Identification of the plasminogen activator inhibitor produced by the day 14 bovine embryo

Graduation date: 2014Day 12-14 bovine embryos produce urokinase-type plasminogen activator (uPA) and\ud an uPA-plasminogen activator inhibitor (PAI) complex. The type of PAI, -1 or -2, in\ud the complex has not been determined therefore the objective of this research was to\ud identify the PAI produced by the Day 14 bovine embryo. In experiment 1, Western\ud blot analysis was used to determine the presence of PAI-1 and/or PAI-2 in the Day 14\ud bovine embryo. In experiment 2, a caseinolytic-agar gel immunoneutralization assay\ud was used to determine the concentration of PA in pooled Day 14 embryo-conditioned\ud medium incubated with primary antibodies directed against PAI-1 or PAI-2\ud previously used in the Western blotting. Western blot analysis detected two PAI-1\ud bands in embryos (61 and 76 kD) and only the high molecular mass form in\ud conditioned medium (75 kD). Likewise, Western blotting detected two PAI-2 bands\ud in embryos (52 and 66 kD) and a high molecular mass form in conditioned medium\ud (61 kD). In the immunoneutralization assay, PA activity was greater (P< 0.05) in\ud conditioned medium following treatment with anti-PAI-1 or anti-PAI-2 compared to\ud treatment with non-specific immunoglobulins. These data suggest Day 14 bovine\ud embryos produce both PAI-1 and PAI-2