Electrophoretic characterization of the plasminogen activator produced by bovine blastocysts

Summary. Sodium dodecyl sulphate-polyacrylamide gel electrophoresis and zymog-raphy were used to determine the tissue source and to characterize the types of plasminogen activator (PA) produced by bovine blastocysts. Day 12\p=n-\14blastocysts were collected at slaughter from oestrus-synchronized, superovulated and artificially inseminated Holstein cows. In Expt 1, blastocysts were cultured for 24 h in Ham's F-12 in a humidified atmosphere of 5 % CO2 in air at 37\s=deg\C.After culture, blastocysts and medium were recovered and stored separately at \m=-\20\s=deg\C.In Expt 2, embryonic discs were separated from trophoblast by microdissection. Intact blastocysts, embryonic discs and trophoblast were then cultured for 24 h and recovered as in Expt 1. In both experiments, embryonic tissues and media were electrophoresed with PA and molecular mass standards. Polyacrylamide gels were laid onto casein\p=n-\agargel plates (zymograms) and incubated at room temperature for 24\p=n-\48h. Caseinolytic zones in zymograms containing plasminogen were evidence of PA. In Expt 1, bovine blastocysts contained and secreted light and heavy forms of PA (47\m=.\0\m=+-\1\m=.\0and 86\m=.\1 \m=+-\0\m=.\7kDa, respectively). In Expt 2, intact blastocysts and trophoblast produced both forms of PA (41\m=.\5\m=+-\1\m=.\5and 92\m=.\2\m=+-\2\m=.\7kDa) but PAs were not detected in embryonic discs. The results suggest that Day 12\p=n-\14bovine blastocysts produce urokinase-type PA (41\m=.\5\p=n-\ 47\m=.\0kDa) and a form of high molecular mass (86\m=.\1\p=n-\92\m=.\2kDa) that is either a novel tissue-type PA or a PA inhibitor which complexes with the lighter form