The involvement of collagenolysis in ovulation in the rat.

Collagenolytic activity in ovarian follicles was previously demonstrated by using synthetic peptides and reconstituted collagen fibers. However, attempts to demonstrate degradation of ovarian collagen and to correlate collagenase activity with ovulation were not successful. By administration of L-(5-3H) proline, we have labeled ovarian and follicular collagen and followed collagenolytic activity by separation of 3H-hydroxyproline (3H-Hyp) from acid hydrolyzates of ovarian tissue by HPLC. The level of ovarian and follicular 3H-Hyp decreased by about 40% on the afternoon of proestrus or after exogenous stimulation of ovulation by human CG (hCG), and this decrease was abolished by blocking the surge of gonadotropins with Nembutal. To verify that the observed reduction in 3H-Hyp was due to the action of a typical collagenase, the collagenous fraction was prepared from ovarian tissue and from preovulatory follicles before and after the ovulatory stimulus. The extracts were treated with trypsin (25 min, 25 C, 0.01 mg/ml) plasmin and p-amino-phenyl-mercuric acetate to fully activate the collagenase extracted along with collagen. Both, enzymatic and chemical activation of collagenase in vitro resulted in degradation of collagen. This degradation could be inhibited by cysteine and EDTA; both are classic inhibitors of mammalian collagenases. The activity of ovarian collagenase increased within 3 h after hCG-stimulation, peaked at 5-fold 6 h after hCG, and declined afterwards. Administration of cysteine (0.001-0.01 mmol) into the bursal cavity of proestrous rats blocked ovulation and breakdown of ovarian collagen in a dose-dependent manner. Cysteine effectively inhibited ovulation even when injected 7 h after the hCG stimulus. Inhibitors of arachidonic acid metabolism prevent ovulation. Indomethacin (inhibitor of cyclooxygenase) and nordihydroguaiaretic acid (inhibitor of lipoxygenase) blocked ovulation and inhibited hCG-induced ovarian collagenolysis. Collectively, these results corroborate the essential role of collagenolysis in follicular rupture in mammals.

[1]  R. Miskin,et al.  Follicular plasminogen activator: involvement in ovulation. , 1985, Endocrinology.

[2]  Z. Naor,et al.  Possible involvement of lipoxygenase products of arachidonic acid pathway in ovulation. , 1983, Prostaglandins.

[3]  S. Murao,et al.  Effect of various proteinase inhibitors on ovulation of explanted hamster ovaries. , 1983, Journal of reproduction and fertility.

[4]  J. M. Marsh,et al.  Collagen, collagenase and collagenolytic activity in rat Graafian follicles during follicular growth and ovulation. , 1983, Biochimica et biophysica acta.

[5]  A. Bleecker,et al.  Automated fluorometric amino acid analysis: the determination of nonprotein cyclic imino acids. , 1982, Analytical biochemistry.

[6]  L. Espey.  Ovulation as an inflammatory reaction--a hypothesis. , 1980, Biology of reproduction.

[7]  J. M. Marsh,et al.  A microassay for the direct demonstration of collagenolytic activity in Graafian follicles of the rat. , 1978, Biochimica et biophysica acta.

[8]  R. Crystal,et al.  Activation of alveolar macrophage collagenase by a neutral protease secreted by the same cell , 1976, Nature.

[9]  L. Espey,et al.  Factors which influence ovulatory regradation of rabbit ovarian follicles. , 1976, Biology of reproduction.

[10]  Sidney Strickland,et al.  Ovarian plasminogen activator: Relationship to ovulation and hormonal regulation , 1975, Cell.

[11]  H. Birkedal‐Hansen,et al.  Trypsin activation of latent collagenase from several mammalian sources. , 1975, Scandinavian journal of dental research.

[12]  J. M. Marsh,et al.  Post ovulatory changes in the concentration of prostaglandins in rabbit Graafian follicles. , 1974, Prostaglandins.

[13]  P Rondell,et al.  Role of steroid synthesis in the process of ovulation. , 1974, Biology of reproduction.

[14]  L. Espey.  Ovarian proteolytic enzymes and ovulation. , 1974, Biology of reproduction.

[15]  H. R. Lindner,et al.  Ovulation rate and serum LH levels in rats treated with indomethacin or prostaglandin E2. , 1973, Prostaglandins.

[16]  U. Zor,et al.  Physiological role of prostaglandins in the induction of ovulation. , 1972, Prostaglandins.

[17]  U. K. Laemmli,et al.  Cleavage of Structural Proteins during the Assembly of the Head of Bacteriophage T4 , 1970, Nature.

[18]  L. Espey.  Ultrastructure of the apex of the rabbit graafian follicle during the ovulatory process. , 1967, Endocrinology.

[19]  J. F. Woessner,et al.  The determination of hydroxyproline in tissue and protein samples containing small proportions of this imino acid. , 1961, Archives of biochemistry and biophysics.

[20]  S. Schochet.  A suggestion as to the process of ovulation and ovarian cyst formation , 1916 .

[21]  D. Armstrong,et al.  Blockade of spontaneous and LH-induced ovulation in rats by indomethacin, an inhibitor of prostaglandin biosynthesis. I. , 1972, Prostaglandins.