The role of ovarian proteases and their inhibitors in ovulation.
To assess the role of inhibitors of proteolytic enzymes, such as plasminogen activator (PA) and collagenase in the ovulatory process, inhibitor activity and mRNA levels were examined in periovulatory rat and human ovaries. In the rat, immature animals received 20 IU of pregnant mare serum gonadotropin (PMSG) followed 52 h later by 10 IU of hCG. Ovaries were removed at intervals from 0 to 20 h after human chorionic gonadotropin (hCG) administration. Inhibitor activity for metalloproteinases, such as collagenase, increased from 60.5 +/- 4.1 inhibitor units/ovary at 0 h (i.e., time of hCG treatment) to a maximum of 218.2 +/- 11.4 units/ovary at 8 h after hCG before decreasing at 12 h (time of ovulation) and 20 h (122.2 +/- 7.9 and 71.6 +/- 8.1 units/ovary, respectively). Human follicular fluid and granulosa cells were obtained from preovulatory follicles of patients in our in vitro fertilization program. Metalloproteinase inhibitor activity was evaluated in follicular fluid as well as the levels of PA and PA inhibitor (PAI) mRNA by Northern analysis. Increasing metalloproteinase inhibitor activity was positively correlated with follicular levels of estradiol (p less than 0.001) and progesterone (p less than 0.02, N = 26). Chromatographic separation of follicular fluid resulted in two peaks of metalloproteinase inhibitor activity. The large molecular weight (MW) inhibitor had an approximate size of 700 kilodaltons (kDa) and may represent alpha 2-macroglobulin, a serum-derived inhibitor. The small MW inhibitor shared many of the characteristics of tissue-derived inhibitors of metalloproteinases. Partial purification of the small MW inhibitor by Concanavalin A-Sepharose and Heparin-Sepharose chromatography demonstrated the inhibitor to be a glycoprotein with an approximate MW = 28-29 K. Northern analysis of human granulosa cell total RNA from preovulatory follicles showed little or no detectable tissue-type PA or urokinase-type PA mRNA. In contrast, two species of PA inhibitor type-1 mRNA were detected in relative abundance. The present findings demonstrate the presence of proteolytic inhibitors in periovulatory ovaries of the rat and human. These ovarian inhibitors may play a role in regulating connective tissue remodeling during follicular rupture.
Curry
T E
TE
Department of Obstetrics & Gynecology, University of Kentucky, Lexington 40536.
Dean
D D
DD
Sanders
S L
SL
Pedigo
N G
NG
Jones
P B
PB
D000818:Animals; D006063:Chorionic Gonadotropin / Q000494:pharmacology; D005260:Female; D015571:Follicular Fluid / Q000201:enzymology; D006064:Gonadotropins, Equine / Q000494:pharmacology; D006107:Granulosa Cells / Q000187:drug effects / Q000201:enzymology; D006801:Humans; D008666:Metalloendopeptidases / Q000378:metabolism; D003012:Microbial Collagenase / Q000378:metabolism; D010053:Ovary / Q000187:drug effects / Q000201:enzymology*; D010060:Ovulation; D010447:Peptide Hydrolases / Q000378:metabolism*; D012333:RNA, Messenger / Q000032:analysis; D051381:Rats; D011919:Rats, Inbred Strains
D006063:Chorionic Gonadotropin; D006064:Gonadotropins, Equine; D012333:RNA, Messenger; D010447:Peptide Hydrolases; D008666:Metalloendopeptidases; D003012:Microbial Collagenase