Plasminogen activators in tissues of the immature and estrogen-stimulated rat uterus and in uterine luminal fluid: characterization and properties.

We have characterized the molecular properties of the plasminogen activators in different cell types comprising the immature and the estrogen-stimulated rat uterus and in rat uterine luminal fluid. There were two plasminogen activators in the immature (day 20) rat uterus with apparent molecular weights, determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, of 70,000 and 46,000. Both plasminogen activators were present in epithelial and in stromal plus myometrial cell fractions of the immature uterus, and after stimulation by 17 beta-estradiol, no new plasminogen activators were detected in either cell fraction. The Michaelis constants (Km) for the activation of dog plasminogen by extracts from epithelial cells and from stromal plus myometrial cells obtained from either immature or 17 beta-estradiol-stimulated uteri were similar (approximately 11 microM). The maximal velocity (Vmax), normalized to protein concentration, increased 2.5-fold in the stromal plus myometrial cell fraction and 6.5-fold in the epithelial cell fraction, upon hormone stimulation (2 micrograms 17 beta-estradiol/day X rat for 3 days). The greatest concentration of plasminogen activator activity was found in the luminal fluid from estrogen-stimulated uteri, where the Vmax per mg protein was more than 10-fold greater than that in the cell fractions from estrogen-stimulated uteri. The plasminogen activator activity of luminal fluid was inhibited by diisopropyl fluorophosphate and rho-nitrophenyl rho-guanidinobenzoate, was not inhibited by human alpha-1-proteinase inhibitor and human antithrombin III, and was inhibited by high, but not low, concentrations of soybean trypsin inhibitor and bovine pancreatic trypsin inhibitor. These studies indicate that the plasminogen activators in different cell types comprising the uterus are similar and show that the estrogen enhancement of uterine plasminogen activator activity is the result of an increase in Vmax. The presence, upon hormone stimulation, of an apparent concentration gradient of increasing plasminogen activator activity through the uterus from myometrium to epithelium to luminal fluid may be a reflection of the dynamic role of this protease in the physiology of the uterus.

Uterine plasminogen activator activity: modulation by steroid hormones.

We have used a sensitive and quantitative assay to investigate the hormonal regulation of plasminogen activator (PA) activity in the rat uterus. PA activity is increased 5-fold (per U protein or DNA) by low physiological (0.1 micrograms) doses of estradiol, with increases in activity first observed at approximately 12 h. The stimulation of PA activity shows strict specificity among the steroid hormones, being stimulated by estrogens only or by high doses of dihydrotestosterone, which are known to affect the estrogen receptor system, and this stimulation is suppressed markedly by triphenylethylene antiestrogens. Comparative dose-response studies with a variety of estrogens of different uterotropic potencies indicate a good correlation between the potencies of different estrogens in stimulating PA activity and uterine growth (diethylstilbestrol = 17 beta-estradiol greater than estrone = 17 alpha-estradiol greater than estriol), with the exception of the zearalanol estrogen P-1496, which was consistently a potent stimulator of PA activity while being a very weak uterotropic agent. These studies suggest that increases in uterine PA levels may serve as a good marker of estrogen action in the uterus. Although the role of PA in uterine function remains unknown at present, its relatively large increase (up to 25-fold increase in content per uterus) may play a role in tissue remodeling during uterine growth.