The effect of prostaglandin I2 on the contractility of the term pregnant human myometrium

Small myometrial strips were dissected from the upper and lower segments of the term pregnant human uterus. The specimens were superfused in organ chambers and contractile activity was recorded isometrically.In strips from the upper segment, prostacyclin (PGI₂), induced an initial excitatory response followed in the majority of experiments by transient inhibition. In the lower segment the response was generally the same although direct inhibition without initial stimulation occurred in some cases.During the period of inhibition the specimens were refractory to iterated exposure to PGI₂. Furthermore, during this period of PGI₂-induced inhibition the muscle strip was also refractory to PGE₂ but responded to PGF_2α and oxytocin by stimulation.After inhibition of spontaneous contractile activity induced by indomethacin PGI₂ induced an excitatory response.The results do not indicate any critical change in the myometrial responsiveness of the upper uterine segment to PGI₂ during labor. In strips from the lower segment obtained before labor there tended to be a dominance of non-responders and inhibition only as compared to the results during labor. Nevertheless, whether or not PGI₂ under physiological or pharmacological conditions has any significant influence on the contractility of the term pregnant human uterus, still remains obscure.As judged from earlier reports from our laboratory and the present study it is evident that the uterine vessels are considerably more sensitive to the action of PGI₂ than the myometrium.     

Leukotrienes and myometrial activity of the term pregnant uterus

Biopsies from different segments of the pregnant human uterus were superfused in organ chambers and contractile activity was registered. Leukotriene C4(LTC4) caused inhibition of spontaneous but not noradrenaline induced contractile activity in strips from the cervix. This effect occurred both in early pregnancy and at term. However, the lower and the upper uterine segment of the term pregnant uterus did not respond to LTC4. The results represent a documentation of the segmental differentiation in the uterine response to eicosanoids.     

Production of peptide leukotrienes in endotoxin shock

Arachidonate metabolites are potent mediators generated in endotoxin shock. Following endotoxin administration (15 mg/kg) into unanesthetized rats, we found a rapid biliary secretion of peptide leukotrienes. Analysis of bile for peptide leukotrienes included organic solvent extractions, reversed phase-HPLC, radioimmunoassay (RIA), and spectrophotometry. The major immunoreactive endogenous leukotriene (LT) from bile was eluted between LTC₄ and LTD₄ in three chromatographic systems. It corresponded thereby to a biliary metabolite of injected LTC₄ and LTD₄ which in turn showed the ultraviolet spectrum of a peptide leukotriene. This demonstration of endotoxin-induced generation of peptide LTs in vivo was possible by sequential HPLC and RIA analyses in bile into which peptide LTs are eliminated from blood.     

Elevated levels of uterine anti-apoptotic signaling may activate NFKB and potentially confer resistance to caspase 3-mediated apoptotic cell death during pregnancy in mice

Preserving the uterus in a state of relative quiescence is vital to the maintenance of a successful pregnancy. Elevated cytoplasmic levels of uterine caspase 3 during pregnancy have been proposed as a potential regulator of uterine quiescence through direct targeting and disabling of the uterine contractile architecture. However, despite highly elevated levels of uterine caspase 3 during pregnancy, there is minimal evidence of apoptosis. This current study defines the mechanism whereby the pregnant uterine myocyte may harness the tocolytic activity of active caspases while avoiding apoptotic cell death. Using the pregnant mouse model, we have analyzed the uterus for changes in pro- and antiapoptotic signaling patterns associated with the advancing stages of pregnancy. Briefly, we have found that members of the IAP family, such as SURVIVIN and XIAP, and the Bcl2 family members, such as MCL1, are elevated in the uterine myocyte during late gestation. The IAP family members are the only endogenous inhibitors of active caspase 3, and MCL1 limits activation of caspase 3 by suppressing proapoptotic signaling. Elevated XIAP levels partner with SURVIVIN, resulting in increased levels of the antiapoptotic MCL1 via NFKB activation; these together have the potential to limit both the activity and level of active caspase 3 in the pregnant uterus as term approaches. We propose that modification of these antiapoptotic signaling partners allows the pregnant uterus to escape the apoptotic action of elevated active caspase 3 levels but also functions to limit the levels of active uterine caspase 3 near term.     

Expression of the calcium-activated potassium channel in upper and lower segment human myometrium during pregnancy and parturition

Background  

Large conductance calcium-activated potassium channel (BKCa) plays an important role in the control of uterine contractility during pregnancy. The change from uterine quiescence to enhanced contractile activity may be associated with the spatial and temporal expression of BKCa within myometrium. The objectives of this study were to examine the expression of BKCa alpha- and beta-subunit in upper segment (US) and lower segment (LS) regions of uterus, and to investigate for the possibly differential expression of these proteins in US and LS myometrium obtained from three functional states: (1) non-pregnant (NP); (2) term pregnant not in labour (TNL) and (3) term pregnant in labour (TL).     

The effect of prostaglandin I on the contractility of the term pregnant human myometrium

Small myometrial strips were dissected from the upper and lower segments of the term pregnant human uterus. The specimens were superfused in organ chambers and contractile activity was recorded isometrically. In strips from the upper segment, prostacyclin (PGI2), induced an initial excitatory response followed in the majority of experiments by transient inhibition. In the lower segment the response was generally the same although direct inhibition without initial stimulation occurred in some cases. During the period of inhibition the specimens were refractory to iterated exposure to PGI2. Furthermore, during this period of PGI2-induced inhibition the muscle strip was also refractory to PGE2 but responded to PGF2 alpha and oxytocin by stimulation. After inhibition of spontaneous contractile activity induced by indomethacin PGI2 induced an excitatory response. The results do not indicate any critical change in the myometrial responsiveness of the upper uterine segment to PGI2 during labor. In strips from the lower segment obtained before labor there tended to be a dominance of non-responders and inhibition only as compared to the results during labor. Nevertheless, whether or not PGI2 under physiological or pharmacological conditions has any significant influence on the contractility of the term pregnant human uterus, still remains obscure. As judged from earlier reports from our laboratory and the present study it is evident that the uterine vessels are considerably more sensitive to the action of PGI2 than the myometrium.     

Estrogen effects on NADH oxidase and superoxide dismutase in prepubertal female rats

Thirty-four day old, ovariectomised rats were treated with increasing doses of estradiol, 2-hydroxyestradiol 2,3-dimethyl ether (23E₂), 4-hydroxyestradiol 3,4-dimethyl ether (34E₂) and 4-methoxy-estradiol (4ME₂) for five days by subcutaneous injection. Superoxide dismutase, phenol activated NADH oxidase and uterine dry weights were determined. Only estradiol was found to be uterotrophic and increased NADH oxidase activity in these experiments. Both 23E₂ and 34E₂ treatment reduced the enzyme activity significantly. Though 4ME₂ showed a decrease in NADH oxidase at 0.05μg/100gm body weight there was no further decrease at higher dose (5μg/100gm). The Superoxide dismutase (SOD) in uterus and liver was unaffected by estradiol, while 23E₂, 34E₂, and 4ME₂ significantly reduced SOD in both liver and uterus. These results indicate that 23E₂, 34E₂ and 4ME₂, in spite of their non-uterotrophic property, affect uterine metabolism. Furthermore, in view of the reports indicating the importance of SOD levels in various tumors and since catecholestrogens are observed to reduce SOD levels in liver and uterus, it is suggestive that catecholestrogens may play an important role in the pathophysiology of certain tumors.     

The effect of the antioxidant, butylated hydroxy anisole, on peroxide-induced and spontaneous activity of the uterus from the pregnant rat

Chorioamnionitis is associated with preterm labor. Leukocytes infiltrate infected tissue and secrete hydrogen peroxide (H2O2) and other reactive oxygen products as part of their bactericidal activity. We have therefore investigated the effect of H2O2 on activity of in vitro uteri from pregnant rats. Uteri from 18-day pregnant rats exposed to H2O2 showed a dose-dependent increase in both contractile activity and production of prostaglandins (PG) E2 and F2 alpha compared to untreated controls. The antioxidant butylated hydroxy anisole (BHA) inhibited the H2O2-induced uterine activity. Furthermore, BHA inhibited contractions and PG production from spontaneously contracting uteri from 21-day pregnant rats. H2O2 increased chemiluminescence of uterine tissue, an index of oxygen or lipid radical formation, whereas BHA inhibited this effect. The BHA inhibition of uterine activity was reversed by addition of PGE2 to the incubation chamber. These data support the hypothesis that reactive oxygen can regulate PG production by the uterus and suggests a role for reactive oxygen in infection-induced labor and perhaps normal term labor as well.     

Rapid extraction of leukotrienes from biologic fluids and quantitation by high-performance liquid chromatography

Previous methods for the recovery and quantitation of leukotrienes have involved tedious extraction procedures, and high-performance liquid chromatographic (HPLC) techniques with significant limitations. We have designed a method to extract leukotrienes from biologic fluids using commercially available silica mini-columns requiring minimal preparation. Sample clarification is followed by a sensitive and reproducible HPLC technique which separates and quantifies the leukotrienes LTC₄, LTD₄, LTB₄ (and at least three of their isomers). The entire procedure requires less than one hour per sample.     

Opposite effects of ecdysone and 20-hydroxyecdysone on in vitro uterus motility of a tsetse fly

In order to examine the possible effects of ecdysteroids on parturition, we studied in vitro the influence of ecdysone and 20-hydroxyecdysone on the motility of isolated uterus from virgin and pregnant female tsetse fly (Glossina fuscipes). Ecdysone initiates phasic uterine contractions or enhances the frequency of preexisting contractile activity. In contrast, uterine contractions are decreased or abolished by 20-hydroxyecdysone. Pharmacological data indicate that tsetse fly uterus exhibits myogenic and nerve-evoked contractions. Ecdysteroids mainly act on nervous structures that control muscle contractions. Our results provide evidence for a specific action of ecdysteroids on a nerve-muscle target involved in female reproduction.     

Thromboxane B2, 6-keto-prostaglandin F1 alpha, and prostaglandin F2 alpha production by contracting pregnant rate uteri in vitro

While prostaglandin production by uterine tissue has been shown to be involved in the contractile mechanism of this tissue, less attention has focused upon the involvement of other prostanoids. We have simultaneously measured in vitro isometric contractility of pregnant rat uteri with the release of prostaglandin F2 alpha (PGF), 6-keto-prostaglandin F1 alpha (6-k-PGF1 alpha) and thromboxane B2 (TXB2) into the bathing medium under various conditions. Frequency of uterine contractions and integrated contractile force (ICF) increased from 15 days of gestation and peaked at the time of parturition. Activity was generally greatest during the first 15 min of incubation except during parturition and on Day 1 postpartum when the uterine segment remained active for 1 h experimental period. Indomethacin (INDO) significantly reduced contractile activity regardless of gestational stage. PGF, TXB2, and 6-k-PGF1 alpha increased with gestational age, peaking at the time of parturition. Production was greatest during the first 15 min of incubation and INDO inhibited production of each prostanoid regardless of gestational stage. Imidazole (100 micrograms/ml) inhibited TXB2 production without affecting PGF or 6-k-PGF1 alpha levels. Frequency of contraction and ICF were not affected by imidazole treatment despite TXB2 reduction. These data demonstrate that the in vitro uterus from pregnant rats is capable of producing prostanoids other than prostaglandins and their production generally parallels uterine contractile activity. Thus, the possibility that these prostanoids are involved in physiologic changes during parturition warrants further investigation.     

Morinda lucida reduces contractility of isolated uterine smooth muscle of pregnant and non-pregnant mice.

The present work investigated the effect of Morinda lucida (M. lucida) extract on isolated uterine smooth muscle of pregnant and non-pregnant mice. Pregnant and non-pregnant mice were pretreated with oral stilboesterol (0.1 mg/kg body weight) and killed by cervical dislocation. Thin strips of the uterus were cut and mounted in a 20ml organ bath containing De Jalon solution bubbled with 95%O2-5% CO2 gas mixture. The strips were connected to a force transducer coupled to a Grass 7D Polygraph for the recording of isometric tension. Effects of graded concentrations of oxytocin (OXY; 10-5-10-2 mol/L), acetylcholine (ACh; 10-9-10-5 mol/L) and M. lucida extract (0.015-1.5 mg/ml) were recorded. Fresh uterine strips were then incubated with M. lucida extract for 5mins and cumulative response to OXY was repeated. Another set of fresh strips was incubated in L-NAME for 15mins and the cumulative responses to M.lucida extract were repeated. OXY resulted in increased contractile responses in both pregnant and non-pregnant uterine muscles. M. lucida resulted in relaxation of the uterine smooth muscle in both pregnant and non-pregnant mice at all doses. However, at 1.5mg/ml, M. lucida completely blocked spontaneous uterine contractions. Following incubation with L-NAME, M. lucida extract led to a slightly greater relaxation of the uterine strips. In conclusion, M. lucida reduced contractility of uterine smooth muscle in both pregnant and non-pregnant mice as well as blocking contractile responses to OXY and Ach in uterine smooth muscle of pregnant and non-pregnant mice. There was no significant alteration of M. lucida activity by L-NAME suggesting that the action of the compound on uterine muscle is not associated with impaired nitric oxide synthase.     

Functional and histochemical characterization of a uterine adrenergic denervation process in pregnant rats

The time course of pregnancy-induced changes in the contractile responses of isolated uterine rings and sympathetic innervation pattern were studied using electric field stimulation and histofluorescence techniques, respectively, in intact and 6-hydroxydopamine-treated rats. Neurally mediated contractions elicited by field stimulation (0.6 msec, 1-70 Hz, 40 V) were measured in uterine preparations obtained from nonpregnant, 6-hydroxydopamine-treated and 5-, 10-, 15-, 18-, and 22-day (term) pregnant rats. At all frequencies, the amplitudes of contractions were highest in nonpregnant uteri. Stimulation at 1-2.5 Hz evoked contractions in 10-day pregnant uteri but failed to cause contractions on Day 5 and from Day 15 onward. In uterine preparations obtained from term and from 6-hydroxydopamine-treated rats, contractions could not be evoked by stimulation at 1-20 Hz. Fluorescence histochemistry of uterine adrenergic nerves revealed rich perivascular and myometrial innervation in nonpregnant and in pregnant rats through Day 10. Degeneration and loss of adrenergic nerve fibers was apparent by Day 15, and fluorescent myometrial and perivascular nerves were practically absent by Day 22. These findings demonstrate a progressive, frequency-related reduction of nerve-mediated uterine contractions beginning in midterm pregnancy, in parallel with a gradual loss of adrenergic nerve fibers. Pregnancy-induced nerve degeneration may promote the development of nonsynaptic alpha-adrenergic uterine contractile activity towards term. The reduced responsiveness of uterine smooth muscle to electric field stimulation in early pregnancy appears to be unrelated to alterations in uterine innervation but may be related to changes associated with implantation.     

The effects on stretching and prostaglandin F2alpha on the contractile and bioelectric activity of the uterus in rat.

The effects of stretching and of prostaglandin F2alpha on spontaneous and induced with local deformation contractile activity of rat uterus were studied. It was found that stretching the uterus up to the length of the decontracted organ in vivo increased the contractile and bioelectric activity. The rise in spontaneous uterine activity was a factor reducing induction of additional contractions. The fall in the level of endogenous prostaglandins in the uterus following administration of indomethacin inhibited the spontaneous contractile activity but was without effect on the contractions induced by local deformation of the myometrium. Prostaglandin F2alpha added to the bath in a concentration of 0.001 ng/ml exerted an inhibitory action on the different tested parameters of the contractile activity. After high doses stimulation of the uterine activity was observed.     

Effects of taurine on calcium binding and accumulation in rabbit hippocampal and cortical synaptosomes

The effect of taurine on Ca²⁺ binding and uptake was studied with rabbit brain cortical and hippocampal synaptosomes. Taurine (25 mM) increased by 25% the high affinity ⁴⁵Ca²⁺ binding in the cortical fraction and by 55% in hippocampal synaptosomes but had no effect on low affinity Ca²⁺ binding. Taurine decreased significantly the fluorescence of the chlorotetracycline-hydrophobic Ca²⁺ chelate probe in both synaptosomal fractions which suggests a shift of bound Ca²⁺ from the hydrophobic to the hydrophilic part of the membranes. The uptake of ⁴⁵Ca²⁺ by rabbit brain synaptosomes, when measured in control and 65 mK K⁺-containing media, was not influenced by taurine. However, taurine inhibited significantly the ⁴⁵Ca²⁺ uptake in synaptosomes incubated in media containing moderately increased K⁺ concentrations (14 and 20 mM K⁺). The effects of taurine are discussed in conjunction with its stabilizing effect on excitable membranes.     

Progesterone concentration and reproduction: An evaluation in pregnant, heat-acclimated, white rats

1. 1.|Progresterone concentrations in peripheral plasma, corpus luteum, uterus and placenta were determined by radioimmunoassay, in pregnant heat-acclimated (HA) and control (C) rats.

2. 2.|Concentration of progesterone in plasma and uterus of HA rats were lower than C from day 5 to 18 and day 5 to mid-gestation respectively.

3. 3.|Number and weight of pups born to HA rats were lower than C by 11.1% and 12.3% respectively.

4. 4.|It is concluded that aberration in pregnancy development observed in HA rats is associated with lower progesterone concentration in plasma and uterus from day of implantation to mid-pregnancy.

Neutral metalloendopeptidase associated with the smooth muscle cells of pregnant rat uterus

The pregnant rat uterus contains a membrane-bound metalloendopeptidase that is biochemically and immunologically similar to kidney enkephalinase (E.C.3.4.24.11). The uterus enzyme readily cleaved specific neutral endopeptidase substrates and oxytocin as well as the synthetic elastase substrate, Suc(Ala)3-pNA, yet did not digest native elastin. Using specific inhibitors, the uterus endopeptidase was identified as a metallopeptidase and not a serine protease, having an absolute requirement for zinc and perhaps calcium for maximal activity. The uterus endopeptidase cross-reacted with polyclonal antiserum to kidney microvillar endopeptidase and a monoclonal antibody to common acute lymphocytic leukemia antigen. Immunohistochemical localization of the enzyme in a 17 day pregnant uterus indicated that the enzyme was localized on the smooth muscle bundles of the myometrium and the endometrial epithelium. Total enzyme activity was 25 times higher in the late-term pregnant uterus (17th day of pregnancy) than in the nonpregnant uterus. Enzyme levels dropped rapidly prior to parturition and within 4 days after delivery the enzyme activity had returned to control levels. Inhibition of NEP in uterine strips with phosphoramidon resulted in a marked potentiation of oxytocin-induced contractions. Our results suggest that the uterine endopeptidase may have an important role in regulating uterine smooth muscle cell contraction during the later stages of pregnancy through its action on oxytocin and perhaps other biologically active peptides.     

Estradiol regulation of secretory component: Expression by rat uterine epithelial cells

Sex hormones are known to play an important role in the regulation of mucosal immunity in the female reproductive tract. The purpose of this study was to examine the effect of estradiol (E₂) on secretory component (SC) expression by epithelial cells in the rat uterus and to determine whether SC mRNA is present in uterine tissues and is under hormonal control. When ovariectomized rats treated with E₂ for 3 days and sacrificed 12 h after the last injection, expression of SC on luminal and glandular epithelial cells, as determined by immunohistochemistry, was elevated when compared to control animals. To determine whether E₂ regulation of SC involves mRNA synthesis, uterine RNA was extracted and analyzed by Northern blot. These experiments demonstrated that SC RNA is present in uteri from intact rats and markedly increased when ovariectomized animals are treated with E₂. In other studies, uterine epithelial cells from adult rats were isolated and grown on permeable membranes for 5 to 10 days. Under these conditions, isolated epithelial cells grow to confluence, form tight junctions, and preferentially secrete SC into the apical medium. These studies identify epithelial cells as a key target cell in the uterus for the regulation of mucosal immunity by E₂, which we postulate will play an important role in studies to prevent and/or control the spread of sexually transmitted diseases.