Krzysztof Chwalisz

Regulation of the Uterus and Cervix during Pregnancy and Labor Role of Progesterone and Nitric Oxide

Preterm labor and subsequent preterm birth remain the major problems of perinatology overall because prematurity is the leading cause of infant mortality and morbidity. One of the reasons why efficacious and safe methods for the prevention and treatment of preterm labor are still not available is that the actual mechanisms responsible for the initiation of both term and preterm labor are unclear in primates. In the past, pregnancy research was focused on mechanisms responsible for the release of uterotonins (i.e., oxytocin, prostaglandins), with little attention paid to relaxation mechanisms which may be even more crucial during the initiation of parturition. Parturition is a relatively long-term process that involves the progression of uterine contractility from an inactive to a vigorously active state, and in addition a softening and opening of the rigid and closed cervix. These crucial steps (preparatory or conditioning phase and active labor) are thought to be regulated by a coordinated change in various systems including the interactions of endocrine, nervous, and immune control mechanisms. Experimental and clinical studies with progesterone antagonists clearly demonstrate the pivotal role of progesterone in establishing and maintaining pregnancy. More recently, nitric oxide (NO) has emerged as a possible element in controlling uterine quiescence and cervical functions (see ref. 1 for review). A major problem that remains in testing new strategies for the appropriate treatment or prevention of preterm labor is related to diagnostic uncertainty-that is, the lack of clear biophysical and biochemical indicators of the conversion from unproductive uterine contractions (false labor, pre-labor) to those capable of inducing progressive cervical dilatation. Moreover, this problem makes the analyses of tocolytic agents complicated and confusing. Hence, further progress in this area will not only depend on new tocolytic agents, but also demands the development of improved diagnostic methods. In this short review we focus on the role of progesterone and NO in controlling

The progesterone antagonist onapristone increases the effectiveness of oxytocin to produce delivery without changing the myometrial oxytocin receptor concentrations

The progesterone antagonist onapristone was used in guinea pigs during late pregnancy (43 +/- 2 days after coitus) and before term (day 61 after coitus) to investigate the role of progesterone on uterine reactivity to exogenous oxytocin, concentration of oxytocin receptors, and gap junctions in the myometrium. Onapristone priming increased the ability of oxytocin to induce delivery during late pregnancy and before term by factors of greater than or equal to 30 and approximately 10, respectively. The intrauterine pressure recording on day 43 after coitus revealed phasic, laborlike contractions in response to oxytocin in onapristone-treated animals, in contrast to tonic reactions in controls. The increase in the oxytocin response in onapristone-treated animals was not associated with an increase in myometrial oxytocin receptor concentrations either during late pregnancy or before term. By contrast, treatment with onapristone significantly decreased the input resistance of myometrial cells in guinea pigs in late pregnancy (43 +/- 1 day after coitus) to the level of animals at term. This was associated with a marked increase in myometrial gap junctions stained with antibodies against connexin 43. These results indicate that progesterone may control myometrial reactivity to oxytocin in pregnant guinea pigs by effects on postreceptor events mainly by suppressing the gap junctions.

Mechanism of Action of Antiprogestins in the Pregnant Uterus

Progesterone plays a crucial role in nearly the entire female reproductive process in mammals. It most likely controls the final stages of folliculogenesis and regulates ovulation. In addition, progesterone stimulates epithelial proliferation in the mammary gland and controls lactogenesis. During the preimplantation phase of the fertile cycle it prepares the endometrium for implantation by inducing endometrial receptivity. The sudden withdrawal of progesterone action at the end of the luteal phase leads to a constriction of spiral arteries and in turn induces menstruation in primates. During early pregnancy, progesterone is essential for the entire implantation process. In more advanced stages progesterone is responsible for pregnancy maintenance, acting predominantly on the myometrium and the uterine cervix. In addition, there is some evidence that progesterone has an immunosuppressive effect and may regulate the cytokine network in the uterus. Finally, progesterone may play an important role in the maternal adaptation of the cardiovascular system during pregnancy. During the preimplantation period and early pregnancy progesterone targets the endometrium. During later stages of pregnancy the myometrium and the uterine cervix become the major targets. There is ample evidence that during advanced pregnancy progesterone predominantly controls myometrial responsiveness by suppressing both the excitability and the propagation of the uterine muscle. The second important function of progesterone is the control of cervical ripening. Progesterone functions during preg-

Distribution of estrogen and progesteron receptors in the uterus: an immunohistochemical study in the immature and adult pseudopregnant rabbit.

SummaryIn order to clarify the distribution and content of estrogen (ER) and progesteron receptors (PR) under changing hormonal influences within the various cell populations of the uterus (glandular and luminal endometrial epithelium, stroma, myometrium), immunohistochemical determinations using specific monoclonal antibodies were made. To correlate the immunohistochemical findings with peripheral hormone levels and specific tasks of the endometrium, 17β-estradiol and progesterone serum levels were measured and cell proliferation determined by use of BrdU-labelling-immunohistochemistry. At the subcellular level ER and PR were located exclusively in the cell nuclei of female rabbits, which were either immature and lacking any peripheral hormone levels or were pseudopregnant (d0–d8 p.hCG). In the immature rabbits a general faint ER and PR immunostaining was found. In addition to a general increase in ER and PR in all cell populations estrous rabbits (d0 p.hCG) showed a significant rise of ER in the epithelial cells and of PR in the myometrium. Within the epithelial cells and the myometrium the ER dropped heavily within a few days of pseudopregnancy. The PR, however, increased sharply during the first two days of pseudopregnancy and decreased gradually following d4 p.hCG. A close relationship was observed between the high PR content and the proliferation rate of the epithelial cells on d2 p.hCG. In spite of the more rapid decrease of ER compared with PR, the glandular epithelium retained positive immunostaining. In the stroma the ER and especially PR content did not change significantly during the course of pseudopregnancy suggesting that some of the well-known differentiation events in the luminal epithelium may be mediated by the stroma.

Antiprogestin-Releasing Intrauterine Devices: A Novel Approach to Endometrial Contraception

Intrauterine devices (IUDs) that release progestins are highly effective contraceptives, but they induce breakthrough bleeding that some women find unacceptable. Because progesterone (P) antagonists [antiprogestins (APs)] are known to suppress the endometrium, induce amenorrhea and inhibit fertility, AP-releasing IUDs (AP-IUDs) may provide an effective contraceptive that also controls endometrial bleeding. Here, we assessed the effects of empty (blank) vs. AP-IUDs (ZK 230 211) on bleeding patterns and endometrial growth in ovariectomized, artificially cycled macaques. The AP-IUDs (but not the blank controls) induced extended, frank menstruation when inserted during the late luteal phase, an indication of local AP action. Over time, endometrial glandular and arterial proliferation were inhibited, steroid receptors were elevated, spiral arteries showed degenerative changes, P withdrawal bleeding was prevented, and estradiol (E2)dependent proliferation was suppressed by the AP-IUDs. In sum, AP-IUDs suppressed the effects of P on endometrial progestational development and blocked the effects of E2 on endometrial proliferation, as previously shown for systemic treatment with APs. Therefore, AP IUDs may provide novel contraceptive devices with minimal breakthrough bleeding.

Inhibition of aromatase activity in human placental microsomes by 13-retro-antiprogestins

Mifepristone (RU 486), used clinically for the termination of early pregnancy, and its acetyl and 13-retro (13 alpha) analogs show potent antiproliferative effects against estrogen-dependent human breast tumors and endometriosis. However, there has been no report on direct inhibition of aromatase by antiprogesterones. Aromatase inhibitors have been shown to be effective against estrogen-dependent breast cancer. We evaluated the inhibition of aromatase by various antiprogestins (ZK 112.993, ZK 98.734, ZK 114.043, ZK 98.299, and ZK 114.863). Human placental microsomes were incubated with [1 beta-3H,4-14C] androstenedione (3-114 nM) in the presence of NADPH, with or without putative inhibitors (10-200 microM). Aromatase activity was assessed by tritium release to water from the 1 beta-position of the substrate. ZK 112.993 and ZK 98.734 did not show any inhibitory effect. The statistical analysis of the data using standard errors was obtained from replicate experiments. ZK 114.043 showed slight inhibition with a Ki of 54.8 +/- 6.4 microM (m +/- SE, n = 6) against androstenedione aromatization. The two 13-retro-steroids, ZK 98.299 and ZK 114.863, showed aromatase inhibition with Ki values of 19.0 +/- 1.5 microM (n = 7) and 12.7 +/- 0.94 microM (n = 7), respectively, which is weak with respect to some known potent inhibitors, but significant when compared with the other antiprogestins which were tested. The results suggest that the unnatural 13-retro-antiprogestin conformation may have a better fit to the aromatase active site than the natural 13 beta-antiprogestin conformation. (Steroids 60:234-238, 1995).

Morphologische Aspekte der Cervixreifung unter Antigestageneinfluß

Der Einflus von ZK 98 299, einem Progesteronantagonisten, auf die Cervixreifung wurde an schwangeren Meerschweinchen elektronenmikroskopisch untersucht. Es zeigte sich eine massive Umgestaltung im kollagenen Bindegewebe mit Verminderung der kollagenen Fibrillen, Zunahme der Grundsubstanz sowie Aktivierung und Vermehrung der zellularen Komponente. Diese ZK 98 299 induzierten Effekte stimmen mit den physiologischerweise auftretenden morphologischen Veranderungen bei der Cervixreifung zum Zeitpunkt der Geburt uberein.