Infection-induced Preterm Labor Research Articles

Inflammatory cytokines in a murine model of infection-induced preterm labor: cause or effect?

To characterize the expression of inflammatory cytokines in a murine model of preterm delivery induced by heat-killed bacteria. The right uterine horns of female CD-1 mice on day 14.5 of 19-20 days of gestation were inoculated with either sterile media or killed Escherichia coli bacteria (10(5)-10(10) organisms per mouse). The incidence of preterm delivery was recorded. The concentrations of cytokines (interleukin [IL-] 1 alpha, IL-1 beta, IL-1 receptor antagonist [IL-1ra], IL-6, and tumor necrosis factor alpha [TNF alpha]) within maternal and fetal tissue homogenates were determined by enzyme-linked immunosorbent assay at various times after inoculation. Killed E. coli induced preterm delivery in a dose-dependent fashion. Inoculation with 10(10) bacteria (sufficient to cause delivery in all mice) produced increases in IL-1 alpha, IL-1 beta, IL-6, and TNF alpha within uteri and fetal membranes, but not within placentas, fetal bodies, and maternal serum. Maximum mean uterine levels of IL-1 and IL-6 exceeded those of fetal tissues (membranes, placentas, and fetal bodies) by greater than 15-fold. Maximal uterine IL-1 and TNF alpha levels following inoculation with 10(10) bacteria exceeded those that followed inoculation with 10(7) bacteria (below the threshold for delivery) by 2.5- to 5-fold. The anti-inflammatory cytokine IL-1ra was expressed in higher concentrations in fetal than in maternal tissues and was unaltered by the bacterial inoculum. E. coli induce labor in mice even in the absence of bacterial viability. Although IL-1 and TNF alpha were upregulated by bacterial inocula causing delivery, peak levels were only 2.5- to 5-fold higher than those that occurred with inocula below the threshold for delivery (1000-fold fewer bacteria). Whether IL-1 and TNF alpha mediate labor during in vivo infection, or whether the upregulation of these cytokines merely represents an epiphenomenon accompanying infection, remains unknown.

Inflammatory Cytokines in a Murine Model of Infection-Induced Preterm Labor: Cause or Effect?

Inflammatory Cytokines in a Murine Model of Infection-Induced Preterm Labor: Cause or Effect?

Basal and interleukin-1beta-stimulated prostaglandin production from cultured human myometrial cells: differential regulation by corticotropin-releasing hormone.

During pregnancy, placental CRH acts on human myometrium via specific receptors and might play a role in the regulation of myometrial contractility and hence human parturition. The myometrium is the site of production and target of several PGs, which can be activated by cytokines, especially during infection-induced preterm labor. We established primary human myometrial cell cultures that express functional CRH receptors (CRH-R1alpha, -R1beta, -Rlc, and -R2beta) to investigate the possible regulation of PG production by CRH. We studied the effect of CRH on the two major myometrial PGs, PGE2 and 6-keto PGF1alpha. Human CRH was able to partially inhibit basal, interleukin-1beta-stimulated, and oxytocin-stimulated PGE2 production (56 +/- 11%, 45 +/- 8%, and 58 +/- 6% inhibition, respectively). This effect was blocked by a specific CRH receptor antagonist in a concentration-dependent manner. Furthermore, CRH had no effect on 6-keto PGF1alpha production, indicating that the CRH inhibitory action does not involve suppression of cyclooxygenase, the enzyme responsible for the production of both PGE2 and 6-keto-PGF1alpha. These data further support the view that during pregnancy, CRH may promote myometrial quiescence and might play an important role in the regulation of human labor.

Modulation of the prostaglandin e receptor: a possible mechanism for infection-induced preterm labor

Objective: To evaluate the modulatory effects of interleukin (IL)-1β and prostaglandin (PG)E 2 on the PGE 2 receptor subtype EP1 in amnion cell cultures. Methods: Amnion cell cultures were incubated in increasing concentrations of (IL)-1β or PGE 2. Cultures were also incubated in high concentrations of IL-1β and PGE 2 in combination. Changes in EP1 receptor levels were evaluated by western and northern blot analysis. Culture fluid PGE 2 levels were measured by enzyme-linked immunosorbent assay. Results: EP1 receptor protein levels decreased with increasing levels of PGE 2 ( r = −0.82, P < .05). EP1 receptor protein ( r = 0.95, P < .05), EP1 mRNA ( r = 0.95, P < .01), and culture fluid PGE 2 levels ( P < .01) were all increased after IL-1β administration. EP1 receptor levels also increased approximately fourfold in response to IL-1β incubation even in the presence of high agonist (PGE 2) concentrations ( P < .01). Conclusion: The results of this study show that IL-1β might be involved in infection-induced preterm labor by interfering with the normal regulation of EP1 receptor levels and with the promotion of increased PGE 2 production in amnion tissue.

Tumor necrosis factor-alpha upregulates the prostaglandin E2 EP1 receptor subtype and the cyclooxygenase-2 isoform in cultured amnion WISH cells.

Recent studies have demonstrated a strong correlation between infection and preterm labor. Preterm delivery is also associated with high levels of cytokines and prostaglandins in amniotic fluid. The purpose of this study was to investigate the effect of tumor necrosis factor-alpha (TNF-alpha) on the levels of cyclooxygenase, prostaglandin E2 production (PGE2), and expression of the PGE2 receptor subtype EP1 in amnion WISH cell culture. Amnion WISH cell cultures were incubated in increasing concentrations of TNF-alpha (0-50 ng/ml). Changes in cyclooxygenase and EP1 receptor proteins were evaluated by Western blot analysis. Changes in EP1 mRNA were evaluated by Northern blot, and culture fluid concentrations of PGE2 were estimated by enzyme immunoassay (EIA). EP1 protein (p<0.01), EP1 mRNA (p<0.05), cyclooxygenase-2 (COX-2) protein (p<0.001), and PGE2 concentrations (p<0.01) all increased with increasing concentrations of TNF-alpha. Changes in COX-1 protein were not observed following TNF-alpha-incubation. The results suggest that TNF-alpha may play a role in infection-induced preterm labor by its pleiotropic ability to simultaneously stimulate COX-2 activity, PGE2 concentrations, and PGE2 EP1 receptor levels in human amnion.

Interleukin-1β and bacterial endotoxin change the metabolism of prostaglandins E 2 and F 2α in intact term fetal membranes

There is strong evidence that prostaglandins E 2 and F 2α (PGE 2 and PGF 2α) are involved in the initiation and maintenance of human parturition and that their production can be stimulated by a number of cytokines and in infection-induced preterm labour by bacterial endotoxin. This study used an intact fetal membrane disk model to investigate the regulation of PGE 2 and PGF 2α metabolism by interleukin-1 beta (IL-1β and bacterial endotoxin [lipopolysaccharide (LPS)]. Fetal membrane explants were incubated with IL-1β (0.1 or 1.0 ng/ml) or LPS (10 ng/ml) for 24 h. A mixture of 3H-prostaglandin (0.1 μCi) and unlabelled prostaglandin (1 μg) was then added at selected times after the addition of inflammatory mediators. The radiolabelled prostaglandins and their metabolites were then extracted from the culture medium and quantified by high-pressure liquid chromatography. Levels of prostaglandin metabolites were generally decreased following incubation with IL-1β or LPS, which is consistent with a decrease in the activity of 15-hydroxyprostaglandin dehydrogenase (PGDH). It is concluded that IL-1β and LPS moderately decrease the metabolism of prostaglandins, which may contribute to increasing the local levels of active prostaglandins induced by these stimuli.

Inflammatory cytokine (interleukins 1, 6, and 8 and tumor necrosis factor-α) release from cultured human fetal membranes in response to endotoxic lipopolysaccharide mirrors amniotic fluid concentrations

OBJECTIVE: This study was conducted to quantitate and compare the amount of cytokines released from human fetal membranes in response to treatment with bacterial lipopolysaccharide and to compare this with amniotic fluid levels. STUDY DESIGN: Amniochorionic membranes were collected from women undergoing elective repeat cesarean section and showing no signs of infection- or pregnancy-related complications. Membranes were maintained in an organ explant system and stimulated with bacterial lipopolysaccharide for 24 hours. Media samples were collected and stored at -20° C until cytokine levels were assayed by enzyme-linked immunosorbent assay. RESULTS: Enzyme-linked immunosorbent assay results demonstrated that lipopolysaccharide stimulated production of interleukins 1, 6, and 8 and tumor necrosis factor-α by the fetal membranes in comparison with the control cultures. A greater release of interleukin-6 and interleukin-8 compared with interleukin-1 and tumor necrosis factor-α was noticed. The relationships between cytokine concentrations observed in culture mirror those seen in amniotic fluid. CONCLUSION: Amniochorionic membranes can respond to an infectious process with increased secretion of interleukins 1, 6, and 8 and tumor necrosis factor-α. Cytokines produced from both amnion and chorion (interleukin-6 and interleukin-8) are released in greater quantities than those cytokines produced from chorion or amnion alone (interleukin-1 and tumor necrosis factor-α). These studies support a major role for amnion in infection-induced preterm labor. (Am J Obstet Gynecol 1996;174:1855-62.)

Reactivity of endothelial cell intermediate filaments with human and mouse monoclonal IgM antiphospholipid antibodies

s A.63 REACTIVITY OF ENDOTHELIAL CELL INTERMEDIATE FILAMENTS WITH HUMAN AND MOUSE MONOCLONAL IgM ANTIPHOSPHOLIPID ANTIBODIES. N.S. Rote, L. Lin, L. Shroyer, T.W. Lyden, and A.K. Ng, Departments of Microbiology & Immunology, and Obstetrics & Gynecology, Wright State University School of Medicine, Dayton, OH, USA. Antibodies against phospholipids (aPLs) have been associated with recurrent thrombosis, possibly through modulation of endothelial cell function. In the current study we investigate the antl-endothelial cell reactivity of IgM monoclonal aPLs and IgM aPL-positive sera from patients with the antiphospholipid antibody syndrome. Using immunofluorescence and acetone fixed human endothelial cells we investigated the reactivity of three monoclonal antibodies that differentiate between cardiolipin (CL) and phosphatidylserine (PS) (BA3B5C4 [CL+/PS+]; 3SB9b [CL-/PS+]; DIIA4 [CL+/PS-]). 3SB9b reacted strongly with cytoskeletal-like cytoplasmic components, whereas DIIA4 and BA3B5C4 were unreactive. The patients' sera also contained IgM that reacted with cytoskeletal-like structures in the same pattern. Blocking experiments were performed using antibodies against actin (microfilaments), tubulin (microtubules), and vimentin (intermediate filaments), proteins that are representative of the three primary endothelial cell cytoskeletal structures. Only anti-vimentin blocked the binding of aPLs, either monoclonal or human, western blots of NP40 extracts of endothelial cells were probed with aPLs. 3SBgb reacted strongly with bands at 112 and 51 kDa, BA3B5C4 reacted with a 50 kDa band, and DIIA4 was unreactive. Sera from four patients also reacted against the 112 kDa, and two also reacted against the 51 kDa band. These results suggest that PS-like epitopes, identified by both monoclonal and patient's antibodies, are associated with endothelial intermediate filaments, aPL-mediated pathophysiology may, therefore, require endothelial cell damage or activation as a necessary cofactor to expose PS to circulating aPLs. EXPRESSION OF IL-IB AND IL-6 PROTEIN AND mRNA IN AMNIOCHORIONIC MEMBRANE. N.S. Rote, R. Menon, K.F. Swan, T.W. Lyden, and S.J. Fortunato, Departments of Microbiology & Immunology, and obstetrics & Gynecology, Wright State University School of Medicine, Dayton, OH, USA. A proposed mechanism for preterm labor is that intraamniotic infections induce cytokines that increase prostaglandin production. The role of fetal membranes in this mechanism has not been determined. We investigated whether cells of the amniochorionic membranes produce cytokines and whether that production is increased with exposure to endotoxin. Membranes collected by elective caesarian section from women without infection or pregnancy-related complications were washed in sterile buffer, cut into 5 mm diameter pieces, and maintained for 72 hr in media (DMEM:Nutrient mixture FI2 Ham) with antibiotics. IL-IB and IL-6 gene expression was measured with or without exposure to endotoxin (i00 pg/ml to i000 ng/ml) by PCR and in situ hybridization using short (20-35 mer) primers/probes for each cytokine and the sense strand as a negative control and by immunocytochemistry using immunoperoxidase labelling. Probes/primers for GAPDH were used as positive controls. Both chorion and amnion cells normally produced IL1B message or protein, or both, but not IL-6. Endotoxin (optimum at i0 ng/ml) induced IL-6 message and protein in both chorion and amnion. This study suggests that fetal membranes may be direct sources of IL-I and IL-6 in situ and contribute to infection-induced preterm labor.

Inflammatory mediators regulate interleukin-8 production by cultured gestational tissues: evidence for a cytokine network at the chorio-decidual interface.

The physiology of parturition, and the pathophysiology of preterm labor, is incompletely understood. Infection of gestational tissues may account for a significant proportion of women who experience preterm labor. Interleukin-8 (IL-8), or neutrophil-activating protein (NAP-1), is a potent chemotactic and activating factor for neutrophils and has been implicated in the pathogenesis of inflammatory injury to skin and lung, but has yet to be described in gestational tissues. Cultured chorion and decidual cells obtained from normal human pregnancies were used to evaluate whether these tissues produce IL-8 basally and in response to inflammatory cytokines. As measured by specific IL-8 RIA, chorion and decidual cells produce IL-8 constitutively and in response to IL-1 beta and tumor necrosis factor. Cotreatment of chorion cells or decidual cells with IL-1 beta and actinomycin D or cycloheximide abrogated IL-8 production. Northern blot analysis confirmed that IL-1 beta stimulation of chorion and decidual cells resulted in increased IL-8 messenger RNA expression. Our data support the concept that a complex cytokine network between maternal and fetal gestational tissues exists, and that activation of inflammatory cytokine production in these tissues, including IL-8, likely contributes to the pathophysiology of infection-induced preterm labor.

A murine model of preterm labor: inflammatory mediators regulate the production of prostaglandin E2 and interleukin-6 by murine decidua.

Inflammatory processes may be important in the initiation and propagation of uterine contractions and preterm labor in human pregnancies. Recently, a murine model of preterm labor has been described. The purpose of our study was to determine whether murine decidua responds to inflammatory mediators, such as lipopolysaccharide (LPS) and the inflammatory cytokines interleukin (IL)-1 beta and tumor necrosis factor (TNF). Allogeneic pregnant mice (C3H/Hen females mated with C57/B6 males) were killed at 12-14 days of pregnancy, decidual tissue was isolated, and explants were placed on the polycarbonate membrane of Costar Transwell inserts. Initial validation studies of this explant system, including biochemical and histologic evaluations, indicated that the decidual tissue remained intact, viable, and responsive to IL-1 beta for at least 5 days in explant culture. Treatment of murine decidual explants with LPS, IL-1 beta, and TNF resulted in significant increases in the production of prostaglandin E2 (PGE2) and IL-6. Thus the regulation of PGE2 and IL-6 production from murine decidua by LPS and inflammatory cytokines is similar to findings previously reported for human decidua. Our findings are consistent with the view that the pathophysiology of infection-induced preterm labor in the mouse may be similar to that in human pregnancy and supports the continued development of murine models of preterm labor.

Cytokines and infection-induced preterm labour

Our present understanding of the physiologic role of prostaglandins during pregnancy and, in particular, the precise function of these agents in the initiation of human parturition, is limited. This uncertain posture persists in spite of evidence (reviewed in detail recently by Challis and Olson 1988) in favour of a fundamental role for prostaglandins in a host of physiological processes in reproductive biology, including the onset and progression of labour. All investigators agree that prostaglandin E2 (PGE2) and prostaglandin F2, (PGF2,) can act, pharmacologically, as uterotonins (to cause myometrial contraction) when administered to pregnant women at any stage of gestation; moreover, the ingestion of inhibitors of prostaglandin synthesis by pregnant women is associated with a delay in the onset of labour-whether spontaneous or induced. There is much evidence in support of the conclusion that there is an increase in the rate of production of prostaglandins during labour: namely, during labour, there are increases in the levels of PGE2, PGF2, and 13,14-dihydro-15-keto-PGF2, (PGFM) in amniotic fluid and increases in PGFM in maternal blood. Also, prostaglandins will effect cervical dilatation. However, it has not been proven that prostaglandins serve as the physiological uterotonins of parturition or that prostaglandins are obligatorily involved in parturition at all stages of pregnancy, i.e. term and preterm labour. The evidence that there is an increase in the levels of prostaglandins in the maternal or fetal compartment prior to the onset of labour is limited. The possibility that the increase in prostaglandins is a consequence of labour (and not an initiating event) has not been excluded. Importantly, based on the findings of studies conducted recently, the possibility exists that labour (and delivery) may occur in some preterm pregnancies in the absence of an increase in prostaglandin production of the same magnitude that occurs during normal labour at term (Cox et al. 1989). Thus, we suggest that it is timely to consider the role of prostaglandins in the parturitional process with a new view, i.e, one that may enable an expansion of our understanding of the biochemical events involved in the initiation of parturition at term and preterm. Nowadays, the statement that human pregnancy is characterized by profound alterations in a host of biochemical and physiologic processes that encompass most all organ systems may rightly be considered timeworn. Yet, recent analyses of this well-recognized premise have led to the concepts that among these pregnancy-altered processes are those that facilitate recognition of pregnancy and, thereby, adaptation to pregnancy and maintenance of pregnancy. Perhaps the most remarkable compliance of normal human pregnancy is the maintenance of uterine quiescence for 38 weeks. For more than 99% of gestation, normal pregnancy is characterized by remarkable uterine repose and equanimity -a state that is essential to the maintenance of pregnancy. This quiescent state is uncommon for this smooth

Cachectin/tumor necrosis factor-alpha formation in human decidua. Potential role of cytokines in infection-induced preterm labor.

This study was conducted as part of an investigation to evaluate the hypothesis that bacterial toxins (LPS or lipoteichoic acid), acting on macrophage-like uterine decidua to cause increased formation of cytokines, may be involved in the pathogenesis of infection-associated preterm labor. We found that cachectin/tumor necrosis factor-alpha (TNF-alpha) was synthesized and secreted into the culture medium by human decidual cells and explants in response to treatment with LPS. LPS treatment also caused an increase in PGF2 alpha production by decidual cells and explants. In amnion cells in monolayer culture, TNF-alpha stimulated PGE2 formation, and TNF-alpha was cytostatic (inhibited [3H]thymidine incorporation into DNA) but not cytolytic in amnion cells. TNF-alpha was not detectable (less than 0.34 ng/ml) in the amniotic fluid of normal pregnancies at midtrimester or at term before or after the onset of labor (n = 44); but TNF-alpha was present at concentrations between 2.8 and 22.3 ng/ml in amniotic fluids of 4 of 20 pregnancies with intact membranes complicated by preterm labor (less than 34 wk gestational age). LPS was present in 10 of the 20 amniotic fluids of preterm labor pregnancies, including all four in which TNF-alpha was present. Bacteria were identified in only one of the four LPS-positive, TNF-alpha-positive fluids. Cytokine formation in macrophage-like decidua may serve a fundamental role in the pathogenesis of preterm labor, including increased prostaglandin formation and premature rupture of the membranes.