Abstract
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<0.001) and progesterone (p<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 α2-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.
| Original language | English |
|---|---|
| Pages (from-to) | 501-521 |
| Number of pages | 21 |
| Journal | Steroids |
| Volume | 54 |
| Issue number | 5 |
| DOIs | |
| State | Published - Nov 1989 |
Bibliographical note
Funding Information:This manuscript represents both original, unpublished data and previously published information presented at the William J. LeMaire Reproductive Symposium, December 2,1988, Miami, Florida. The authors would like to thank Williams and Wilkins for the release of copyright information which has been included in this article. The didactic criticism of Dr. 1. Frederick Woessner throughout our studies on ovulation is kindly appreciated. The authors would like to thank Dr. Michael Vernon for performin the steroid RIA analysis. The expertise of Dr. Kathy Sharpe, Dr. Colom %o Giovanna, and Ms. Gail Sievert in rotein separation by SDS-el electrophoresis is ratefully acknowledge I! . The authors would also Bik e to ratefully than & Mr. Scott Estes, Ms. Marie Selzer, and Ms. Carol n Taplin 3o r their technical assistance and Ms. Darleen Chamberlain fort K e preparation of this manuscript. This work was supported by NIH grants HD-08747 and HD-06773 and by BRSG RR05374. Also, a note of tribute to Dr. David Puett for undertaking this compilation of scientific information which has been influenced by Dr. LeMaire. In closin a very heartfelt thank you to “Wim” for stimulating my personal an c3‘s cientific growth (TEC).
ASJC Scopus subject areas
- Biochemistry
- Molecular Biology
- Endocrinology
- Pharmacology
- Clinical Biochemistry
- Organic Chemistry