The influence of interleukin-1beta on oxytocin signalling in primary cells of human decidua.

OBJECTIVE: Oxytocin (OT) and its corresponding receptor (OTR), synthesized within the pregnant uterus, play a key role in the process of (preterm) labor as part of a paracrine system that regulates symmetrical contractility. In the setting of intrauterine infection, a major cause of preterm labour and birth, decidua serves as a major source of cytokine production. The present study evaluates the time-dependent effect [0-24 h] of the inflammatory cytokine Interleukin-1beta (IL-1beta) treatment on OT signalling and OT stimulated prostaglandin release in primary cultures of human decidua. STUDY DESIGN: Primary cultures of human decidua (n=6) were treated with IL-1beta [5 ng/ml] for 0-24h and or indomethacin [100 microM]--an inhibitor of the prostaglandin synthesis--for 0-24 h or for 24 h. OT peptide expression, OTR binding, Inositol triphosphate production (IP(3)), and arachidonic acid (AA) as well as prostaglandin (PGE(2)) release were measured. RESULTS: IL-1beta transiently reduced cytoplasmic OT peptide at 4-6 h of IL-1beta incubation, while its secretion into the media was increased after 6 h of stimulation. The later was completely blocked by indomethacin. A decrease in OTR mRNA expression and a loss of OTR binding were detected after 8 h and 16 h of IL-1beta treatment, respectively. IL-1beta also decreased IP(3) production and AA release, but significantly enhanced OT mediated PGE(2) production. This effect was completely suppressed by the cyclooxygenase-2 (COX-2) inhibitor NS-398. CONCLUSION: Our data suggest, that IL-1beta indirectly increases OT secretion in primary cultures of human decidua in a time dependent fashion through the production of prostaglandins through COX-2 and that this increase in OT peptide may secondarily down-regulate the OTR and its signalling cascade. These findings might explain the poor effectiveness of oxytocin receptor antagonists as tocolytic agents in the setting of intrauterine infection.

Effect of IL-1beta and IL-6 on oxytocin secretion in human uterine smooth muscle cells.

PROBLEM: Infection-mediated preterm labor results in the production of inflammatory cytokines, including interleukin-1beta (IL-1beta) and interleukin-6 (IL-6). Oxytocin (OT) plays a key role in the process of labor. This study investigates the effect of IL-1beta and IL-6 on intra-and extracellular OT in human smooth muscle cells and evaluates IL-1beta induced changes in IL-6 production. METHOD OF STUDY: Primary cultures of human myometrium, obtained from term pregnant women (n = 7) were incubated with either IL-1beta or IL-6 for 0-24 hr. Intra- and extracellular OT peptide concentrations were measured by radioimmunoassay and IL-6 mRNA and protein were evaluated by reverse transcription polymerase chain reaction and enzyme-linked immunosorbent assay. RESULTS: Both, IL-1beta and IL-6 led to an increase in OT secretion, which was accompanied by a reduction of the intracellular OT peptide pool. IL-1beta significantly induced IL-6 mRNA expression and protein secretion, which did not further enhance IL-1beta induced OT secretion. CONCLUSIONS: The induction of OT secretion by proinflammatory cytokines in human myometrium in vitro, supports the concept of a thus regulated infection-triggered preterm labor process in vivo.

Interleukin-6 up-regulates the oxytocin receptor in cultured uterine smooth muscle cells.

PROBLEM: Intra-uterine infection results in the production of inflammatory cytokines, including interleukin-6 (IL-6). Increased oxytocin-receptor (OTR) concentrations are associated with the onset of preterm labor. We hypothesize that infection up-regulates OTR expression through IL-6-induced transcription factors. METHOD OF STUDY: Primary cultures of human myometrium were treated for various time periods or with different concentrations of IL-6 and OTR mRNA as well as OTR binding were measured by means of reverse transcription polymerase chain reaction and 125I-ornithine-vasotocin-binding assay. To study underlying mechanisms of OTR changes with IL-6 treated, cells were also incubated with genistein or H7 (tyrosine and serine phosphorylation inhibitors), respectively. RESULTS: OTR mRNA increased 2.5-fold after 4 hr of IL-6 treatment and OTR binding 1.4-fold after 8 hr of cytokine stimulation. The IL-6-induced increase in binding was blocked by genistein and H7. CONCLUSIONS: IL-6 up-regulates uterine OTR mRNA expression and binding capacity in cultured human myocytes most likely through tyrosine and serine phosphorylation pathways involving the nuclear factor STAT-3.

Oxytocin signaling in human myometrium is impaired by prolonged exposure to interleukin-1.

Intra-amniotic infection leads to preterm labor and is associated with the local release of inflammatory cytokines by fetal membranes, resulting in the production of uterotonic prostaglandins. Oxytocin, however, also plays a key role in the initiation of labor. Short-term exposure of myometrium to interleukin (IL)-1 enhances oxytocin signaling and contractility. With intrauterine infection, however, myometrium is exposed to inflammatory cytokines for prolonged periods. The present study was conducted to demonstrate that myometrial oxytocin signaling is significantly impaired following prolonged exposure to IL-1. Myometrial cells were treated with IL-1 for 24 h. Oxytocin-stimulated inositol trisphosphate (IP(3)) production was measured in tritiated myoinositol-loaded myometrial cells. Arachidonic acid (AA) release was measured in tritiated AA-loaded myometrial cells. Increases in intracellular calcium were measure with fluo-3. Prostaglandin (PG) F(2alpha) and 6-keto-PGF(1alpha) were measured by ELISA assay. Prolonged exposure of myometrial cells to IL-1 resulted in a significant reduction in oxytocin-mediated signaling as measured by IP(3) production and AA release, as well as a decrease in intracellular calcium. Prolonged exposure of myometrial cells to IL-1, however, resulted in enhanced PG release. Oxytocin may not contribute significantly to the labor-inducing action of IL-1 in the setting of preterm labor with prolonged infection.

Candida albicans chorioamnionitis associated with preterm labor and sudden intrauterine demise of one twin. A case report.

BACKGROUND: Although cervicovaginal Candida infections occur in 20-25% of pregnancies, the incidence of ascending infection in these cases is only 0.8%, and such infection rarely causes chorioamnionitis. CASE: Sudden intrauterine fetal demise (IUFD) of twin A occurred in a diabetic primigravida presenting with a twin pregnancy and preterm labor at 33 weeks of gestation. Placental pathology and autopsy of the stillborn twin revealed extensive chorioamnionitis and fetal sepsis in the presence of Candida albicans. Twin B was unaffected. CONCLUSION: In this case, C albicans chorioamnionitis seemed to be associated with sudden IUFD.

Interleukin-1 stimulates human uterine prostaglandin production through induction of cyclooxygenase-2 expression.

PROBLEM: Uterine infection occurs in as much as 20% of preterm labor and results in increased decidual cytokines. The objective of this study was to examine the effect of interleukin-1 (IL-1) and the cyclooxygenase-2 (COX-2) inhibitor, NS-398, on myometrial prostaglandin (PG) production and COX-2 expression. METHOD OF STUDY: Human uterine myocytes were stimulated with IL-1 (0-50 ng/mL) over 24 hr. PGE2, PGF2alpha, and 6-keto F1alpha were measured by enzyme-linked immunosorbent assay. Both COX-1 and COX-2 proteins and mRNA were measured by western and northern blot, respectively. RESULTS: IL-1 increased PG production beginning at 6 hr, COX-2 protein increased beginning at 4 hr and continued to increase at 24 hr. COX-2 mRNA increased at 2 hr and peaked at 4 hr. NS-398 blocked PG production but had no effect on COX-2 protein or mRNA. CONCLUSIONS: IL-1 increases PG production by myometrium by increased COX-2 expression. NS-398 completely blocks IL-1-induced PG production. With intrauterine infection, IL-1 may induce labor through the autocrine production of uterotonic PGs.

Interleukin-1 beta down-regulates the oxytocin receptor in cultured uterine smooth muscle cells.

PROBLEM: Intrauterine infection accounts for 20% of preterm labor and results in the production of decidual inflammatory cytokines, including interleukin-1 (IL-1). The oxytocin receptor plays a key role in the onset of preterm labor. Cytokines likely regulate oxytocin receptor expression through several cytokine-induced DNA-binding proteins. METHOD OF STUDY: The objective of this study was to evaluate the effect of the IL-1 alone on oxytocin receptor number as measured by radioligand binding and immunocytochemistry, and oxytocin receptor mRNA as measured by reverse transcriptase-polymerase chain reaction (RT-PCR) in cultured uterine myocytes. RESULTS: Unexpectedly, IL-1 treatment decreased oxytocin receptor number from 111,067 to 23,941 receptors/cell. Loss of oxytocin receptor binding began after 8 hr of IL-1 treatment and was reversible after IL-1 removal. Immunocytochemistry confirmed a loss of cellular oxytocin receptors. Oxytocin receptor mRNA decreased beginning after 2 hr of IL-1 treatment. CONCLUSIONS: IL-1 down-regulates the uterine oxytocin receptor in a time- and dose-dependent fashion.

Mitogenic effect of basic fibroblast growth factor and estradiol on cultured human myometrial and leiomyoma cells.

OBJECTIVE: We compared the mitogenic effect of basic fibroblast growth factor with and without estradiol on myometrial and leiomyometrial cells. STUDY DESIGN: The mitogenic effect of basic fibroblast growth factor on myometrial cells was measured by thymidine incorporation and cell count. The mitogenic effect of basic fibroblast growth factor with and without estradiol as measured by thymidine incorporation was compared between myometrial and leiomyometrial cells. RESULTS: Both human myometrial and leiomyometrial cells showed significant (p = 0.004 and p = 0.001, respectively), dose-dependent incorporation of thymidine in response to basic fibroblast growth factor. Leiomyometrial cells showed significantly (p = 0.04) less thymidine incorporation compared with matched normal myometrial cells. The addition of estradiol with basic fibroblast growth factor did not result in a further increase in thymidine incorporation. CONCLUSIONS: Both myometrial and leiomyometrial cells respond to basic fibroblast growth factor with increased thymidine incorporation; however leiomyometrial cells are less responsive than are matched normal myometrial cells. The addition of estradiol is not synergistic with basic fibroblast growth factor.

Ritodrine increases leukotriene B4 concentrations in pregnant sheep.

OBJECTIVES: Our goals were (1) to determine if beta-adrenergic receptor stimulation leads to 5-lipoxygenase metabolism of arachidonic acid and (2) to determine if inhibition of prostaglandin synthase alters 5-lipoxygenase metabolism of arachidonic acid during beta-adrenergic stimulation. STUDY DESIGN: We infused saline solution, ritodrine (4 micrograms/kg/min), and a combination of ritodrine (4 micrograms/kg/min) and ketorolac (1.2 microgram/kg/min) into chronically catheterized pregnant sheep (gestational ages 110 to 120 days, term 147 days). With a radioimmunoassay we measured concentrations of leukotriene B4, a 5-lipoxygenase metabolite of arachidonic acid, in uterine venous and arterial plasma at 0, 2, and 4 hours during the infusion. RESULTS: Both uterine venous and arterial leukotriene B4 were increased during ritodrine infusion (mean uterine venous increase at 2 hours 218%, p < 0.05; mean arterial increase at 2 hours 280%, p < 0.05). Concentrations during combined infusion of ritodrine and ketorolac increased significantly but were not different than concentrations observed during ritodrine infusion. CONCLUSION: Infusion of the beta-agonist ritodrine leads to 5-lipoxygenase metabolism of arachidonic acid and increased concentrations of leukotriene B4. The increased concentration in both uterine venous and arterial plasma suggests a systemic source of leukotriene B4 production. Concurrent inhibition of prostaglandin synthase during ritodrine infusion does not change 5-lipoxygenase metabolism in this model.

The prostaglandin synthesis inhibitor ketorolac blocks ritodrine-stimulated production of prostaglandin F2 alpha in pregnant sheep.

OBJECTIVE: To determine whether ritodrine-stimulated production of prostaglandin (PG) F2 alpha by pregnant uterine tissues can be blocked by the concurrent administration of ketorolac, a PG synthesis inhibitor. METHODS: We infused saline, ritodrine, ketorolac, or a combination of ritodrine and ketorolac into chronically catheterized pregnant sheep. Concentrations of PGF2 alpha in uterine venous plasma were measured by radioimmunoassay at 0, 1, 2, 3, 4, and 24 hours during the infusions. RESULTS: Ritodrine significantly increased uterine venous PGF2 alpha; mean percent increases at 4 hours were 330% and 380%, and at 24 hours 370%, compared with controls. During concurrent ritodrine and ketorolac infusion, there was no increase in uterine venous PGF2 alpha at any time. CONCLUSIONS: Ketorolac completely blocks ritodrine-stimulated production of PGF2 alpha in pregnant uterine tissues. We conclude that ritodrine stimulates PG production through mobilization of arachidonic acid, and this can be effectively blocked with a PG synthesis inhibitor. This finding may have important clinical applications in the treatment of preterm labor.

Use of the fetal chest in estimating fetal weight.

OBJECTIVE: This study was designed to develop formulas using the chest circumference instead of the abdominal circumference for estimating fetal weight. STUDY DESIGN: Ultrasonographic measurements of the chest circumference, biparietal diameter, abdominal circumference, humeral length, and femoral length were obtained in 75 term fetuses of uncomplicated pregnancies within 24 hours of delivery. Three equations for fetal weight estimation that used the chest circumference, instead of the abdominal circumference, in combination with the biparietal diameter or the humeral length were developed by regression analysis. RESULTS: The average mean errors of fetal weight estimation for these equations vary from 7.1% to 7.6%. CONCLUSIONS: These equations may be used in predicting the birth weight when the fetal abdomen is altered by certain fetal abnormalities.