Activation of Plasminogen by the Early Bovine Embryo1

Abstract

Activation of the plasma zymogen plasminogen to the enzyme plasmin by the early bovine embryo was evaluated. Sixteen-cell embryos to early morulae were collected at death from handmated synchronized and superovulated crossbred beef cows. Embryos were cultured in Ham's F-12 medium supplemented with 15 mg/ml bovine serum albumin containing 0, 15, 30, 60 or 120 micrograms/ml plasminogen in a humidified atmosphere of 5% CO2 in air at 37 degrees C. Cultures were observed every day, and stage of development was recorded. Medium was collected at 24-h intervals, starting at initiation and continuing through 288 h of culture. Plasminogen activator and plasmin levels in the culture media were determined, using a caseinolytic assay. The percentages of embryos developing to the initiating hatching blastocyst, hatched blastocyst, attached blastocyst, and attached blastocyst with trophoblastic outgrowth stages were not significantly different between the five levels of plasminogen. Initiation and completion of hatching, however, accelerated as plasminogen concentration increased in the culture media. Plasminogen activator production, expressed as milliunits X ml-1 X h-1 X viable embryo-1, was low for the first 48 h of culture, increased between 48-120 h, and tended to plateau thereafter. Plasminogen activation, measured indirectly as the plasmin concentration in a microdrop of medium and expressed as microgram plasmin X ml-1 X h-1 X viable embryo-1, followed plasminogen activator production, and was consistently low for the first 48-72 h of culture. Embryonic activation of plasminogen increased sharply thereafter, and also plateaued after 120 h.