Journal: PLoS ONE

Article Title: Mechanisms of thrombin-Induced myometrial contractions: Potential targets of progesterone

doi: 10.1371/journal.pone.0231944

Figure Lengend Snippet: (A-C) Immunofluorescence of PAR1 (green) in myometrium from a pregnant woman (A) , non-pregnant woman ( B ), and fetal membrane (C) . Nuclei were stained with DAPI (blue). Am, amnion, Cho, chorion, Deci, decidua. (D-G) Localization of hemorrhage, thrombin, and PAR1 in placental abruption at 25 weeks of gestation (D and E) , and 33 weeks gestation (F and G) resulting in disseminated intravascular coagulopathy and uterine bleeding requiring hysterectomy for hemostasis. (D and F) Hematoxylin and eosin staining of the myometrium adjacent to the placenta. Note that hemorrhage infiltrated the myometrium. Bars, 50 μm. (E and G) Immunofluorescence of PAR1 (green), thrombin (red), and DAPI (blue) at the same location of (A) . Bars, 50 μm.

Article Snippet: Primary antibodies used and concentration were as follows: thrombin (coagulation factor II, Novus Biologicals, NBP1-58268, Research Resource Identifier (RRID): AB_11023777, 1:100) and PAR1 (N2-11, Novus Biologicals, NBP1-71770, RRID: AB_11027203, 1:100).

Techniques: Immunofluorescence, Membrane, Staining

Journal: PLoS ONE

Article Title: Mechanisms of thrombin-Induced myometrial contractions: Potential targets of progesterone

doi: 10.1371/journal.pone.0231944

Figure Lengend Snippet: Thrombin increased contraction of primary human myometrial cells through PAR1. (A and B) (Left images) Representative images of collagen lattice assay of human myometrial cells at 30 min treated with PBS (Ctl) and thrombin (A) or PAR1 activating peptide, TFLLR (B) . (Right graphs) Quantification of myometrial contractions in collagen lattice assays (n = 3). **, p < 0.01 at each time point. (C) Collagen lattice assay of myometrial cells at 30 min with 2 U/mL of thrombin (Thr) pretreated with or without 100 nM PAR1 selective inhibitor (SCH79797, PAR1-i) for 1 h (n = 3). Representative image (upper panel) and quantification of gel areas (lower graph). The experiments were repeated three times. *, p < 0.05, and **, p < 0.01.

Article Snippet: Primary antibodies used and concentration were as follows: thrombin (coagulation factor II, Novus Biologicals, NBP1-58268, Research Resource Identifier (RRID): AB_11023777, 1:100) and PAR1 (N2-11, Novus Biologicals, NBP1-71770, RRID: AB_11027203, 1:100).

Techniques:

Journal: PLoS ONE

Article Title: Mechanisms of thrombin-Induced myometrial contractions: Potential targets of progesterone

doi: 10.1371/journal.pone.0231944

Figure Lengend Snippet: (A) Immunocytochemistry of PAR1 (red) and phosphorylated MLC2 (Ser19, green) in human myometrial cells. (B, C) Immunoblots of phosphorylated MLC2 (p-MLC2), total MLC2, and β-actin of myometrial cells. Myometrial cells were treated with thrombin (2 U/mL) as a function of time (B) , or pretreated with 100 nM PAR1 inhibitor (SCH79797) for 1 h, and then treated with 2 U/mL of thrombin for 30 min (C) . The experiments were repeated three times.

Article Snippet: Primary antibodies used and concentration were as follows: thrombin (coagulation factor II, Novus Biologicals, NBP1-58268, Research Resource Identifier (RRID): AB_11023777, 1:100) and PAR1 (N2-11, Novus Biologicals, NBP1-71770, RRID: AB_11027203, 1:100).

Techniques: Immunocytochemistry, Western Blot

Journal: PLoS ONE

Article Title: Mechanisms of thrombin-Induced myometrial contractions: Potential targets of progesterone

doi: 10.1371/journal.pone.0231944

Figure Lengend Snippet: (A) Collagen lattice assay of myometrial cells at 30 min with 2 U/mL of thrombin (Thr) pretreated with 1 μM progesterone (P4) for 1 h. Representative image (upper panel) and quantification of gel areas (lower graph). (B) Inhibition of thrombin-induced increases of PTGS2 , IL1B , and F2R mRNA by P4. Myometrial cells were pretreated with 1 μM of P4 for 1 h, and then treated with 2 U/mL of thrombin. (C) Gene expressions of progesterone receptor-A and–B ( PgR-A and PgR-B ) with 24 h treatment of 1 U/mL of thrombin, 10 nM of PGE2, and 10 nM of PGF2α (upper graphs) and PgR-A to PgR-B ratio (lower graphs). n = 3 in each group. *, p < 0.05, and **, p < 0.01. The experiments were repeated three times.

Article Snippet: Primary antibodies used and concentration were as follows: thrombin (coagulation factor II, Novus Biologicals, NBP1-58268, Research Resource Identifier (RRID): AB_11023777, 1:100) and PAR1 (N2-11, Novus Biologicals, NBP1-71770, RRID: AB_11027203, 1:100).

Techniques: Inhibition

Journal: Frontiers in Bioengineering and Biotechnology

Article Title: Novel Designed Proteolytically Resistant VEGF-B186R127S Promotes Angiogenesis in Mouse Heart by Recruiting Endothelial Progenitor Cells

doi: 10.3389/fbioe.2022.907538

Figure Lengend Snippet: Characterization of the designed proteolytically resistant VEGF-B186R127S. (A) Graphical representation of VEGF-B186R127S. (B) Representative image of VEGF-B protein expression following HeLa cell transduction. (C) BaF3-VEGFR1 cell survival and proliferation assay. Each dot represents mean ± SD from two technical replicates from a representative experiment . (D) Receptor binding properties of different VEGF-B forms. Each lane represents the precipitation of ligands with hVEGFR1, hNRP1, hNRP2, or no receptor (blank). Red arrow signs indicate the full-length and proteolytically processed VEGF-B186, VEGF-B186R127S, or VEGF-A165.

Article Snippet: VEGF-B, VEGF-A165, or CMV (control)–containing media from adenoviral vector transduced HeLa cells was pre-cleared with Pierce Protein A/G Magnetic Beads (Thermo Scientific, 88802) and then combined with Fc-tagged soluble hVEGFR1 (R&D systems, 321-FL-050/CF), hNRP1 (R&D systems, 10455-N1-050), or hNRP2 (R&D systems, 2215-N2-025) recombinant proteins and incubated overnight at 4°C.

Techniques: Expressing, Transduction, Proliferation Assay, Binding Assay