Restriction of placental vasculature in a non-human primate: a unique model to study placental plasticity.

The limits of placental plasticity, i.e., the ability of the placenta to adapt and alter its growth trajectory in response to altered fetal requirements, are not known. We report fetal and placental hemodynamic adaptations in a novel non-human primate model in which the fetal inter-placental bridging vessels were surgically ligated. Doppler ultrasound studies showed that the rhesus placenta compensates for an approximate 40% reduction in functional capacity by increased growth and maintenance of umbilical volume blood flow. This unique experimental animal model has applications for mechanistic studies of placental plasticity and the impact on fetal development.

Relaxin stimulates interleukin-6 and interleukin-8 secretion from the extraplacental chorionic cytotrophoblast.

In the absence of infection, decidual relaxin (RLN) expression is increased in patients with preterm premature rupture of the membranes (PPROM) resulting in preterm birth, but it is not known whether inflammation stimulates RLN expression or vice versa. This study examined the effect of lipopolysaccharide (LPS) on the expression of RLN mRNA and secreted protein and whether RLN treatment influences secretion of proinflammatory cytokines from the fetal membranes. Explants of human fetal membranes in vitro and rhesus monkey fetal membranes in vivo were treated with LPS, which increased expression of IL-6 but had no effect on RLN. RLN treatment stimulated IL-6 and IL-8 secretion from choriodecidual explants in a subset of patients, as well as from isolated chorionic cytotrophoblast cells but not decidual cells. In vivo results obtained in rhesus monkeys after intra-amniotic infusion of RLN demonstrated increased IL-6 and IL-8 concentrations in amniotic fluid. Our results indicate that increased decidual RLN expression is independent of LPS but may induce a local sterile inflammatory process which potentially contributes to extracellular matrix degradation and weakening of the fetal membranes.

Localization of prostaglandin H synthase, prostaglandin dehydrogenase, corticotropin releasing hormone and glucocorticoid receptor in rhesus monkey fetal membranes with labor and in the presence of infection.

Prostaglandins (PGs) play a central role in primate parturition by their actions on uterine contractility and on cervical ripening. Rhesus monkey placentation is hemochorial and the endocrine events surrounding parturition are qualitatively similar to human pregnancy. Although there is an increase in PG production before the onset of labor, little is known about the cellular localization of the PGH synthase (PGHS) or the 15-hydroxy PG dehydrogenase (PGDH) in the fetal membranes of nonhuman primates and whether it changes at term in spontaneous labor or during preterm labor associated with infection. Placental corticotropin releasing hormone (CRH) and the glucocorticoid receptor (GR) have also been implicated as mediators in parturition by virtue of their roles in PG production. We utilized immunohistochemical methods to localize the inducible isoform PGHS-2, PGDH, GR and CRH in rhesus monkey amnion, chorion and attached decidua. Tissues were obtained at cesarean section during late pregnancy, in spontaneous labor at term and in premature labor induced by Group B streptococcal intraamniotic infection. Specific staining for immunoreactive (ir)-PGHS-2 was observed in amnion epithelial and mesenchymal cells and to a lesser extent in chorion and decidua. In contrast, ir-PGDH was localized primarily to the extravillous trophoblast layer of chorion. GR was localized to both the cytoplasm and nucleus of amnion epithelial cells, subepithelial fibroblasts, chorion trophoblasts and in decidua. Immunostaining for CRH was found in amnion and in scattered decidual cells but was most intense in the chorion trophoblast layer. There was no demonstrable change in this overall pattern of immunostaining in association with the onset of labor at term except for a decrease in staining for ir-PGDH in chorion. Experimental Group B streptococcal chorioamnionitis resulted in preterm labor and extensive necrosis of extravillous trophoblast cells with subsequent loss of chorionic ir-PGDH and relative sparing of ir-PGHS-2 in amnion epithelium which favors the net production of PGs. The expression pattern of these effectors in the rhesus monkey fetal membranes points to a functional role of PGs and glucocorticoids in the process of term and preterm parturition which is similar to that in human pregnancy.

Progesterone receptor localization and isoforms in myometrium, decidua, and fetal membranes from rhesus macaques: evidence for functional progesterone withdrawal at parturition.

OBJECTIVE: It is not known whether withdrawal of progesterone (P) action is a prerequisite for parturition in women or in nonhuman primates because concentrations of circulating progesterone or progesterone receptors (PR) in myometrium and decidua do not decrease before delivery. To examine this potentially important regulatory mechanism, we determined PR isoforms, PR localization, and mRNA in myometrium, decidua, and fetal membranes from rhesus monkeys during pregnancy and in spontaneous labor at term. METHODS: Gestational tissues were obtained midpregnancy (day 80-100), late pregnancy (day 130-145), and during spontaneous labor at term (day 161-167). Samples of rhesus monkey myometrium, decidua, chorion-decidua, and amnion were collected and analyzed for total nuclear and cytosolic PR by competitive binding assay. Progesterone receptor isoforms were identified and quantified by Western blot analysis, and PR mRNA was determined by a specific ribonuclease protection assay. Nuclear PR was localized by immunohistochemistry with monoclonal anti-PR (JZB39) after microwave stabilization. RESULTS: Myometrium and decidua showed no change in total PR during pregnancy and labor. Nuclear PR was not detected in fetal membranes by binding assay but was localized in amnion epithelial and mesenchymal cells and in chorion laeve cytotrophoblasts by immunohistochemistry. Staining for PR was substantially less by serial antibody dilution in fetal membranes than in decidua. Message for PR was confirmed in all tissues analyzed. A significant (P <.05) shift in the ratio of PR isoforms (from PR-B dominance at midpregnancy to PR-A dominance in labor) was observed in myometrium but not in decidua. Both PR-A and PR-B isoforms and PR nuclear staining were nearly undetectable in amnion obtained during labor. CONCLUSION: A shift to PR-A dominance in myometrium at term together with a loss of PR in fetal membranes provides evidence for a functional progesterone withdrawal mechanism, which may facilitate the initiation of parturition in primates.

Maternal or fetal origin of rhesus monkey (Macaca mulatta) amniotic fluid leukocytes can be identified by polymerase chain reaction using the zinc finger Y gene.

Leukocytes can be found in substantial numbers within the intrauterine tissues and amniotic fluid of women, and play a central role in the pathophysiology of infection-related preterm labor by their production of proinflammatory mediators. It remains unclear whether these leukocytes represent a fetal immune response, a maternal response, or a combination of the two. The objective of this study was to develop a test in the rhesus monkey (Macaca mulatta) suitable for determining the percentage of male fetal cells present in a population of leukocytes recovered from blood or amniotic fluid. We found inadequate specificity for rhesus monkey cells using commercial human Y-chromosome paint kits (fluorescence in situ hybridization (FISH)). Human-specific primers for the repetitive Y chromosome DYZ-1 locus employed in the polymerase chain reaction (PCR) produced an unacceptable percentage of false positives. However, we successfully developed a PCR-based test using rhesus-specific primers for the zinc finger Y (ZFY) locus. Densitometry of PCR products from known ratios of male and female adult peripheral leukocytes generated a linear standard curve which provided quantitative results and required only 400 cells per sample. The rhesus beta globin (RBG) gene served as an internal control. The PCR test correctly discriminated the sex of peripheral leukocytes in 20 adult males, 20 adult females, two male fetuses, and one female fetus. Serial samples of amniotic fluid from four chronically catheterized rhesus monkeys bearing male fetuses were used to confirm the utility of this assay for quantifying fetal cells in amniotic fluid. In conclusion, we have developed a PCR test which is suitable for distinguishing male from female cells in adult and fetal blood and in amniotic fluid, which lends itself to a variety of diagnostic and biologic applications in the rhesus monkey and potentially in other nonhuman primates.

Cervical cerclage in the second trimester of pregnancy: a historical cohort study.

OBJECTIVE: The purpose of this study was to compare second-trimester transvaginal cervical cerclage with conservative management on duration of pregnancy and perinatal outcome in patients with early or advanced cervical changes. STUDY DESIGN: A historical cohort analysis was performed. Maternal and neonatal records between 1995 and 1999 were retrospectively reviewed for women presenting between 18 and 27 weeks of gestation with early cervical changes (length <3 cm, dilatation <2 cm, funneling of fetal membranes shown by transvaginal ultrasonography) (group 1, n = 31) and for women with advanced cervical effacement and dilatation (cervical dilatation > or =2 cm but < or =5 cm, fetal membranes visible) (group 2, n = 39). In each group, patients who underwent Shirodkar or McDonald cerclage were compared with patients treated conservatively with bed rest. Both groups also received multifactorial treatment with tocolytic agents, broad-spectrum antibiotics, and indomethacin. Outcome variables were analyzed for statistical significance by parametric and nonparametric methods. RESULTS: Regardless of treatment method, patients with early cervical changes (group 1) were given a diagnosis earlier and delivered later in pregnancy compared with their counterparts who had advanced cervical changes (group 2) (P <.05). In both patients who underwent cerclage and those treated conservatively, the mean birth weight among surviving infants was higher and the mean neonatal intensive care unit stay was shorter in group 1 than in group 2 (P <.02). However, duration of maternal hospital stay and neonatal survival rates were not different. In both groups 1 and 2, the interval from treatment to delivery, the mean gestational age at delivery, and mean birth weight were increased, whereas neonatal intensive care unit stay was decreased by cerclage treatment (P <.05). In group 1, a higher percentage of patients treated with cerclage received antibiotics and indomethacin than did control subjects (P <.01), whereas in group 2, the use of multifactorial treatment was not different (P =.5). The duration of maternal hospital stay and neonatal survival did not differ significantly among patients treated conservatively or with cerclage. CONCLUSIONS: Diagnosis of premature cervical changes by ultrasonography was correlated with treatment earlier in gestation and with a favorable impact on perinatal outcome in both patients treated with cerclage and those treated conservatively. Cervical cerclage was associated with an improved perinatal outcome (in comparison with conservative therapy) in women with early cervical changes detected by ultrasonography and in patients with advanced cervical dilatation and visible membranes. However, the apparent therapeutic effect of cerclage in patients with mild cervical incompetence may be due in part to an increased use of antibiotics and indomethacin in conjunction with cerclage.

Indomethacin blocks interleukin 1beta-induced myometrial contractions in pregnant rhesus monkeys.

OBJECTIVE: We sought to determine whether blockade of prostaglandin synthesis with indomethacin prevents interleukin 1beta-induced increases in uterine contractions in a nonhuman primate model. STUDY DESIGN: Maternal and fetal vascular catheters, intra-amniotic fluid pressure catheters, and fetal electrocardiographic and myometrial electromyographic electrodes were implanted in 11 rhesus monkeys at 124 +/- 2 days' gestation (term, 167 days). After postsurgical stabilization (136 +/- 2 days) indomethacin 50 mg was administered orally twice daily for 5 days (n = 6). On day 3 human recombinant interleukin 1beta 10 microg was infused into the amniotic cavity over 2 hours. Five days after the last indomethacin dose the study was repeated without indomethacin treatment. Uterine activity was continuously monitored and quantified as the hourly contraction area (millimeters of mercury. seconds per hour) in the experimental group and a control group (n = 5) that received interleukin 1beta alone. At timed intervals amniotic fluid was sampled for leukocyte counts and assayed for prostaglandin E(2) and F(2alpha), the inflammatory cytokines interleukin 1beta, interleukin 6, interleukin 8, tumor necrosis factor alpha, and interleukin 1 receptor antagonist by specific assays. RESULTS: Uterine activity was increased severalfold from baseline after interleukin 1beta infusion alone and in the absence of indomethacin treatment (P <.05). There was no increase in uterine contractility when interleukin 1beta was infused concurrently with indomethacin treatment. Concentrations of amniotic fluid leukocytes and cytokines increased significantly after interleukin 1beta infusion in both the presence and absence of indomethacin. Amniotic fluid prostaglandins E(2) and F(2alpha) were suppressed during indomethacin treatment but rose significantly after interleukin 1beta infusion in the absence of indomethacin. Except for higher interleukin 6, cytokine levels were unaltered by indomethacin. CONCLUSIONS: After interleukin 1beta infusion, indomethacin blocked the development of uterine activity. Amniotic fluid prostaglandins were suppressed by indomethacin treatment, but cytokines and leukocytes were not. These results suggest that prostaglandins or possibly other indomethacin-suppressible compounds stimulate uterine activity after interleukin 1beta infusion in late-gestation rhesus monkeys or that indomethacin has direct tocolytic effects.

In vitro modulation of primate coronary vascular muscle cell reactivity by ovarian steroid hormones.

Susceptibility to drug-induced coronary vasospasm in rhesus monkeys increases after removal of the ovaries and can be normalized by adding back physiological levels of estradiol-17ss (E2) and/or natural progesterone (P) in vivo as reported recently by our group. Furthermore, the reactivity status (Ca2+ and protein kinase C responses) of freshly isolated and primary culture coronary artery vascular muscle cells (VMC) mimic the intact coronary artery responses to 5-HT + U46619. Since coronary reactivity is maintained in the isolated VMC, we hypothesized that the reactivity state inherent in the VMC was modulated directly by ovarian steroids in vitro as in the whole animal. To test this hypothesis, we treated hyperreactive VMC from ovariectomized (ovx) monkeys in vitro with E2 or P and measured VMC reactivity to combined stimulation with 5-HT and U46619, as determined by the amplitude and especially the duration of intracellular Ca2+ signals, as well as protein kinase C (PKC) activation/translocation. VMC were treated for 12 96 h with 3 100 pg/ml E2 (10 365 pM) and/or 0.3 3 ng/ml P (0.95 9.5 nM). Hyperreactive responses to the combination of 5-HT and U46619 in untreated VMC were significantly and dose-dependently reduced by treatment in vitro with physiological levels of either E2 or P for at least 24 h. Both the early transient and late sustained increases in intracellular Ca2+ and PKC translocation were blunted, and the effects of 0.2 nM E2 and 3.2 nM P were specifically antagonized by the receptor blockers ICI 182,780 (200 nM) and RU486 (15 nM), respectively. Antibodies to the estrogen receptor and progesterone receptor labeled nuclei in VMC, which were also positively labeled by a smooth muscle myosin heavy chain monoclonal antibody. These data indicate that natural ovarian steroids directly reduce hyperreactive 5-HT and thromboxane A2-stimulated Ca2+ and PKC responses of coronary artery VMC from surgically menopausal rhesus macaques. We hypothesize that vascular hyperreactivity, which may be a critical factor involved in the increased incidence of coronary artery vasospasm and ischemic heart disease in postmenopausal women, can be normalized by E2 and/or P through direct actions on coronary artery vascular muscle cells.

Amniotic fluid prolactin is decreased by experimental intrauterine infection or interleukin-1beta infusion but not via prostaglandins in pregnant rhesus macaques.

Amniotic fluid contains a high concentration of prolactin produced and secreted by the decidua. In vitro models have suggested that bacterial products inhibit prolactin secretion by decidual cells. To further examine this potentially important regulatory mechanism in the whole animal, chronically instrumented pregnant rhesus monkeys were prepared. Experimental infection was induced by intraamniotic or choriodecidual inoculation of 10(3)-10(6) group B streptococcus. Alternatively, interleukin (IL)-1beta was infused into the amniotic cavity. Finally, indomethacin was coadministered with IL-1beta to block the production of prostaglandins (PGs). The average prolactin level prior to inoculation (0 h) equaled 34.0 +/- 6.4 microg/ml. There was a 40% decrease in prolactin by 37 h postinfection (n = 6) and a 71% decrease between 61 and 72 h postinfection (n = 3, p < 0.01 vs. before infection). Infusion of IL-1beta also caused a decrease in amniotic fluid prolactin. There was a 42% decrease in prolactin between 0 and 24 h postinfusion (p < 0.05) and a 66% decrease between 25 and 72 h after IL-1beta infusion (p < 0.05; n = 6). Coadministration of indomethacin with IL-1beta prevented the accompanying increase in PGs but did not prevent the decrease in prolactin (n = 5). Amniotic fluid prolactin levels in untreated monkeys were stable and without a prepartum decline during the sampling period from 130 to 166 days of gestation. In summary, intrauterine bacterial infection decreases amniotic fluid prolactin, and IL-1beta mimics this effect. The effect of IL-1beta on amniotic fluid prolactin does not appear to be mediated by PGs and may involve a direct effect of IL-1beta on decidual cells.

Hysteroscopy, hysterosalpingography and tubal ostial polyps in infertility patients.

OBJECTIVE: To compare the findings in infertility patients who underwent preoperative hysterosalpingography (HSG) followed by hysteroscopy and to determine the incidence of tubal ostial polyps, their HSG appearance and the results of hysteroscopic resection in our patient population. STUDY DESIGN: Sixty-eight infertility patients were evaluated by HSG followed by hysteroscopy. HSG diagnoses were divided into groups: group 1, normal; group 2, bilateral tubal occlusion; group 3, unilateral tubal occlusion; group 4, filling defects; and group 5, abnormal cavity. HSG findings were compared to the hysteroscopy findings. For patients in whom tubal ostial polyps were found, the findings were described, including postsurgical interval to conception. RESULTS: The agreement rates were 90%, 50%, 69%, 73% and 71% for groups 1-5, respectively. The positive predictive value of an abnormal HSG was 65%, and the negative predictive value of a normal HSG was 90%. Six of 68 patients (11.3%) had polyps at the fallopian tube ostium. Three of these patients (50%) had had the finding of proximal tubal occlusion on the ipsilateral side predicted by HSG; three had had normal HSGs. Four of the six conceived following polypectomy. The mean interval from surgery to conception was 4.5 months. CONCLUSION: HSG was a specific but not sensitive predictor of uterine pathology in our patient population. Tubal ostial polyps may occur in a significant proportion of infertility patients and can cause proximal tubal occlusion on HSG. Their possible contribution to infertility and clinical significance deserve further investigation.

Inhibition and augmentation of progesterone production during pregnancy: effects on parturition in rhesus monkeys.

OBJECTIVES: Uterine quiescence during mammalian pregnancy is attributed to progesterone. However. systemic progesterone levels remain elevated in primates before parturition. Epostane, a selective 3beta-hydroxysteroid dehydrogenase inhibitor, and progesterone (with or without epostane) were administered to late pregnant rhesus monkeys to clarify the role of progesterone in primate parturition. STUDY DESIGN: On days 122 to 132 of gestation (term 167 days), 11 rhesus monkeys (Macaca mulatta) with timed pregnancies were divided into three treatment groups: (1) epostane alone (10 mg/kg subcutaneously), (2) epostane with progesterone subcutaneously in Silastic silicone rubber capsules, and (3) progesterone implants only with no surgical instrumentation. Maternal and fetal blood and amniotic fluid were sampled for progesterone, estrone, estradiol, cortisol, testosterone, androstenedione, dehydroepiandrosterone, dehydroepiandrosterone sulfate, and amniotic fluid was sampled for prostaglandins E2 and F2alpha. Uterine activity was monitored continuously by electromyography and intraamniotic pressure. Cervical status was assessed by a modified Bishop's score. Production of prostaglandins E2 and F2alpha by amnion was determined by tissue superfusion. The group of three noninstrumented monkeys, which received only progesterone Silastic silicone rubber implants subcutaneously at 146 to 148 days, were observed until spontaneous vaginal delivery. RESULTS: Epostane reduced maternal and fetal progesterone levels by 75% and 50%, respectively, followed by increased uterine activity and cervical ripening within 24 hours and vaginal delivery within 48 hours. Amniotic fluid progesterone decreased to undetectable levels. Progesterone implants prevented the epostane-induced decrease in maternal and fetal progesterone levels and the associated myometrial and cervical changes until the implants were removed. Alterations in other steroid hormones were consistent with inhibition of 3beta-hydroxysteroid dehydrogenase. Amniotic prostaglandin E2 production was increased sixfold by epostane (p < 0.05) but did not reach the high levels normally seen at spontaneous parturition. Animals that received progesterone implants alone had markedly elevated circulating progesterone concentrations yet were delivered spontaneously at term (range 163 to 167 days). CONCLUSIONS: Progesterone withdrawal induces preterm labor and delivery (which can be blocked by progesterone substitution) but exogenous progesterone, even in substantial quantities, does not prevent parturition at term.

Endocrine-immune interactions in pregnant non-human primates with intrauterine infection.

Preterm birth remains the most common cause of perinatal mortality. Although the causes of preterm labor are multifactorial and vary according to gestational age, preterm labor and term labor share common cellular and molecular mechanisms, including stimulation of the fetal hypothalamic-pituitary-adrenal (HPA) axis and endocrine/immune system interactions. We have developed a non-human primate experimental model for intrauterine infection and preterm labor using chronically instrumented rhesus monkeys (Macaca mulatta) with timed gestations. We have documented the temporal and quantitative relationships among intrauterine infection, the synthesis and release of proinflammatory cytokines, prostaglandins, and fetal-placental steroid biosynthesis in this model. Infection-induced preterm parturition is characterized by significant elevations in amniotic fluid proinflammatory cytokines and by increases in fetal adrenal steroid biosynthesis, but not by corresponding increases in placental estrogen biosynthesis characteristic of spontaneous parturition. This suggests that activation of the fetal HPA axis by the stress of infection is accompanied by placental dysfunction and also that infection-induced preterm parturition is not dependent upon the increased estrogen biosynthesis observed in spontaneous parturition. These different endocrine and immune responses have important diagnostic and therapeutic implications in the management of preterm labor.

Rapid loss of oestrogen and progesterone receptors in human leiomyoma and myometrial explant cultures.

Oestrogen and progesterone are promoters of uterine leiomyoma growth: oestrogen receptors (ER) and progesterone receptors (PR) are over-expressed in these tumours. Paradoxically, there is a heterogeneity in responsiveness of leiomyoma growth to oestrogen and progesterone in culture. In this study, leiomyoma and adjacent myometrium were obtained at hysterectomy. The effect of oestrogen and progesterone on steroid receptor maintenance was examined using minced explants. Quantitative enzyme-linked immunoassay and Northern analysis were performed to assess ER and PR protein and mRNA content respectively. There was an approximately 75% decrease in ER and PR protein content within 8 h of incubation in both leiomyoma and myometrium. The presence or absence of oestrogen and/or progesterone had no effect on receptor protein loss. Northern analysis indicated a parallel loss of ER and PR mRNA transcripts. These findings suggest that the ER and PR expression in leiomyoma may require other extracellular factors. In-vitro studies designed to test the effects of sex steroids and their respective inhibitors on growth and function of leiomyoma and myometrial cells should consider this phenomenon.

Fetal and maternal endocrine responses to experimental intrauterine infection in rhesus monkeys.

OBJECTIVE: Our purpose was to describe the temporal and quantitative relationships among intrauterine infection, fetal-placental steroid biosynthesis, and preterm labor in a nonhuman primate model. STUDY DESIGN: On approximately day 130 of gestation (term 167 days) chronically instrumented rhesus monkeys (Macaca mulatta) were infected with 10(6) colony-forming units of group B streptococci either by intraamniotic (n = 4) or choriodecidual (n = 2) inoculation. As controls, four additionally chronically instrumented noninfected monkeys were followed up to spontaneous parturition. Amniotic fluid and maternal and fetal arterial blood were serially sampled in all monkeys (both before and after infection) for progesterone, estrone, estradiol, dehydroepiandrosterone, dehydroepiandrosterone sulfate, androstenedione, and cortisol by specific radioimmunoassays, and uterine activity was continuously recorded. RESULTS: Spontaneous parturition was preceded by gradual and significant increases in the plasma concentrations of fetal dehydroepiandrosterone, dehydroepiandrosterone sulfate, and androstenedione and fetal and maternal levels of estrone, estradiol, and progesterone but not by changes in cortisol. In contrast, infection-associated parturition (either intraamniotic or choriodecidual) was characterized by abrupt increases in fetal dehydroepiandrosterone, dehydroepiandrosterone sulfate, androstenedione, progesterone, and cortisol but not by increases in maternal or fetal estrone or estradiol. Infection-associated steroid changes occurred concurrently with or after increases in uterine activity. CONCLUSION: Infection-associated preterm parturition is associated with dramatic increases in fetal adrenal steroid biosynthesis but not by corresponding increases in placental estrogen biosynthesis. This suggests that fetal stress in accompanied by placental dysfunction and that infection-associated parturition is not dependent on the increased estrogen biosynthesis observed in spontaneous parturition.

Interleukin-1 beta intra-amniotic infusion induces tumor necrosis factor-alpha, prostaglandin production, and preterm contractions in pregnant rhesus monkeys.

OBJECTIVE: To describe the temporal and quantitative consequences of intra-amniotic interleukin-1 beta infusion in a nonhuman primate model. METHODS: On days 128-138 of gestation (term 167 days), four chronically instrumented rhesus monkeys (Macaca mulatta) underwent serial intra-amniotic infusions of 2, 5, and 10-20 micrograms recombinant human interleukin-1 beta. Each infusion was for 2 hours, and subsequent infusions were at least 48 hours later. Amniotic fluid was sampled serially both before and after infusion for interleukin-1 beta, tumor necrosis factor-alpha (TFN-alpha), and prostaglandin (PG) E2 and F2 alpha by specific assays, and uterine activity in each monkey was recorded continuously. RESULTS: Intra-amniotic concentrations of interleukin-1 beta rose dramatically after infusion. This rise was rapidly followed by the appearance of TNF-alpha in the amniotic cavities of all animals, with maximal levels reached 5 hours after the initiation of the infusion. Both interleukin-1 beta and TNF-alpha were rapidly cleared from the amniotic fluid and returned to baseline levels by 24-48 hours. Increases in PGE2 and F2 alpha paralleled those of the two cytokines but remained elevated for the duration of the experiments. The stimulation of uterine contractility from a pre-infusion level of 200 mmHg. seconds/hour to 6000 mmHg. seconds/hour occurred an average of 6-10 hours after interleukin-1 beta infusion. These stimulations were transient, usually abating by 22 hours after infusion, and did not result in frank labor. CONCLUSION: In the rhesus monkey, intra-amniotic infusion of interleukin-1 beta rapidly induces production of intra-amniotic TNF-alpha as well as PGE2 and F2 alpha, followed by uterine contractility. Uterine activity diminishes as cytokine levels return to pre-infusion levels, even in the presence of elevated intraamniotic PG levels. Tumor necrosis factor-alpha may act synergistically with interleukin-1 beta in the pathophysiology of cytokine-related preterm labor.

Tubal surgery of IVF--making the best choice in the 1990s.

Although there are tubal causes of infertility for which surgery offers little or no chance of successful treatment, there are at least two situations-sterilization reversal and microsurgical or laparoscopic adhesiolysis in the absence of fimbrial damage and/or male factor-in which subsequent live birth rates are excellent, 60-80% for the former and 45-65% for the latter. An advantage of tubal reconstruction over IVF-ET, which is the only viable alternative in tubal infertility, is avoidance of the risks of the stimulated ovulation protocol and multifetal gestation. Of course, the demands of microsurgery or operative laparoscopy are stringent; and the decision to undertake tubal reconstruction instead of IVF-ET must be coupled with appropriate patient selection.

Insulin-like growth factor I promotes leiomyoma cell growth in vitro.

OBJECTIVE: Our purpose was to determine whether insulin-like growth factors I and II preferentially stimulate uterine leiomyoma cells versus myometrial cells in monolayer culture. STUDY DESIGN: Leiomyomas and normal myometrium were obtained at hysterectomy from five premenopausal women. Specimens were enzymatically digested for use in primary monolayer cell cultures. By use of serum-free media, insulin-like growth factor I or II was added in 1, 10, and 100 ng/ml concentrations to both cell types with the patient serving as her own control. Cell number, prolactin production, and proliferative index values were measured on day 15 of cell culture. RESULTS: Significant increases in cell number were found in the leiomyoma cultures (p < 0.05) treated with 10 and 100 ng/ml insulin-like growth factors I but not with insulin-like growth factors II. Neither factor exerted a stimulatory effect on myometrial cells. CONCLUSION: Insulin-like growth factors I preferentially stimulates leiomyoma cells in monolayer culture. These results suggest an autocrine-paracrine role in vivo for this factor in conjunction with gonadal steroids in promoting leiomyoma growth.

Estrogen receptor gene expression in human uterine leiomyomata.

Estrogen and progestin are believed to be important physiological regulators of uterine leiomyoma growth. We recently showed that progesterone receptor messenger ribonucleic acid (mRNA) and protein levels are increased in human uterine leiomyomas compared with those in myometrial biopsy tissue obtained from the same patient. To further characterize the molecular mechanisms underlying abnormal growth of uterine leiomyomas, we analyzed biopsy samples of tumor and adjacent normal myometrium for estrogen receptor (ER) gene expression. Northern analysis indicated that ER mRNA levels were increased 1.4-to 12.6-fold in leiomyoma compared with myometrium in all patients examined (n = 11), whereas beta-actin mRNA was not different between the two groups. The size of the primary ER mRNA transcript was 6.2 kilobases in both leiomyoma and myometrium, indicating no gross mutation of the ER gene. An ER protein of 66 kilodaltons was detected by Western blot analysis, and quantitative immunoassay of ER revealed 9448 +/- 1955 fmol/mg DNA in leiomyoma compared to 2827 +/- 979 fmol/mg DNA in myometrial tissue. Scatchard analysis of 17 beta-estradiol binding to cell-free extracts revealed enhanced binding capacity (per mg DNA) in leiomyoma tissue (n = 6) of about 6-fold, whereas ER binding affinity was not substantially different between the leiomyoma and adjacent myometrial tissues. We propose that increased expression of progesterone receptor in leiomyoma is most likely a consequence of overexpression of functional ER that results in increased end-organ sensitivity to estradiol.