

Techniques: In Vitro, Recombinant, Western Blot, Control, Immunostaining

Journal: eBioMedicine

Article Title: The therapeutic effects of the VEGF decoy receptor fusion protein VEGF-Grab in chronic inflammatory diseases

doi: 10.1016/j.ebiom.2026.106216

Figure Lengend Snippet: Inhibitory effects of PB101 on angiogenesis. ( A ) Endothelial cell chemotactic migration assay. EA.hy926 cells (1 × 10 5 , n = 3) were loaded in the upper chambers of Transwell inserts and chemoattracted by VEGF (100 ng/mL) or PlGF-2 (40 ng/mL) with or without PB101 (2 or 5 μg/mL) in the lower chambers. The cells that migrated to the underside of the Transwell chamber were manually counted, and the results are presented as a percentage of the untreated control (none). ( B ) Wounding migration of endothelial cells. EA.hy926 cells (4 × 10 5 , n = 5) seeded on a 6-well plate were wounded using pipette tips and treated with VEGF (200 ng/mL) or PlGF-2 (100 ng/mL) with or without PB101 (2 or 5 μg/mL). The cells that crossed the reference lines (red dashed lines) were counted as migrated cells. ( C ) Capillary tube formation by endothelial cells. EA.hy926 cells (3 × 10 4 , n = 3–4) were seeded on a Matrigel-coated 96-well plate and incubated with VEGF (500 ng/mL) or PlGF-2 (500 ng/mL) with or without PB101 (10 or 50 μg/mL). The tube formation area was measured and is presented as an arbitrary unit (a.u.). The cellular images are representative of three independent experiments. Scale bar, 200 μm. The bar graphs present the mean and SEM. The p-values were determined via one-way ANOVA with Tukey's multiple comparisons test. ∗p < 0.05, ∗∗p < 0.01, and ∗∗∗p < 0.001, ∗∗∗∗p < 0.0001 vs. untreated controls. # p < 0.05, ## p < 0.01 vs. PlGF- or VEGF-treated positive controls. ( D and E ) Suppression of recombinant PlGF-induced angiogenesis by PB101 in a Matrigel plug assay. C57BL/6 mice (n = 3, total 12 mice) were injected subcutaneously with Matrigel mixed with VEGF (500 ng/mL) or recombinant PlGF-2 (200 ng/mL) with or without PB101 (50 μg/mL) (D). Mice were injected with vehicle or PB101 (50 mg/kg) every day. To assess the effects of PlGF-overexpressing T cells, CD4 + T cells isolated from PlGF transgenic (PlGF Tg) mice were stimulated with anti-CD3/CD28 Abs for 48 h. PlGF Tg CD4 + T cells (5 × 10 5 ) and their culture supernatants were incorporated into Matrigel with or without PB101 (50 μg/mL) (E). C57BL/6 mice (n = 3) were subcutaneously injected with these Matrigel plugs and received daily injections of Fc vehicle or PB101 (50 mg/kg). After 14 days, the vascularity of the Matrigel plugs was evaluated through visual assessment and H&E staining of the tissue sections. Scale bars, 500 nm (top) for Matrigel plugs, and 100 (middle) and 1000 (bottom) μm for H&E. The boxed areas in the upper panels of the H&E-stained images are shown at higher magnification in the lower panels. ( F ) Immunohistochemical staining of the Matrigel plugs was performed using an anti-F4/80 Ab to assess macrophage infiltration. The macrophages were manually counted in three randomly selected fields per section of total nine sites per group. Scale bars, 100 μm. The bar graphs present the mean and SEM. ∗∗∗∗p < 0.0001 vs. untreated controls. ### p < 0.001 and #### p < 0.0001 vs. PB101-untreated positive controls. The p-values were determined via Kruskal–Wallis with Dunn's multiple comparisons test.

Article Snippet: Next, the cells were treated for 12 h with medium containing VEGF (200 ng/mL; catalog 100-20; Peprotech, Cranbury, NJ) or PlGF-2 (100 ng/mL; catalog 465-PL; R&D Systems, Minneapolis, MN) with or without PB101 (2, 5, or 10 μg/mL) and supplemented with 1% FBS for EA.hy926 cells and 0.1% FBS for RA-FLSs.

Techniques: Migration, Control, Transferring, Incubation, Recombinant, Matrigel Assay, Injection, Isolation, Transgenic Assay, Staining, Immunohistochemical staining

Journal: eBioMedicine

Article Title: The therapeutic effects of the VEGF decoy receptor fusion protein VEGF-Grab in chronic inflammatory diseases

doi: 10.1016/j.ebiom.2026.106216

Figure Lengend Snippet: Suppressive effects of VEGF-Grab on the aggressiveness of RA-FLSs. ( A ) Chemotactic migration of RA-FLSs in Transwell inserts. RA-FLSs (1 × 10 5 , n = 4) were loaded in the upper chambers of Transwell inserts. Culture media supplemented with 1% FBS and recombinant PlGF-2 (100 ng/mL), with or without PB101 (5 or 10 μg/mL), were loaded in the lower chamber. After 12 h, cells that migrated to the lower side of the Transwell membrane were manually counted. Scale bars, 1000 μm (upper panel) and 200 μm (lower panel). ( B ) Wounding migration assay with RA-FLSs. RA-FLSs (1.5 × 10 5 , n = 4) were seeded on 6-well plates, scratched with pipette tips, and treated with recombinant PlGF-2 (100 ng/mL), with and without PB101 (2 or 10 μg/mL), for 12 h. The cells that migrated across the reference lines (red dashed lines) were manually counted. Scale bars, 200 μm. The bar graphs present the mean and SEM. ∗∗p < 0.01 vs. untreated controls. # p < 0.05 and ## p < 0.01 vs. PlGF-treated positive controls. The p-values were determined via one-way ANOVA with Tukey's multiple comparisons test. ( C ) RA-FLS invasion into cartilage in a humanised synovitis model. RA-FLSs (2 × 10 6 ) were resuspended in a medium containing PlGF-2 (100 ng/mL) or a medium with PlGF-2 (100 ng/mL) and PB101 (2 μg/mL). These RA-FLSs were co-implanted with human cartilage into the left flanks (primary) of SCID mice (n = 3, total 6 mice). In the right flanks (contralateral), cartilage of the same size was implanted without RA-FLSs. For 60 days after cartilage implantation, the mice were injected intraperitoneally with vehicle or PB101 (2 mg/kg) twice a week. ( D ) Following excision, the cartilage samples were processed for H&E staining to evaluate invasion. The scoring protocol is detailed in the Materials and Methods section. For each implanted cartilage, three distinct regions were evaluated independently, and all regional values are shown as individual data points. Scale bars, 100 μm. The bar graphs present the mean and SEM. ∗p < 0.05 vs. vehicle controls. The p-values were determined via an unpaired t -test.

Article Snippet: Next, the cells were treated for 12 h with medium containing VEGF (200 ng/mL; catalog 100-20; Peprotech, Cranbury, NJ) or PlGF-2 (100 ng/mL; catalog 465-PL; R&D Systems, Minneapolis, MN) with or without PB101 (2, 5, or 10 μg/mL) and supplemented with 1% FBS for EA.hy926 cells and 0.1% FBS for RA-FLSs.

Techniques: Migration, Recombinant, Membrane, Transferring, Injection, Staining