| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
|
|
|
|||||||
|
|||||||||
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Pregnancy |
Fetal and Neonatal Research Group,3 Department of Physiology, Monash University, Clayton, Victoria 3800, Australia
Centre for Biotechnology,4 University of Melbourne, Melbourne, Victoria 3052, Australia
Institute of Reproduction and Development,5 Monash Medical Centre, Clayton, Victoria 3168, Australia
 |
ABSTRACT |
|---|
 TOP ABSTRACT INTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
|---|
cortisol, cytokines, parturition, placenta, uterus
|
INTRODUCTION |
|---|
|
TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
|---|
It is believed that infections within the uterus can occur between the choriodecidual space, within the fetal membranes (the amnion and chorion), in the placenta or amniotic fluid, or within the umbilical cord or fetus [1]. It has been proposed that organisms first ascend from the vagina into the choriodecidual space. In women, the organisms are then believed to cross the intact chorioamniotic membranes into the amniotic fluid [5]. Recent evidence suggests that intrauterine infection may occur quite early in pregnancy and remain undetected for months [6], thus exposing the fetus to the effects of endotoxin for a considerable period. Identifying women with intrauterine infections therefore, is a major obstetric challenge, because intrauterine infection is often asymptomatic [7].
A number of studies have used the administration of bacteria or bacterial products to pregnant animals as a model of human premature labor [8–11]. In a nonhuman primate model, intra-amniotic inoculation with group B Streptococcus increases amniotic fluid cytokines, particularly interleukin (IL)-6, in parallel with increases in amniotic fluid prostaglandins [12]. Both these changes precede increases in uterine contractility. The elevation in amniotic fluid cytokines observed after experimental infection are consistent with those reported in women, in whom IL-6, IL-1, and tumor necrosis factor (TNF
) have been found in elevated concentrations in the amniotic fluid of women with intra-amniotic infection and premature labor [13, 14].
Lipopolysaccharide (LPS), a pyrogenic component of Gram-negative bacteria cell walls, induces a downstream cascade of inflammatory responses, stimulating macrophages to produce large amounts of cytokines such as TNF-, IL-1, and IL-6 [15, 16], similar to that observed with infection, but without causing invasion or multiplication of microorganisms within the body. Most previous studies have focused on endotoxin exposure within the intra-amniotic cavity, where the amniotic fluid is believed to provide a conduit through which the infection, or the infective agents, then gain access to the fetus, initiating responses that may either harm or protect the fetus [17]. Work by Kallapur et al. [18] showed that intra-amniotic LPS administration in sheep resulted in chorioamnionitis, an elevation in proinflammatory cytokine expression in the amnion/chorion, and lung inflammation, similar to that observed in premature infants with associated chorioamnionitis. Thus, LPS was considered as an ideal experimental tool for investigating inflammatory responses during pregnancy, but the effect on uterine activity and fetal endocrine parameters were not measured in this study. Many other studies have used LPS as an agent to induce premature labor, which have also been accompanied by fetal death. A study by Fidel et al. [19] showed that i.p. LPS administration (50 µg) in mice at 70% gestation results in an 87% incidence of premature labor, yet these authors did not describe fetal or neonatal outcome. Similarly, Kaga et al. [10] demonstrated a 100% incidence of premature delivery in mice treated i.p. with LPS (50 µg, twice at 3-h intervals), however, all fetuses that delivered prematurely were dead in utero. Fetal death was also observed in a study by Schlafer and colleagues [11], in which LPS administration (1 µg/kg) to the ewe resulted in fetal death and the onset of premature labor. Collectively, these data support the view that an inflammatory challenge induced by LPS can lead to higher concentrations of proinflammatory cytokines and, subsequently, premature labor. However, because most of these studies have been undertaken in species in which it is not possible to monitor the fetal effects of chronic endotoxin exposure, little is known of the fetal responses during this inflammatory process or of the effects of these responses on fetal well-being and neonatal outcome. Often, studies administering LPS to the amniotic fluid have used doses in excess of those that would be severely toxic if administered to the mother [20] or the fetus [18]. We hypothesize that the inflammatory effects of endotoxins that are produced as a result of ascending intrauterine infections during pregnancy are suppressed, in comparison with those observed in response to a vascular insult, through the barrier provided by the amniotic fluid.
Our aim, therefore, was to use an ovine model that allows continual fetal monitoring to investigate the fetal effects of an inflammatory challenge during pregnancy using maternal systemic, intra-anmiotic, and extra-amniotic administration of LPS, thus mimicking the processes believed to be involved in premature labor associated with infection in women. A greater understanding of the consequences of uterine infections during pregnancy and the mechanisms by which the fetus and mother respond to an inflammatory challenge is crucial to developing better approaches for the detection and treatment of premature labor. This may also provide an indication as to when it may be appropriate to delay cases of premature labor that may be associated with infection.
|
MATERIALS AND METHODS |
|---|
|
TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
|---|
Animals and Surgery
Border Leicester-Merino crossbred ewes were used in this study. Animals were housed indoors in individual metabolism cages, fed between 0900 h and 1200 h daily in a 12L:12D cycle. Water was provided ad libitum. Surgery was performed on pregnant ewes at 123 ± 1 day gestation (GA) to insert vascular catheters in the maternal and fetal carotid artery (CA) and jugular vein (JV). An intra-amniotic fluid catheter (basket) with a perforated end to avoid surrounding membranes from occluding the terminal ports, was sutured to the nape of the fetal neck to allow sampling directly from the amniotic fluid. A separate intra- or extra-amniotic catheter was inserted for LPS administration. The extra-amniotic catheter was placed between the chorioamnion [21] and the endometrium and secured to the uterine wall. Uterine electromyographic (EMG) leads were sutured to the external surface of the myometrium for continual monitoring of uterine activity before and during LPS treatment.
Experimental Design
Lipopolysaccharide derived from Escherichia coli (serotype 0127:B8; Sigma Chemical Co., St Louis, MO) was used as the inflammatory agent and was diluted in sterile saline (David Bull Laboratories, Melbourne, Victoria, Australia). Control values were taken from each animal at -12, -0.5, and -0.25 h before LPS treatment, allowing each animal to be used as its own control. Results for -12, -0.5, and -0.25 h before LPS treatment were thus combined to give pretreatment values for each animal.
Maternal systemic LPS administration Ewes were administered LPS as a bolus injection via the maternal JV catheter (2 µg/kg of maternal body weight in 5 ml of saline, n = 5) at 130 days GA. Ewes and their fetuses were monitored before, during, and up to 48 h after maternal LPS treatment.
Intra-amniotic LPS administration
At 125 ± 1 day GA, ewes were administered LPS as a bolus injection via the intra-amniotic catheter attached to the fetus (400 µg/kg of maternal body weight, a total of 
20 mg, in 10 ml of saline; n = 4). Ewes and their fetuses were monitored before, during, and up to 48 h after intra-amniotic LPS treatment.
Extra-amniotic LPS administration At 132 ± 2 days GA, ewes received a constant infusion of LPS via the extra-amniotic catheter at three doses (0.1, 1.0, and 10 µg/kg of maternal body weight per day, in saline 0.5 ml/h; n = 4). Ewes and their fetuses were monitored before and during the extra-amniotic infusion for 72 h.
Sample Collection
Blood samples were obtained from the fetal CA, maternal CA, and amniotic fluid in all ewes before, during, and after the administration of LPS at regular intervals. At the completion of each experiment, ewes and their fetuses were killed by barbiturate overdose (Lethabarb; Arnolds of Reading Pty. Ltd., Boronia, Victoria, Australia), administered via the maternal JV catheter.
Maternal and fetal blood respiratory gases (PaO2 mm Hg, PaCO2 mm Hg, percent O2 saturation), hemoglobin, and pH were measured using an ABL5 blood gas analyzer and OSM2 hemoximeter (Radiometer, Copenhagen, Denmark). Blood samples were collected into EDTA tubes containing indomethacin (10 µM/ml blood). Aliquots of plasma for PGFM (13, 14-dihydro-15-keto PGF2, the stable metabolite of PGF2), cortisol, and IL-6 assays were stored at -20°C until further analysis. Plasma for PGE2 assay was diluted 1:1 with 0.12 M methyloxyamine hydrochloride (Sigma) in sodium acetate buffer (1 M, pH 5.6) containing 10% ethanol, and left overnight at room temperature then stored at -20°C.
PGE2 and PGFM Radioimmunoassay
PGE2 and PGFM concentrations in fetal and maternal plasma and amniotic fluid were measured by RIA as described previously [22, 23]. Both [5,6,8,11,12,14,15-3H(n)]PGE2 (DuPont-NEN Products, Boston, MA) and standard PGE2 (0.5 mg; Sigma) were methyloximated as previously described [23]. The PGE2 and PGFM antisera were raised in sheep and donated by Dr. R.I. Cox (Commonwealth Scientific and Industrial Research Organisation, Prospect, NSW, Australia). The intraassay and interassay coefficients of variation of the PGE2 RIA were 18% and 21%, respectively. The mean sensitivity of the assay was 0.26 ± 0.08 nmol/L. The intraassay and interassay coefficients of variation of the PGFM RIA were 4% and 15%, respectively, with a mean sensitivity of 0.30 ± 0.08 nmol/L.
Cortisol Radioimmunoassay
Cortisol was measured in fetal and maternal sheep plasma after extraction with dichloromethane as previously described [24]. Antiserum #3368 raised in sheep was kindly supplied by Dr. R.I. Cox. The intraassay and interassay coefficients of variation were 10% and 15%, respectively. The mean sensitivity of the assay was 1.75 ± 0.58 nmol/L.
IL-6 ELISA
IL-6 concentrations in fetal and maternal plasma and amniotic fluid samples were measure by ELISA, as previously described [25]. The standard was ovine recombinant IL-6 [25] and the intraassay and interassay coefficients of variation were 7% and 24%, respectively. The mean sensitivity of the assay was 0.39 ± 0.17 ng/ml.
Uterine Electromyographic Activity
Uterine EMG activity was recorded continuously before, during, and after administration of LPS using an ML135 dual bioamplifier connected to an ML795 PowerLab/16sp data recording system (ADInstruments Pty. Ltd., Castle Hill, NSW, Australia). Uterine EMG activity was analyzed using Chart v4.02 (ADInstruments). For each animal, the number of discrete bursts of uterine activity occurring within each 2-h period were measured as defined previously [26].
Statistical Analysis
All data are presented as the mean ± SEM. Data were first tested for homogeneity of variance, with heterogeneous data rendered homogeneous by square root or logarithmic transformation. All results were grouped into 2-, 6-, or 12-h time periods after LPS treatment, and the data were averaged for collective overall analysis. Basal values taken at -12, -0.5, and -0.25 h before LPS treatment were averaged for analysis and used as pretreatment values. Differences were identified by comparison with mean basal values before LPS treatment. Differences in uterine activity were identified by comparison with basal uterine activity 12 h before LPS treatment. Statistical analysis was performed by identifying differences between means using a one-way repeated measures ANOVA, time being the variable factor (SPSS Inc., Chicago, IL). Least significant difference tests were used to identify significant differences between pairs of mean values. Significance is reported at the 5% level (P < 0.05).
|
RESULTS |
|---|
|
TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
|---|
Premature delivery did not occur in any of the animals studied, despite uterine activity significantly increasing after LPS administration into the maternal circulation and directly into the amniotic fluid. Fetal or maternal death did not result with any of the doses of LPS used or the experimental conditions employed in this study.
Fetal blood gases were monitored at regular intervals before and after the administration of LPS into the maternal circulation or directly into the amniotic fluid, and are presented in Table 1. Fetal O2 saturation was significantly decreased, reaching a minimum value of 35.75% ± 4.31%, at 6–12 h after maternal LPS administration. Correspondingly, fetal PaCO2 was slightly elevated and fetal pH was significantly decreased. Fetal PaO2 also decreased at this time, although it was not significantly different from pretreatment PaO2 values (P = 0.064). Direct administration of LPS into the amniotic fluid resulted in a small but significant decrease in fetal O2 saturation 6–12 h after treatment (Table 1). No significant changes were observed in the maternal O2 saturation after systemic LPS administration (data not shown), however, there was a significant reduction in maternal PaCO2 levels from 30.56 ± 0.66 mm Hg to 25.85 ± 0.94 mm Hg, 2–4 h after maternal systemic LPS administration. PaCO2 remained significantly reduced for the subsequent 8 h. A significant increase in maternal pH was also observed 2–4 h after maternal systemic LPS administration. These ewes displayed an increase in body temperature (39.26 ± 0.04°C to 41.05 ± 0.25°C) 3.5 h after maternal LPS treatment. These changes were also associated with mild respiratory distress (tachypnea, nasal flaring, and grunting) in all animals, lasting for approximately 8 h.
|
A small but significant reduction in O2 saturation was observed in fetuses of ewes that received LPS into the extra-amniotic compartment at the dose of 10 µg/kg per day. There were no significant changes in the other fetal blood gases measured before, during, or after the administration of LPS into the extra-amniotic compartment. There were also no significant changes in maternal blood gas parameters measured after intra-amniotic LPS administration or into the extra-amniotic compartment (data not shown).
Uterine EMG Activity
Uterine EMG activity in animals treated with LPS via the maternal circulation significantly increased within the first 2 h of treatment (Fig. 1A). The elevated uterine EMG activity levels were consistently maintained for more than 30 h, before returning to basal levels by 48 h after treatment. There was a small but significant increase in uterine EMG activity commencing 2 h after intra-amniotic LPS administration (Fig. 1B). The extra-amniotic infusion of LPS had no observable effect on uterine EMG activity at any of the doses of LPS administered, as shown in Figure 1C. Uterine activity in these animals remained at basal levels (<5 bursts/2 h) for the entire study period.
|
Prostaglandin Concentrations
PGE2 and PGFM concentrations in the fetal and maternal circulations and amniotic fluid before and after LPS administration into the maternal circulation or directly into the amniotic fluid are presented in Figure 2. There was a significant increase in fetal plasma PGE2 concentrations observed by 4–6 h after maternal systemic LPS administration, and this increase was maintained for up to 48 h after maternal systemic LPS treatment (Fig. 2A). No changes in maternal plasma PGE2 or amniotic fluid PGE2 concentrations were observed during the entire experimental period (Fig. 2A).
|
There was a significant increase in PGFM concentrations in the fetal circulation within 4–6 h after maternal systemic LPS administration with concentrations continuing to rise until 12–24 h after treatment (Fig. 2B). There was a small, but significant increase in PGFM concentrations in the maternal circulation 2–4 h after maternal systemic LPS administration, with a further increase at 12–24 h after treatment, (Fig. 2B). There was also a significant increase in amniotic fluid PGFM concentrations observed from 6–12 h after maternal systemic LPS treatment, which rose further over the following 48 h.
Plasma PGE2 concentrations in the maternal and fetal circulations remained unchanged after LPS administration directly into the amniotic fluid (Fig. 2C). However, the amniotic fluid PGE2 concentrations increased within 2–4 h after intra-amniotic LPS administration, and continued to rise over the subsequent 48 h (Fig. 2C). A small increase in amniotic fluid PGFM concentrations was also noted 24–48 h after intra-amniotic LPS administration (Fig. 2D).
No significant changes were observed in fetal and maternal plasma or amniotic fluid PGE2 concentrations during the infusion of LPS (0.1, 1.0, or 10 µg/kg per day) into the extra-amniotic compartment (data not shown). Small increases in PGFM concentrations were observed in the fetal plasma samples during the infusion of 1.0 µg/kg per day of LPS (1.15 ± 0.48 nmol/L, P < 0.05, n = 4) and 10 µg/kg per day (2.12 ± 0.50 nmol/L, P < 0.05, n = 4) into the extra-amniotic compartment compared with that of basal concentrations (0.71 ± 0.50 nmol/L). PGFM concentrations in the maternal circulation and amniotic fluid remained unchanged during extra-amniotic LPS infusion.
Cortisol Concentrations
Maternal systemic LPS administration resulted in a dramatic increase in cortisol concentrations in the maternal circulation within 2 h and peaked at 4 h after maternal systemic LPS administration (Fig. 3A). Importantly, there was also a significant rise in cortisol concentrations in the fetal circulation within 2 h after maternal systemic LPS administration and remained elevated over the subsequent 24 h.
|
There was a significant rise in cortisol concentrations in the fetal circulation 4–6 h after intra-amniotic LPS administration, (Fig. 3B), however, there were no significant changes in maternal cortisol concentrations. Also, there were no changes in fetal and maternal plasma cortisol concentrations during the continuous infusion of LPS (0.1, 1.0, or 10 µg/kg per day) to the extra-amniotic compartment (data not shown).
IL-6 Concentrations
Interleukin-6 concentrations increased dramatically in the maternal circulation by 3 h after systemic LPS treatment and remained significantly above basal IL-6 concentrations for up to 24 h after maternal systemic LPS treatment (Fig. 4A). No significant differences were found in IL-6 concentrations in the fetal, maternal, or amniotic fluid after intra-amniotic LPS administration (Fig. 4B) or during LPS infusions into the extra-amniotic compartment (data not shown).
|
|
DISCUSSION |
|---|
|
TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
|---|
The principal finding of this study was that exposure of the maternal side of the placenta to LPS resulted in a marked rise in fetal plasma PGE2 and cortisol concentrations. Importantly, these increases were of a sustained nature, persisting for up to 48 h after maternal LPS treatment. There were also significant alterations in the fetal blood gases, consistent with previous observations [11]. Fetuses became hypoxemic, hypercapnic, and acidemic at 6–12 h after maternal systemic LPS treatment. Ewes treated with systemic administration of LPS displayed an increase in body temperature 3.5 h after LPS administration, a significant decrease in maternal arterial PaO2, and, consistent with the observed maternal hyperventilation, ewes were mildly hypocapnic and alkalotic. These changes were also associated with clinical signs of mild respiratory distress, which lasted for approximately 8 h. These effects are in agreement with previous observations of the pulmonary effects after systemic LPS administration in the ewe [28]. Thus, whereas ewes showed the usual signs of a mild inflammatory response, the present findings indicate that this maternal response may have marked effects on fetal prostaglandin and cortisol concentrations as well as fetal well-being.
The rise in maternal cortisol and IL-6 concentrations are consistent with the induction of a maternal immune response following LPS administration [11, 19]. The mechanisms that initiate these fetal changes after maternal systemic LPS exposure remain unclear. Maternal LPS administration may stimulate the endometrium to secrete cytokines that then act on the placenta, increasing prostaglandin secretion. Alternatively, it is possible that the fetus is affected by the placental passage of monocyte-derived cytokines produced in the maternal compartment [19]. These cytokines would then be expected to enhance prostaglandin production by stimulating prostaglandin synthase type-2 (PGHS-2) within the placental trophoblast [29]. The finding that both PGE2 and PGFM were elevated in the fetus, whereas in the maternal circulation and amniotic fluid only PGFM rose, suggests that stimulation of PGHS-2 within fetal fibroblasts or cells on the fetal side of the placenta may lead to the secretion of predominantly PGE2 [30]. The moderate rise in maternal PGFM in the plasma likely results from higher production within the uterine tissue. This rise would account for the increase in uterine activity that was observed following maternal LPS treatment. In addition, maternal LPS administration could also interfere with placental progesterone secretion causing prostaglandins to increase in the placenta and particularly in the endometrium. The finding that the animals did not deliver following maternal systemic or intra-amniotic LPS administration, despite increases in uterine activity, is consistent with the potent suppressive influence of progesterone produced by the placenta in this species, and which may not have been reduced sufficiently to cause premature labor. In contrast, intrauterine infection has been associated with a fall in luteal progesterone production and plasma concentrations before the onset of parturition in rabbits [9], and results in the onset of premature delivery in many other species [10, 12, 27].
The acute administration of E. coli endotoxin (0.2–0.5 mg/kg) to pregnant sheep has been shown to cause a significant drop in maternal arterial blood pressure as well as utero-placental blood flow, and therefore, a reduction in oxygen delivery to the fetus [31]. The induction of a mild maternal immune response following systemic LPS administration may be sufficient to cause a reduction in maternal arterial blood pressure and utero-placental blood flow, and thus, may account for the fetal blood gas responses observed in the study. The increase in fetal cortisol concentrations may have resulted from factors induced by LPS and released into the fetal circulation by the placenta that may stimulate the fetal adrenals. The latter possibility is supported by the finding that maternal cortisol concentrations were not affected by injection of LPS into the amniotic fluid, but fetal concentrations were significantly increased by this treatment. Indeed, intrauterine infections have been shown to increase fetal hypothalamic and placental production of corticotropin-releasing hormone, causing an increase in fetal corticotropin secretion, which in turn increases fetal adrenal production of cortisol [3]. In addition, previous studies have shown a correlation between fetal oxygen saturation and hypothalamic-pituitary-adrenal activation, in which fetal cortisol concentrations increase when the decrease in oxygenation is close to that associated with the onset of metabolic acidosis [32]. The reduction in fetal oxygenation and pH observed in this study were in the ranges previously reported to increase cortisol concentrations [32], thus, the increase in fetal cortisol concentrations observed in this study may have resulted from the fetal hypoxia associated with decreased utero-placental blood flow caused by maternal systemic LPS treatment. Moreover, the fetal cortisol concentrations in this physiological response were clearly not high enough or sustained long enough to initiate parturition in these ewes [33, 34]. The finding that fetal cortisol concentrations are elevated during both maternal and intra-amniotic LPS treatment may have significant clinical implications. Recent evidence from animal experiments suggests that repeated doses of antenatal corticosteroids to fetuses at risk for premature labor may have beneficial effects for fetal lung function, but may also have adverse effects on fetal brain function and fetal growth [35]. Therefore, elevation of fetal cortisol concentrations in response to maternal or intra-amniotic infection, coupled with subsequent antenatal corticosteroid treatment, may be detrimental to the fetus.
Previous studies by Kallapur et al. [18] showed that an intra-amniotic LPS injection of 20 mg resulted in chorioamnionitis, an elevation in proinflammatory cytokine expression in the amnion/chorion, and lung inflammation. We therefore used this model of intra-amniotic LPS administration to further investigate the fetal and maternal responses to endotoxin within the intrauterine environment. In the current study, the presence of LPS in the amniotic fluid induced responses that were mainly restricted to the amniotic and fetal compartments. Intra-amniotic administration of LPS, 400 times the amount that would be sufficient to kill the fetus if given by i.m. injection [18], caused only a moderate increase in uterine activity, and amniotic fluid PGE2 and fetal cortisol concentrations. These observations suggest that a mild fetal inflammatory response occurs following entry of endotoxins into amniotic fluid, although the route of entry of LPS, proinflammatory agents, or both into the fetus remains to be determined. The fetus may swallow or aspirate the amniotic fluid containing proinflammatory agents, and once in the circulation, could lead to the fetal response and possibly sepsis [1]. A number of studies have suggested that the fetus is very sensitive to infectious challenges. The fetal sheep is much more sensitive to LPS compared with that of the adult, with doses of LPS >1 µg/kg often being fatal [11], whereas doses of 1–100 µg/kg are used in studies with adult animals [10, 19]. Our finding that fetal responses were moderate, despite the relatively high dose of LPS administered, may indicate a potential role for the amniotic fluid in protecting the fetus from endotoxin exposure during pregnancy. Preliminary studies investigating the effect of a continuous infusion of a lower dose of LPS into the amniotic fluid (1 mg/h for 24 h, data not shown), to mimic that of a continuous inflammatory challenge, resulted in no change in prostaglandin concentrations, uterine activity, or blood gas parameters. These observations suggest that the infusion of LPS into the amniotic fluid is not as effective in producing the inflammatory responses as seen after bolus intra-amniotic LPS administration. The factors responsible for the higher prostaglandin concentrations in the amniotic fluid after intra-amniotic LPS administration probably involve direct stimulation of the amnion and chorion by proinflammatory cytokines [8]. Alternatively, LPS administration may decrease the metabolism of prostaglandins by decreasing prostaglandin dehydrogenase expression and activity within uterine tissues [36].
To assess the accessibility of endotoxins produced in the fetal membranes to the amniotic fluid and fetus, we created a space between the chorioamnion and the endometrial tissues to allow LPS to enter directly between the fetal membranes and maternal endometrium. Extra-amniotic placement of catheters, similar to that used in the present study, have been successfully used without destroying the integrity of the fluid sacs, and are a useful delivery route to test the effects of substances on myometrial activity [37, 38]. Extra-amniotic administration of LPS at the doses of 0.1, 1.0, or 10 µg/kg per day resulted in no overt fetal or maternal inflammatory responses. In this group of animals we chose to administer LPS as a continuous infusion, because prolonged exposure to bacterial products at sublethal doses have been suggested to be effective in triggering the onset of premature labor [10], and a more appropriate way to investigate fetal inflammatory responses during pregnancy. The small infusion volume was chosen because we did not wish to disrupt the chorioamnion membranes. The finding that no changes were observed in the uterine EMG activity or maternal prostaglandins in these ewes, suggests any inflammatory process induced may have been too limited in area to have created a systemic response. It is interesting that extra-amniotic infusion of PGF2 to late-gestation sheep using the same procedure as that used in this study, increases fetal and maternal plasma PGFM concentrations, but does not alter myometrial activity, suggesting that the ovine myometrium, when under the influence of progesterone, is relatively insensitive to prostaglandins [37]. Although an increase in fetal PGFM concentrations were observed during the infusion of 1.0 µg/kg per day and 10 µg/kg per day of LPS into the extra-amniotic compartment, this was of insufficient magnitude to increase myometrial activity under the influence of progesterone at this time. The lack of effect of LPS within the extra-amniotic compartment may also be explained by the differences in the placentation of sheep, compared with that of humans. In sheep, placentation is less invasive than in humans [39], with the uterine epithelium remaining intact, and is only in loose contact with the chorioamnion. This difference may explain why infectious agents associated with the fetal membranes are more likely to be transmitted to the decidua and subsequently result in premature labor.
The present findings show that maternal LPS treatment caused marked changes in fetal blood gas parameters and sustained increase in fetal prostaglandins and cortisol concentrations. The presence of LPS in the amniotic fluid induces responses that are restricted to the amniotic and fetal compartments, with no overt responses detected in the maternal circulation. The attenuated fetal responses to intra-amniotic LPS administration, despite infusions of LPS at doses 200 times those used in the adult, may indicate a role of the amniotic fluid in protecting the fetus from endotoxin exposure during pregnancy.
|
ACKNOWLEDGMENTS |
|---|
|
FOOTNOTES |
|---|
2 Correspondence: Jonathan Hirst, Department of Physiology, 13F Monash University, Wellington Road, Clayton, Victoria 3800, Australia. FAX: 613 9905 2547; jon.hirst{at}med.monash.edu.au
Received: 25 July 2002.
First decision: 23 August 2002.
Accepted: 26 November 2002.
|
REFERENCES |
|---|
|
TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
|---|
and parturition in the sheep. In: Shizume K, Imura H, Shimuza N (eds.), Proceedings of the 7th Asia and Oceania Congress of Endocrinology. Tokyo: International Congress Series 598; 1982:376–380 This article has been cited by other articles:
|
|
 |
|
  G. Garcia-Lopez, F. Vadillo-Ortega, H. Merchant-Larios, R. Maida-Claros, M. Osorio, D. Soriano-Becerril, H. Flores-Herrera, J. Beltran-Montoya, Y. Garfias-Becerra, and V. Zaga-Clavellina Evidence of in vitro differential secretion of 72 and 92 kDa type IV collagenases after selective exposure to lipopolysaccharide in human fetal membranes Mol. Hum. Reprod., June 1, 2007; 13(6): 409 - 418. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 L. Kabaroff, H. Boermans, and N. A. Karrow Changes in ovine maternal temperature, and serum cortisol and interleukin-6 concentrations after challenge with Escherichia coli lipopolysaccharide during pregnancy and early lactation J Anim Sci, August 1, 2006; 84(8): 2083 - 2088. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 L McClure, A E O'Connor, S Hayward, G Jenkin, D W Walker, and D J Phillips Effects of age and pregnancy on the circulatory activin response of sheep to acute inflammatory challenge by lipopolysaccharide J. Endocrinol., April 1, 2005; 185(1): 139 - 149. [Abstract] [Full Text] [PDF]
|
|
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
