BOVINE EMBRYOS PRODUCE A UROKINASE-TYPE PLASMINOGEN-ACTIVATOR

The type of plasminogen activator (PA) secreted by bovine embryos was identified. Day 12-14 embryos were collected from estrus-synchronized, superovulated, and naturally mated crossbred beef cows. Embryos were left intact (E) or microdissected into component embryonic discs (ED) and trophoblastic vesicles (TV). Intact embryos, ED, and TV were pre-cultured for 2 days in Minimum Essential Medium Alpha (MEM-alpha) with 10% heat-inactivated fetal calf serum, washed in serum-free MEM-alpha, and cultured individually for 5 days in 50-mu-l microdrops of MEM-alpha with 15 mg/ml bovine serum albumin. At 24 hr intervals, E, ED, and TV were observed for tissue morphology and transferred to fresh microdrops, and medium was recovered and frozen at -20-degrees-C. At the end of culture, blastocoelic fluid (BF) and embryonic tissues were recovered and frozen at -20-degrees-C. Plasminogen activator concentrations in medium, tissues, and BF were determined by using a caseinolytic assay. Antibodies to urokinase-type PA (anti-uPA) and tissue-type PA (anti-tPA), and the urokinase inhibitor, amiloride (AMR), were used to identify the type of PA produced by bovine embryonic tissues. Intact embryos and TV released more PA (P < 0.05) than ED, and tissues exhibiting expanded blastocoels released less PA (P < 0.05) than tissues with collapsed blastocoels. Blastocoelic fluid from TV exhibited more PA (P < 0.05) activity than from ED. Treatment with anti-uPA decreased PA activity (P < 0.05) in pooled medium and tissues from E compared to treatment with nonspecific immunoglobulins and anti-tPA. Amiloride completely eliminated PA activity (P < 0.05) in tissues, medium, and BF. These results suggest that day 12-14 bovine embryos secrete a urokinase-type PA.