Effects of phorbol myristate acetate (PMA), dibutyryl cyclic
AMP (dbcAMP), 6-dimethylaminopurine (6-DMAP), okadaic acid
(OA), cycloheximide (CHX), actinomycin D (AcD) and tunicamycin
(TuM) on plasminogen activator (PA) activity and maturation rate
in bovine and porcine oocyte-cumulus cell complexes (BOCC and
POCC, respectively) in vitro were determined. Plasminogen
activator activity was measured by SDS-PAGE, casein-agar
zymography and densitometry. Three plasmingen-dependent lytic
zones (92-95, 71-73 and 49-51 kD) and one PA inhibitor (52 kD)
were observed in BOCC. Immunoprecipitation and amiloride
sensitivity suggested that the 49-51 kD protease is a urokinase
type PA (uPA), the 71-73 and 92-95 kD proteases are a tissue
type PA (tPA) and tPA-PAI complex, respectively, and the PAI is
PAI-1. In POCC, two plasminogen activators (71-78 and 93-96 KD)
were observed. Lack of amiloride sensitivity suggested that the 71
-78 kD band is a tPA and the 93-96 kD band is possibly a tPA-PAI
complex. Increasing dbcAMP in the culture medium increased
activity in both BOCC and POCC in dose-dependent fashion (P<0.05).
In BOCC cultured with PMA, total PA activity increased, however
high concentrations of PMA (10 and 100 ng/ml) decreased tPA
activity in matured POCC. Plasminogen activator activity
decreased in 6-DMAP, actinomycin D and cycloheximide and oocyte
maturation was also inhibited in these treatments. When POCC
were treated with 25 nM OA, uPA activity was observed.
Plasminogen activator activity increased in either BOCC or POCC
treated with up to 25 nM OA, however PA activity decreased at
concentrations greater than 75 nM (P<0.05). Incubation of BOCC
with tunicamycin reduced the molecular mass of tPA and tPA-PAI
complex and PAI-1 by 5-10%, however PA activity was not
inhibited. These data suggest that BOCC matured in vitro produce
uPA, tPA and PAI-1 however POCC produce only tPA and PAI. The
production of PA and PAI by either BOCC or POCC is associated
with oocyte maturation and influenced by stimulators of the
protein kinase A and C, modulators of intracellular phosphorylation
and metabolic inhibitors. / Graduation date: 1993
| Identifer | oai:union.ndltd.org:ORGSU/oai:ir.library.oregonstate.edu:1957/35942 |
| Date | 06 May 1993 |
| Creators | Kim, Nam-hyung |
| Contributors | Menino, Alfred R. Jr |
| Source Sets | Oregon State University |
| Language | en_US |
| Detected Language | English |
| Type | Thesis/Dissertation |
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