Journal: Biomimetics

Article Title: Developing New Peptides and Peptide–Drug Conjugates for Targeting the FGFR2 Receptor-Expressing Tumor Cells and 3D Spheroids

doi: 10.3390/biomimetics9090515

Figure Lengend Snippet: Determination of K D values from SPR Analysis.

Article Snippet: In our studies, SPR was used to investigate the binding interactions of the three peptides as well as that of the known FGF Receptor Tyrosine Kinase Inhibitor—CAS 192705-79-6 (Santacruz Biotechnologies, Dallas, TX, USA) with the kinase domain of FGFR2.

Techniques: Control

Journal: Experimental Physiology

Article Title: Platelet‐rich plasma affects gap junctional features in myofibroblasts in vitro via vascular endothelial growth factor (VEGF)‐A/VEGF receptor

doi: 10.1113/ep090052

Figure Lengend Snippet: FIGURE 1 Morphofunctional evaluation of NIH/3T3 fibroblast-to-myofibroblast transition. (a) Typical fibroblast (in PM, proliferation medium) impaled by the patch electrode (scale bar represents 20 μm). (b) Resting membrane potential (RMP, in millivolts; n = 4–8 cells; one-way ANOVA with Bonferroni’s correction, P = 0.074, d.f. = 56). Exact P-values of any comparison are reported in Table A1 for clarity. (c–m) Representative immunofluorescence analysis of α-smooth muscle actin (α-sma) and vinculin expression. Nuclei are counterstained with propidium iodide. Scale bars represent 10 μm. (n,o) Densitometric analysis of the fluorescence intensity (FI, in arbitrary units, a.u) of indicated markers. (p) Morphometric analysis of mean cell surface area. Abbreviations: Anti VEGF-A (vascular endothelial growth factor-A) Ab, anti-VEGF-A neutralizing antibodies; DM, differentiation medium; IgG, irrelevant isotype-matched IgG; KRN633, selective pharmacological VEGFR inhibitor; PM, proliferation medium; PRP, platelet-rich plasma; VEGF-A 2, 20, soluble VEGF-A 2 or 20 ng/mL. Values are the mean ± SD. Significance of differences in (n) (α-sma): *** versus PM (P < 0.0001); * versus PM (P = 0.05); §§§ versus DM (P < 0.0001); §§ versus DM (P = 0.007); ### versus DM+PRP (P < 0.0001); @@@ versus DM+DMSO (P < 0.0001); ˆˆˆ versus DM+PRP+DMSO (P < 0.0001); & versus DM+PRP+KRN (P = 0.019); ◦◦◦versus DM+PRP+IgG (P < 0.0001); + versus VEGF-A 2 (P = 0.033). Significance of differences in (o) (vinculin): *** versus PM (P < 0.0001); §§§ versus DM (P < 0.0001); § versus DM (P = 0.049); ### versus DM+PRP (P < 0.0001); @@@ versus DM+DMSO (P < 0.0001); ˆˆˆ versus DM+PRP+DMSO (P < 0.0001); ◦◦◦versus DM+PRP+IgG (P < 0.0001). In (n) and (o), one-way ANOVA with Tukey’s post hoc test, n = 60 regions of interest. Significance of differences in (p) (cell surface area, one-way ANOVA with Tukey’s post hoc test, n = 36 cells): *** versus PM (P < 0.0001); * versus PM (P = 0.026); §§§ versus DM (P < 0.0001); ### versus DM+PRP (P < 0.0001); @@@ versus DM+DMSO (P < 0.0001); ˆˆˆ versus DM+PRP+DMSO (P < 0.0001); ◦◦◦versus DM+PRP+IgG (P < 0.0001)

Article Snippet: In parallel, cells were cultured: (1) in DMplus a specific ATP-competitive inhibitor of VEGFR tyrosine kinase activity, KRN633 (SantaCruz Biotechnology, Santa Cruz, CA, USA; IC50 = 170 nM, Cat# sc-204379, Lot# B1810, dilution in DMSO, Sigma, Cat# 472301) in the absence or presence of PRP; (2) in DM in the presence of PRP plus mouse monoclonal anti-VEGF-A neutralizing antibodies (10 μg/mL; Sigma, Cat# V4758, Lot# 066K1428, RRID:AB_477621).

Techniques: Membrane, Comparison, Immunofluorescence, Expressing, Fluorescence, Clinical Proteomics

Journal: Experimental Physiology

Article Title: Platelet‐rich plasma affects gap junctional features in myofibroblasts in vitro via vascular endothelial growth factor (VEGF)‐A/VEGF receptor

doi: 10.1113/ep090052

Figure Lengend Snippet: FIGURE 2 Morphofunctional evaluation of human skin primary fibroblasts. (a) Typical human primary skin fibroblasts (in PM) impaled by a patch electrode (scale bar represents 10 μm). (b) Resting membrane potential (RMP, in millivolts; n = 4–7 cells; one-way ANOVA with Bonferroni’s correction, P = 0.007, d.f. = 29; **P < 0.01 and ***P < 0.001 vs. PM; §§P < 0.01 and §§§P < 0.001 vs. DM; ##P < 0.01 and ###P < 0.001 vs. DM+PRP) in the indicated experimental conditions. Exact P-values of any comparison are reported in Table A6 for clarity. (c,e,g,i,k,m) Representative immunofluorescence analysis of α-sma and Cx43 expression in human cells cultured as specified in Figure 1. Merge: superimposed DIC and fluorescence images (scale bar represents 25 μm). (d,f,h,j,l,n) Representative time course of Ij (in picoamperes) from cell pairs in the indicated experimental conditions. Abbreviations: α-sma, α-smooth muscle actin; Cx43, connexin 43; DIC, differential interference contrast; DM, differentiation medium; Ij, transjunctional current; KRN633, selective pharmacological VEGFR inhibitor; PM, proliferation medium; PRP, platelet-rich plasma; VEGF, vascular endothelial growth factor-A 2 ng/ml

Article Snippet: In parallel, cells were cultured: (1) in DMplus a specific ATP-competitive inhibitor of VEGFR tyrosine kinase activity, KRN633 (SantaCruz Biotechnology, Santa Cruz, CA, USA; IC50 = 170 nM, Cat# sc-204379, Lot# B1810, dilution in DMSO, Sigma, Cat# 472301) in the absence or presence of PRP; (2) in DM in the presence of PRP plus mouse monoclonal anti-VEGF-A neutralizing antibodies (10 μg/mL; Sigma, Cat# V4758, Lot# 066K1428, RRID:AB_477621).

Techniques: Membrane, Comparison, Immunofluorescence, Expressing, Cell Culture, Fluorescence, Clinical Proteomics

Journal: Breast Cancer Research : BCR

Article Title: erbB3 recruitment of insulin receptor substrate 1 modulates insulin-like growth factor receptor signalling in oestrogen receptor-positive breast cancer cell lines

doi: 10.1186/bcr3018

Figure Lengend Snippet: Western blot analysis . (a) Total and phosphorylated insulin-like growth factor type I receptor (IGF-IR), insulin receptor substrate 1 (IRS-1), erbB3, Akt and extracellular-signal regulated kinase 1/2 (ERK1/2) protein expression following incubation of MCF-7 and T47D cells in medium containing either the specific IGF-IR/IR tyrosine kinase inhibitor 4-anilino-5-bromo-2-[4-(2-hydroxy-3-( N , N -dimethylamino)propoxy)anilino]pyrimidine (ABDP) (1 μM) or appropriate vehicle control for 24 hours and subsequently challenged with increasing concentrations of heregulin β1 (HRGβ1) (0.1 to 10 ng/ml) or vehicle control for 5 minutes. Densitometric analysis of phosphorylated Akt and ERK1/2 protein levels in (b) MCF-7 and (c) T47D cells treated with HRGβ1 in the absence and presence of ABDP. The results are expressed as the means ± standard errors of the mean of at least three separate experiments from the 1 ng/ml HRGβ1-primed groups. (d) Western blot analysis of total IGF-IR, erbB3 and IRS-1 expression following immunoprecipitation (IP) with total IRS-1 antibody in MCF-7 and T47D cells incubated for 24 hours in medium containing ABDP (1 μM) or vehicle control. * P ≤ 0.05 versus HRGβ1 for both cell lines.

Article Snippet: To examine the effects of pharmacological blockade of IGF-IR, cells were incubated in phenol red-free (white) RPMI medium supplemented with 5% FCS and either the IGF-IR/IR tyrosine kinase inhibitor 4-anilino-5-bromo-2-[4-(2-hydroxy-3-( N , N -dimethylamino)propoxy)anilino]pyrimidine (ABDP) (1 μM in dimethyl sulphoxide, AstraZeneca, Macclesfield, UK) [ ] or appropriate vehicle control for 1 to 2 days.

Techniques: Western Blot, Expressing, Incubation, Control, Immunoprecipitation

Journal: Breast Cancer Research : BCR

Article Title: erbB3 recruitment of insulin receptor substrate 1 modulates insulin-like growth factor receptor signalling in oestrogen receptor-positive breast cancer cell lines

doi: 10.1186/bcr3018

Figure Lengend Snippet: Western blot analysis . (a) Total and phosphorylated insulin-like growth factor type I receptor (IGF-IR), insulin receptor substrate 1 (IRS-1), Akt, extracellular signal-regulated kinase 1/2 (ERK1/2) and β-actin protein expression following incubation of MCF-7 and T47D cells in medium containing either lipid and control siRNA (C si) mix (100 nM) or lipid and IRS si mix (100 nM) in the absence or presence of 4-anilino-5-bromo-2-[4-(2-hydroxy-3-( N , N -dimethylamino)propoxy)anilino]pyrimidine (ABDP) (1 μM) for 2 days and subsequently challenged with heregulin β1 (HRGβ1) (10 ng/ml) for 5 minutes. Data are representative of at least three separate experiments. The effects of 4-day treatment with HRGβ1 (10 ng/ml), ABDP (0.1 μM) or a combination of HRGβ1 and ABDP on the growth of (b) MCF-7 cells or (c) T47D cells. The results are expressed as the means ± standard errors of the mean of triplicate wells and are representative of at least three separate experiments. * P ≤ 0.01 versus control, † P ≤ 0.001 versus ABDP.

Article Snippet: To examine the effects of pharmacological blockade of IGF-IR, cells were incubated in phenol red-free (white) RPMI medium supplemented with 5% FCS and either the IGF-IR/IR tyrosine kinase inhibitor 4-anilino-5-bromo-2-[4-(2-hydroxy-3-( N , N -dimethylamino)propoxy)anilino]pyrimidine (ABDP) (1 μM in dimethyl sulphoxide, AstraZeneca, Macclesfield, UK) [ ] or appropriate vehicle control for 1 to 2 days.

Techniques: Western Blot, Expressing, Incubation, Control