Changes in the bovine zona pellucida induced by plasmin or embryonic plasminogen activator

Effects of bovine plasmin and plasminogen activator recovered from bovine embryo-conditioned medium (bePA) on the polypeptide profile and solubility of bovine zonae pellucidae (ZP) were evaluated. ZP were isolated from bovine ovarian oocytes and incubated at 39 degrees C with 0, 100, or 200 microg/ml plasmin for 0, 24, or 48 hr or bePA with 0 or 100 microg/ml human plasminogen for 0 or 48 hr. ZP were evaluated either by SDS-PAGE or for changes in solubility using a zona pellucida dissolution time (ZPDT) assay. Two prominent polypeptides, molecular weight (MW) 76,000 and 65,000, and two minor polypeptides, MW 23,000 and 22,000, were resolved by SDS-PAGE. No changes occurred in the polypeptide profile for ZP incubated with 0 microg/ml plasmin for 0, 24, or 48 hr, and ZPDT did not differ (P > 0.10). Treatment with 100 or 200 microg/ml plasmin induced reductions in the MW 76,000, 23,000, and 22,000 polypeptides and the appearance of MW 45,000 and <10,000 polypeptides. ZPDT were less (P < 0.05) in 100 and 200 microg/ml compared with 0 microg/ml plasmin. Polypeptide profiles and ZPDT for ZP incubated with bePA were similar (P > 0.10) to ZP incubated with unconditioned medium. Addition of human plasminogen to ZP incubated with bePA reduced the MW 76,000, 23,000, and 22,000 polypeptides, caused the appearance of MW 45,000 and 20,000 polypeptides, and decreased ZPDT (P < 0.05). These results demonstrate that bovine plasmin is capable of proteolytically degrading the bovine ZP and that bePA can indirectly affect the ZP by converting plasminogen to plasmin.