Inhibitors of Mammalian Tissue Collagenase and Metalloproteinases Suppress Ovulation in the Perfused Rat Ovary*

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Abstract
We have examined the effects of a new synthetic inhibtor of mammalian tissue collagenase, CI-1 (N-[3-N-(benzyloxycarbonyl)amino-1-(R)carboxypropyl]L-leucyl-O-methyl-L-tyrosine N-methylamide; G. D. Searle SC 40827), and a general metalloproteinase inhibitor, 1,10-phenanthroline, on ovulation, as judged by the observation of follicular rupture, and on progesterone production of the perfused rat ovary. Ovaries of PMSG (20 IU)-primed rats were perfused for 21 h, and samples of medium were taken for analysis of progesterone concentration. The number of ovulations was estimated by counting the number of oocytes released into the perfusion chamber. Ovaries were stimulated with LH (0.1 .mu.g/ml) plus 3-isobutyl-1-methylxanthine (IBMX; 0.2 mM), and this treatment resulted in a mean of 17.2 ovulations/treated ovary. 1,10-Phenanthroline dose-dependently inhibited ovulation, with 0, 0.2, and 12.5 ovulations/treated ovary at 1.0, 0.1, and 0.01 mM, respectively. This inhibition of ovulation closely paralleled the inhibition of extracted collagenase from uterus and ovary. However, 1,10-phenanthroline also suppressed progesterone release in a dose-dependent manner. Addition of the collagenase inhibitor (CI-1; 25 .mu.M) 1 h after LH plus IBMX inhibited ovulation (6.3 ovulations/treated ovary). Its relatively inactive stereoisomer (CI-2; 25 .mu.M) did not suppress ovulation(20.0 ovulations/treated ovary). CI-1 inhibited extracted uterine collagenase 50% at a concentration of 2 .mu.M, whereas CI-2 was only 1/15th as effective. There was an 80% loss of CI-1 from the medium during the perfusions. Neither CI-1 nor CI-2 had any effect on LH plus IBMX-stimulated progesterone release. These data demonstrate that the general metalloproteinase inhibitor 1,10-phenanthroline is able to inhibit ovulation, but also inhibits steroidogenesis. The more specific binding of collagenase, CI-1, can inhibit ovulation without affecting steroid production. These data indicate an important role for collagenase in the ovulatory process.