recombinant active human pai 1  (Innovative Research Inc)

Journal: bioRxiv

Article Title: The apoptotic, inflammatory, and fibrinolytic actions of vasoinhibin are in a motif different from its antiangiogenic HGR motif

doi: 10.1101/2023.08.18.553934

Figure Lengend Snippet: (a) Diagram of the secondary structure of prolactin with 199 residues (PRL), the 123-residue (Vi1-123), and the 48-residue (Vi1-48) vasoinhibin isoforms. The location and sequence of the linear HGR-containing vasoinhibin analog (Vi45-51) and the cyclic retro-inverse HGR-containing vasoinhibin analog (CRIVi45-51) are illustrated. The antiangiogenic HGR motif is highlighted in cyan. ( b ) Effect of 100 nM PRL, Vi1-123, Vi1-48, Vi45-51, or CRIVi45-51 on the proliferation of HUVEC in the presence or absence of 25 ng mL −1 VEGF and 20 ng mL −1 bFGF. Values are means ± SD relative to total cells ( n = 9). (c) Effect of 100 nM PRL, Vi1-123, Vi1-48, Vi45-51, or CRIVi45-51 on the invasion of HUVEC in the presence or absence of 25 ng mL −1 VEGF. Values are means ± SD relative to VEGF stimulated values ( n = 9). * P <0.0001 vs. VEGF+bFGF (-) or VEGF (-) controls (Two-way ANOVA, Dunnett’s). (d) Apoptosis of HUVEC in the absence (-) or presence of 100 nM of PRL, Vi1-123, Vi1-48, Vi45-51, or CRIVi45-51 ( n = 6 ). (e) Leukocyte adhesion to a HUVEC monolayer in the absence (-) or presence of 100 nM of PRL, Vi1-123, Vi1-48, Vi45-51, or CRIVi45-51 ( n = 9 ). (f) Lysis of a plasma clot by tissue plasminogen activator (tPA) alone or together with the plasminogen activator inhibitor-1 (PAI-1) in the presence or absence of PRL, Vi1-123, Vi45-51, or CRIVi45-51 ( n = 3 ). (g) Fibrinolysis relative to tPA and tPA+PAI-1 calculated with the area under the curve (AUC) of (f) . (h) Binding of PAI-1 to immobilized PRL, Vi1-123, Vi1-48, Vi45-51, or CRIVi45-51 in an ELISA-based assay ( n = 6 ). Individual values are shown with open circles. Values are means ± SD, * P <0.001 vs. control without treatment (-) (One-way ANOVA, Dunnett’s).

Article Snippet: Human recombinant plasminogen activator inhibitor 1 (PAI-1) was from Thermo Fisher Scientific (Waltham, MA) and human tissue plasminogen activator (tPA) from Sigma Aldrich (St. Louis, MO).

Techniques: Sequencing, Lysis, Binding Assay, Enzyme-linked Immunosorbent Assay

Journal: bioRxiv

Article Title: The apoptotic, inflammatory, and fibrinolytic actions of vasoinhibin are in a motif different from its antiangiogenic HGR motif

doi: 10.1101/2023.08.18.553934

Figure Lengend Snippet: (a) Diagram of the sequence and localization of the oligopeptides scanning the 48-residue vasoinhibin isoform (Vi1-48). The α-helix 1 (H1), the inflammatory, apoptotic, and fibrinolytic sequence (HNLSSEM), and the antiangiogenic HGR motif are highlighted in bold. (b) Effect of 100 nM 123-residue vasoinhibin isoform (Vi1-123) or the scanning oligopeptides on the proliferation of HUVEC stimulated with 25 ng mL −1 of VEGF and 20 ng mL −1 of bFGF. Values are means ± SD relative to total cells (n=9). (c) Effect of 100 nM Vi1-123 or the scanning oligopeptides on the invasion of HUVEC stimulated with 25 ng mL −1 of VEGF. Values are means ± SD relative to VEGF stimulated values ( n = 9 ). * P <0.0001 vs. VEGF+bFGF (-) or VEGF (-) controls. (d) Effect of 100 nM of Vi1-123, Vi1-48, or the scanning oligopeptides on HUVEC apoptosis ( n = 6 ). (e) Effect of 100 nM of Vi1-123, Vi1-48, or the scanning oligopeptides on the leukocyte adhesion to a HUVEC monolayer ( n = 9 ). (f) Dose-response of the leukocyte adhesion to HUVEC after treatment with the 20-35 or the 30-45 oligopeptides. Curves were fitted by least square regression analysis ( n = 6 , * P = 0.02, Paired T-test). (g) Lysis of a plasma clot by tissue plasminogen activator (tPA) alone or together with plasminogen activator inhibitor-1 (PAI-1) in the presence or absence of the scanning oligopeptides ( n = 3 ). (h) Fibrinolysis relative to tPA and tPA+PAI-1 calculated with the area under the curve (AUC) of (g) . (i) Binding of PAI-1 to the immobilized scanning oligopeptides in an ELISA-based assay ( n = 6 ). Individual values are shown with open circles. Values are means ± SD of at least 3 independent experiments, * P <0.001 vs. without treatment (-) controls (One-way ANOVA, Dunnett’s).

Article Snippet: Human recombinant plasminogen activator inhibitor 1 (PAI-1) was from Thermo Fisher Scientific (Waltham, MA) and human tissue plasminogen activator (tPA) from Sigma Aldrich (St. Louis, MO).

Techniques: Sequencing, Lysis, Binding Assay, Enzyme-linked Immunosorbent Assay

Journal: bioRxiv

Article Title: The apoptotic, inflammatory, and fibrinolytic actions of vasoinhibin are in a motif different from its antiangiogenic HGR motif

doi: 10.1101/2023.08.18.553934

Figure Lengend Snippet: Apoptosis of HUVEC (a) and leukocyte adhesion to a HUVEC monolayer (b) in response to the 123-residue vasoinhibin isoform (Vi1-123), the HNLSSEM-containing 20-35 or 30-45 oligopeptides in the absence (-) or presence of antibodies against β-tubulin (anti-β-tub), PAI-1 (anti-PAI-1) or uPAR (anti-uPAR), or the NF-κB inhibitor BAY117085. Values are means ± SD, n ≥ 3 , # P <0.001 vs. respective untreated (-) group, * P <0.0001 vs. absence of antibodies (-) (Two-way ANOVA, Dunnett’s). (c) Schematic representation to illustrate that the HNLSSEM motif promotes fibrinolysis by binding to PAI-1 and endothelial cell apoptosis and inflammation via the activation of uPAR and NF-κB.

Article Snippet: Human recombinant plasminogen activator inhibitor 1 (PAI-1) was from Thermo Fisher Scientific (Waltham, MA) and human tissue plasminogen activator (tPA) from Sigma Aldrich (St. Louis, MO).

Techniques: Binding Assay, Activation Assay