Myometrial Activation and Preterm Labour: Evidence Supporting a Role for the Prostaglandin F Receptor—A Review

Placenta (2003), 24, Supplement A, Trophoblast Research, Vol. 17, S47–S54 doi:10.1053/plac.2002.0938

Myometrial Activation and Preterm Labour: Evidence Supporting a Role for the Prostaglandin F Receptor—A Review D. M. Olsona,b,c, D. B. Zaragozaa, M. C. Shallowc, J. L. Cooka, B. F. Mitchella, P. Grigsbyd and J. Hirstd The Perinatal Research Centre, the CIHR Group in Perinatal Health and Disease, a Departments of Obstetrics and Gynaecology, b Pediatrics and c Physiology, The University of Alberta, Edmonton, Alberta, Canada, d Department of Physiology, Monash University, Clayton, Victoria, Australia Paper accepted 30 December 2002

An increase in the myometrial expression of the prostaglandin (PG) receptors, and especially the PGF2
receptor (FP), may be an important component of the process initiating preterm labour. In this review of the literature and presentation of new possibilities, evidence will be discussed that demonstrates an increase in mouse uterine FP mRNA occurs at preterm birth whereas uterine PGF2
concentrations do not increase, suggesting elevated uterine receptor expression and sensitivity is a mechanism for preterm labour initiation. The first examination of the complete human myometrial FP promoter will be described and evidence presented that demonstrates the pro-inflammatory cytokine, interleukin-1, stimulates FP mRNA expression. Finally new data showing that administration of a specific FP antagonist delays preterm birth in sheep will be presented.  2003 IFPA and Elsevier Science Ltd. All rights reserved. Placenta (2003), 24, Supplement A, Trophoblast Research, Vol. 17, S47–S54

INTRODUCTION The prevention of preterm birth is considered the most important perinatal challenge facing industrialized countries (Moutquin and Papiernik, 1990; Berkowitz and Papiernik, 1993). Sadly, after more than 40 years of research, effective therapy has not been developed due to many reasons, including maternal and foetal side effects of many treatments. In this review we will discuss the evidence that highlights a role for prostaglandin (PG) receptors in preterm labour initiation and present new concepts for further experimental and possibly therapeutic consideration.

UTERINE ACTIVATION, STIMULATION AND PRETERM BIRTH It is well accepted that uterine contractile activity at birth is the product of enhanced uterine activation and increased levels of contractile stimulators (Lye and Challis, 1989). Several uterine activation proteins (UAPs) enhance the ability of the myometrium to respond to contractile agonists and to develop and propagate the depolarization signal efficiently between smooth muscle cells. These UAPs include the oxytocin receptor e

To whom correspondence should be addressed at: 220 HMRC, The Perinatal Research Centre, The University of Alberta, Edmonton, Alberta, Canada T6G 2S2. Tel.: 780-492-2765; Fax: 780-492-1308; E-mail: [email protected] 0143–4004/03/$-see front matter

(OTR), PGF2
and PGE2 receptors (FP, EP1–4), and the primary gap junction protein, connexin-43 (Cx-43). More recently we have suggested that prostaglandin endoperoxide H synthase (PGHS-2) be added to this group due to the similarities with the other UAPs in its expression (Cook et al., 2000). The stimulators include oxytocin and PGs (Lye and Olson, 1996). The PGs are now recognized as the ‘triggers’ of labour (Challis and Olson, 1988) because the myometrium contracts in response to exogenous PGs, in vivo and in vitro (Senior et al., 1991, 1992, 1993; Crankshaw and Gaspar, 1992), PG synthetic enzymes and levels in tissues and fluids increase before or at the time of labour (Hirst et al., 1995a, 1995b, 1998; Mijovic et al., 1997, 1998a, 1998b, 1999; Slater, Allport and Bennett, 1998; Slater et al., 1999), and inhibitors of PG synthesis delay birth and prolong pregnancy (Challis and Olson, 1988; Poore, Young and Hirst, 1999; Grigsby et al., 2000).

PROSTAGLANDIN RECEPTORS Prostaglandins were originally thought to interact directly with the cell membrane, thereby altering its characteristics and modulating cell physiology. However their different activity profiles and generation of second messengers, principally cyclic AMP (cAMP), phosphatidylinositol turnover and Ca2+ , suggested that they interact with discreet receptors and that different receptors exist for each PG (Kennedy et al., 1982).  2003 IFPA and Elsevier Science Ltd. All rights reserved.

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In the 1980s, studies by Coleman and colleagues using specific antagonists further characterized these receptors pharmacologically (Coleman, Smith and Narumiya, 1994). They classified the receptors as one each for PGs D2, F2
, I2, and TXA2, as DP, FP, IP, and TP, and four for PGE2, as EP1–4. The first receptor that was isolated and cloned was the TP receptor (Ushikubi et al., 1989; Hirata et al., 1991). The protein is a G-protein linked member of the rhodopsin family of receptors with seven transmembrane domains. Subsequently, homology screening using mouse cDNA libraries led to the identification of the structures of all eight receptors (Narumiya, Sugimoto and Ushikubi, 1999), each encoded by a different gene. More recently, several splice variants have been identified for human EP3 (8 isoforms), sheep and bovine FP (2 isoforms) (Pierce et al., 1997; Ishii and Sakamoto, 2001), rat EP1 (2 isoforms) and human TP (2 isoforms) (Pierce and Regan, 1998). They vary largely in their cytoplasmic C-terminal regions where coupling with G proteins occurs. The amino acid sequences have been fully described for human and mouse receptors, and partially described for rat and sheep. There is only 20–30 per cent homology between amino acid sequences of various receptors within a species, but 79–89 per cent homology exists between like receptors of different species. Prostaglandin F2
action is mediated by the FP receptor, which is coupled to G-protein mediated signal transduction pathways leading to the mobilization of intracellular Ca2+ . The human FP receptor consists of 359 amino acid residues with a predicted molecular mass of 40 060 Da (Abramovitz et al., 1994). In human myometrium, PGE2 interacts with EP1 receptors, which elevate [Ca2+ ]i independently from PLC, but involving a Gi protein and plasma membrane calcium channels; EP2 receptors stimulate adenylate cyclase; EP3A receptors inhibit adenylyl cyclase activity through Gi activation; and EP3D receptors activate PLC through a pertusis toxininsensitive pathway and also elevate [Ca2+ ]i (Asboth, Phaneuf and Lopez Bernal, 1997). Overall, the receptors can be grouped into two categories; the EP1, EP3 FP and TP stimulate contraction, and the relaxatory receptors are DP, EP2, EP4 and IP.

PG RECEPTORS IN PREGNANCY AND BIRTH PG receptor expression varies considerably from the nonpregnant state through pregnancy to birth, and the relative level or type of receptors may dictate the degree of uterine quiescence or contractility. The expression of the contractile EP3 gene in pregnant human myometrium was 40 per cent less than that in non-pregnant myometrium, and expression of the FP gene in human myometrium also decreased during pregnancy by 45 per cent compared to levels in non-pregnant myometrium (Matsumoto et al., 1997). Similarly, uterine FP mRNA expression declined significantly with gestational age in patients not in labour and then at term increased significantly with labour (Brodt-Eppley and Myatt, 1999). These data

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correlate very well with the in vitro human myometrial contractile data. The contractile ED25 for PGF2
administered to nonpregnant human myometrium, in vitro, was 0.04 nmol versus the much higher 0.5 nmol in non-labouring term tissue. The values were similar for contractile PGE2 responses suggesting the number of contractile receptors is reduced during pregnancy to maintain uterine quiescence (Senior et al., 1991, 1992, 1993). Conversely, labour in the pregnant baboon myometrium was associated with a lower abundance of the relaxatory EP2 receptor mRNA (Smith, Wu and Nathanielsz, 1999). In the myometrium of pregnant rats, expression of FP mRNA increased significantly from late gestation until delivery, returning to prepartum levels by one day postpartum while the levels of the relaxatory EP2 receptor were the inverse to those of FP (Brodt-Eppley and Myatt, 1998; Al-Matubsi et al., 2000). In one exception, rat myometrial FP mRNA levels were higher on GD18 than in nonpregnant tissues, but were statistically similar to high term levels and may have already begun the labour-associated rise (Dong and Yallampalli, 2000). The myometrium of the sheep also displays a significant increase in the mRNA expression of FP and EP3 with the onset of labour at term, but interestingly, there is also an increase in the relaxatory EP2, which may have some role for passage of the foetus (see below) (Wu et al., 1999). The maintenance of pregnancy is likely related to an increase in relaxatory EP receptor responses (primarily EP2) (Senior et al., 1993) as well as increased coupling of G
s to adenylate cyclase (Europe-Finner et al., 1994). There is much less of this coupling at parturition (Europe-Finner et al., 1994). These events correlate with an increase in EP2 mRNA expression in the myometrium of women late in gestation, and not in labour, when compared to women at term not in labour (Brodt-Eppley and Myatt, 1999). Data from pregnant baboons (Smith, Wu and Nathanielsz, 1999) (as mentioned above) and mice (Docherty et al., 1999) support these findings. Thus, myometrial active quiescence may change to an active contractile state in concert with an up-regulation of contractile receptors and loss of relaxatory receptors and decreased cAMP production. The location of receptors within the tissue may also be important. Contraction-promoting EP1 and EP3 receptors are found in greater abundance in the fundus whereas relaxationpromoting EP2 and EP4 receptors are found in the lower uterine segment (Smith, Wu and Nathanielsz, 2001), where they may be involved in uterine relaxation to allow passage of the foetal head and shoulders. However, the lower segment responds by contraction to PGF2
challenge and lower doses of PGE2 in in vitro studies (Senior et al., 1993; Crankshaw and Dyal, 1994), so it may have the dual function of contraction and relaxation. Only one study has examined the relative levels of PG receptors at human preterm birth (Brodt-Eppley and Myatt, 1998). FP mRNA levels from lower uterine segment myometrium were higher at preterm birth with and without labour than at term without labour. The EP2 mRNA levels were highest at preterm not in labour and decreased to term. None

Olson et al.: FP Receptor at Preterm Labour

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of the patients had clinical signs of infection, but a direct examination of tissues or amniotic fluid for signs of infection (Armer and Duff, 1991; Romero et al., 1993) or stratification of patients to gestational age is warranted to identify more subtle changes in the gestational situation. Such information may provide clues as to potential regulation of the genes for the PG receptors.

MECHANISMS REGULATING PG RECEPTOR EXPRESSION Surprisingly, there is little work published on the regulation of the PG receptors in uterine myometrium. In rat uterus, EP2 expression increased during pregnancy then decreased in labour, in accordance with its role as a relaxatory receptor (Dong and Yallampalli, 2000). Interestingly, FP rose in pregnancy compared to the nonpregnant state, but rose even more during labour. Administration of the progesterone receptor antagonist, RU486, increased FP and decreased EP2 mRNA. Further, antagonism of estrogen action decreased FP mRNA whereas administration of estradiol enhanced FP expression. Progesterone administration alone promoted EP2 expression, and when this was supplemented with estradiol, the increase was greater. Hence steroids regulate PG receptors in the rat. Similarly, little is known of the cellular regulation of PG receptors. Some information may be available from studies examining the other UAPs that have implicated proinflammatory cytokines in their regulation. For instance, PGHS-2 mRNA increases in human myometrium prior to the onset of term labour (Slater et al., 1999), and transcription of PGHS-2 is stimulated by IL-1 through activation of the transcription factor, NFB (Belt et al., 1999). In cultured human ULTR myometrial cells, IL-1 (10 ng/ml for 24 h) stimulated an 8-fold increase in FP mRNA abundance compared to untreated control levels (6.871.69 units normalized to GAPDH vs. 0.850.12 units, n=4, P<0.05) (Zaragoza and Mitchell, unpublished observation). This is the same timecourse as for IL-1-stimulated FP mRNA expression in human granulosa-luteal cells (Narko, Ritvos and Ristimaki, 1997). A potential negative regulator of FP expression is its own ligand, PGF2
. Both exogenous and endogenous PGs decreased the number of binding sites in rat myometrium, suggesting ligand-induced receptor down regulation (Molnar and Hertelendy, 1990). Srinivasan demonstrated that PGF2
induced the internalization of FPA in HEK-293 cells (Srinivasan, Fujino and Regan, 2002). The FP gene has been sequenced from human (Abramovitz et al., 1994), bovine (Sakamoto et al., 1995), mouse (Sugimoto et al., 1994), and sheep (Graves et al., 1995), and there is general agreement in the literature on its structure. The gene is approximately 10 kb in length and is composed of 3 exons separated by 2 introns. The translation initiation start site is located within exon 2, while exon 1 is non-coding and exon 3 also contains large regions of non-coding sequence. Two

Figure 1. Structure of the 5 end of the human FP gene including potential 5 regulatory domains. This genomic fragment of 4107 bp was isolated and completely sequenced. The transcription initiation start site of the human FP receptor (exon 1) was identified using RNA-Ligase Mediated 5 -Rapid Amplification of cDNA Ends (FirstChoice RLM-RACE Kit from Ambion). This technique provides positive selection for amplification products that contain the true 5 end of the desired mRNA and prevents the detection of false 5 ends due to RNA degradation. Genomic DNA containing the 5 end of FP was amplified by nested PCR and fragments were cloned, sequenced and analysed for possible transcription factor binding sites found to be important for regulation of other uterine activation genes using MatInspector.

mRNA species are reported to arise from alternative splicing within exon 3 of the ovine FP gene, and have been called the FPA and FPB isoforms (Pierce et al., 1997). In contrast to the FP gene, the FP promoter has not been well characterized. Four groups have published the DNA sequence from the 5 flanking region of the human (Betz et al., 1999), mouse (Hasumoto et al., 1997), bovine (Ezashi et al., 1997), and rat (Neuschafer-Rube, Moller and Puschel, 2000) FP gene. A sequence of 421 bp was reported from the 5 flanking region of the human FP gene (Betz et al., 1999). No attempt was made to identify the transcription initiation start site or transcription factor binding sites in this report, and the authors did not suggest that this sequence contains the entire FP promoter. The 1500 bp sequence was also reported for the 5 flanking region of the mouse FP gene (Hasumoto et al., 1997), however this report also contained minimal information regarding transcription factor binding sites and no conventional transcription start site was identified. The characteristics of the rat (Neuschafer-Rube, Moller and Puschel, 2000) promoter were similar to the mouse (Hasumoto et al., 1997). Ezashi and colleagues analysed 1818 bp upstream of the bovine FP gene, and sequences internal to intron 1, for consensus transcription factor binding sequences (Ezashi et al., 1997). Unlike the human and mouse studies, the bovine FP promoter was reported to contain a number of putative transcription factor binding sequences and two transcription start sites (Ezashi et al., 1997). Thus, the potential species differences in the promoter sequence, the short amount of sequence reported and the lack of detailed information on cis-acting regulatory elements for the human and mouse FP gene promoter all indicated the necessity of characterizing and analysing the FP promoter. Therefore a 4.1 kb fragment containing intron 1, exon 1 and 2.5 kb upstream of the human FP gene was isolated, cloned and sequenced (Figure 1) (Zaragoza & Olson, unpublished). The transcription start site at the beginning of exon 1,

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Figure 2. Uterine levels of PGF2
and FP mRNA across late gestation and at the time of preterm labour induced by ethanol treatment (EtOH, 6 mg/kg, s.c., on gestational day (GD) 16.0), ovariectomy (OVX on GD 16.0), or lipopolysaccharide treatment (LPS (Sigma, serotype 026), 50 µg, s.c., twice on GD 16 at 09 : 00 and 11 : 00). PGF2
concentrations in whole uterine tissue were determined by ELISA and normalized to protein (pg PGF2
/mg protein), and FP mRNA concentrations were determined by ribonuclease protection assay and normalized to cyclophilin mRNA. Mean, n=3–12 for each bar, significance achieved at Pc0.05. * Denotes a significant difference from gestational day (GD) 16 and GD 17 (ANOVA followed by Tukey’s test). # Denotes significant difference from gestational age-matched control (GD17.75 for EtOH induced preterm labour and GD17 for OVX or LPS induced preterm labour) (Student’s t-test). Data modified from Cook et al., 2000 (left panel) and Cook et al., 2003 (middle panel).

several AP-1 sites, a STAT-1 site, a potential Y estrogen response element (ERE), and a Y progesterone response element (PRE) were identified. In addition, a NFB site was identified. Hence cytokines and steroids are possible regulators of FP expression at term and preterm.

THE FP RECEPTOR IN PRETERM BIRTH IN MICE The relationship of the FP receptor to other UAPs at preterm birth was studied in C57BL/6J mice because the UAPs are easily regulated in this model. Three separate means were used to induce preterm birth: the administration of ethanol (6 mg/ kg) on gestational day 16.0 (GD16), ovariectomy (OVX) on GD16, or administration of lipopolysaccharide (LPS) on GD16. Each manipulation caused a loss of ovarian progesterone (P4) that led to birth during GD17, about 2 days ahead of normal term on GD 19.30.2 (Cook et al., 1999, 2000, 2003; Mijovic et al., 2002). The relative abundance of mRNA expression of the uterine FP receptor and the content of PGF2
in uterine tissue was determined on GD16 and GD17 in non-labouring control pregnant mice, in mice at term birth on GD 19.3, and in mice undergoing preterm labour on GD17 (Figure 2). In control mice, FP mRNA increased at term birth in relation to non-labouring control dams at GD16 and GD17, and similarly, levels of FP mRNA in mice undergoing preterm labour were higher than gestational age-matched controls. The uterine concentrations of PGF2
increased steadily from GD16 to term labour, but there was no increase at preterm birth relative to non-labouring control dams at GD17. Yet, even though the uterine PGF2
concentrations did not increase at

preterm birth, they were sufficiently high to stimulate contraction of the myometrium because administration of inhibitors to the PG synthesizing enzymes, PGHS-1 or PGHS-2, which lower endogenous PG levels, prevented preterm birth (Cook, Randall and Olson, 1998; Cook et al., 1999). These data suggested that increased uterine sensitivity to PGs rather than an increase in the levels of PGs was sufficient for preterm birth to occur.

EFFECT OF FP KNOCKOUT IN MICE Mice in which the FP gene is deleted become pregnant, but hold their pups past normal term because luteolysis is prevented and the fall in P4 does not occur (Sugimoto et al., 1997; Cook et al., 2003). However, ovariectomy on GD19 in FP knockout mice leads to a precipitous drop in circulating P4 concentrations and normal birth at term on GD20 (Tsuboi et al., 2000) that is associated with mRNA increases in both OTR and PGHS-2. These data appear to be contradictory to the notion that FP is a key component to labour because it is dispensable to the birth process. However, when the other uterine activation proteins are examined, none can be proven to be essential to the birth process. The oxytocin knockout mouse delivers at the usual time of term birth (Nishimori et al., 1996; Young et al., 1996); the OTR knockout is lethal to the embryonic mouse (Lou Muglia—personal communication); the PGHS-1 knockout mouse does not deliver on time, but that is due to the loss of PGF2
for luteolysis (Langenbach et al., 1995; Gross et al., 2000); the PGHS-2 knockout mouse rarely survives to adulthood, but when it does, it cannot reproduce because PGHS-2 is required for ovulation (Dinchuk

Olson et al.: FP Receptor at Preterm Labour

Figure 3. Relative changes in uterine activation protein mRNA and PGF2
concentrations in the mouse uterus. Comparisons of relative mRNA values (ribonuclease protection assays normalized to cyclophilin mRNA) or uterine PGF2
concentrations at preterm birth GD 17.00.2, induced by OVX on GD 16), term birth or post-term birth [GD 20.90.4, delayed by progesterone pellet implant (released 2 mg/day)] that were all made to GD 17 control, Student’s t-test, P<0.05 (actual data not shown). Comparisons of relative mRNA or uterine PGF2
concentrations at P4 Term delayed birth were made to normal term birth values, Student’s t-test, P<0.05. An up arrow indicates an increase relative to control, a down arrow indicates a decrease, and a dash represents no change in expression or concentration from the appropriate control value. Reproduced with permission from Cook et al., 2003.

et al., 1995; Morham et al., 1995); and connexin-43 (CX-43) knockout foetuses die at birth as a result of a failure in pulmonary gas exchange caused by a swelling and blockage of the right ventricular outflow tract from the heart (Reaume et al., 1995). We used other means, therefore, to address the need for the four uterine activation proteins in mice by studying their mRNA levels and uterine PGF2
concentrations at term birth, preterm birth induced by ovariectomy, and under conditions in which birth was delayed at term by an implanted pellet that released progesterone at a low rate (Cook et al., 2003). These treatments created conditions of variable expression of the UAPs (summarized in Figure 3). These data demonstrate that the FP receptor mRNA increases at each birth—preterm, term and postterm, that its expression is decreased at expected term birth when elevated P4 levels blocked birth, and that it is the only UAP that exhibits these characteristics. Further, birth occurs if expression of three of the four UAPs are increased, suggesting that redundant mechanisms exist to effect birth. But when only two of the UAPs increase their expression, birth does not occur. Thus it may be possible to delay preterm birth by blocking the expression or action of two of the UAPs. By extension, antagonizing the activity or action of either PGHS-2 or FP, effectively blocks two of the UAPs, and this may be the reason selective PGHS-2 inhibitors are so effective in delaying preterm labour (Poore, Young and Hirst, 1999). This concept also predicts that antagonizing the FP receptor will delay preterm labour. The following studies tested this concept using a specific antagonist of FP action. EFFECT OF A SPECIFIC FP ANTAGONIST IN MICE AND SHEEP THG113 is a specific, noncompetitive, reversible peptide inhibitor of the FP receptor that blocks the interaction of the

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Figure 4. THG113.31 delays RU486-induced preterm labour in sheep. When the uterine EMG activity reached a level of twice the average 12 h control (pre-RU486) levels for two-2 h epochs, THG113.31a was infused in a stepwise fashion: 53 µg/kg bolus followed by an infusion of 53 µg/kg/h for 2 h, then 105 µg/kg bolus and 105 µg/kg/h infusion for 2 h, then 210 µg/kg bolus and 210 µg/kg/h infusion for 2 h, then 420 µg/kg bolus followed by 420 µg/kg/h infusion for 8 h during which uterine EMG activity maintained a constant level. Approximately 2–3 h after the cessation of the THG113.31a infusion, EMG activity rose quickly and the ewe delivered at 41.3 h after the RU486 injection.

receptor with G
q, ultimately preventing the increase in intracellular [Ca2+ ]. It binds specifically to FP-expressing but not to native (non FP-expressing) HEK293 cells (Peri et al., 2002). In FP-expressing HEK293 cells, THG113 significantly reduces PGF2
-induced IP3 generation with an IC50 of 30 n. Similarly, PGF2
-stimulated retinal arteriole contraction is non-competitively blocked (ca. 90 per cent, IC50 340 n) by THG113 whereas neither agonists requiring homologous receptors (PGE2 (EP1 receptor) and U46619 (TP receptor)) nor numerous other contractile agents (e.g. PAF, endothelin, or angiotensin II) are affected by THG113. Only 1 per cent or less crosses the mouse placenta (K. Peri, personal communication), a labyrinthine, hemotrichorial model (two layers of syncytiotrophoblast and a single mononuclear cell layer) (Beaudoin, 1980; Leiser and Kaufmann, 1994). The rate of transfer across other placenta types, especially the hemochorial human placenta is unknown, but hopefully is small, thereby minimizing the effect of THG113 upon the foetus and qualifying THG113 as a candidate tocolytic. THG113 was effective in delaying preterm birth induced by administration of LPS (50 g, ip, twice at 3 h interval) on GD 16 to pregnant mice (Quiniou et al., 2001). In control dams, preterm birth occurred within 18 h of LPS administration in 100 per cent of cases. THG113 administered (10 mol/kg bolus followed by 0.8 mol/kg/h) 4–6 h after administration of LPS, when the PG effect of luteolysis was past, delayed preterm birth >40 h in LPS-treated dams by antagonizing PGF2
responses in the uterus. Fetuses from THG113 treated dams were born alive with higher birth weights than controltreated dams and appeared healthy. Increases in the abundance of contractile FP and EP3 mRNA and in the relaxatory EP4 mRNA were evident in sheep myometrium at term in one study (Wu et al., 1999) but not in

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another study (Gyomorey et al., 2000). No studies have explored the changes in receptors at preterm birth in sheep to our knowledge. We therefore determined that a version of the FP receptor antagonist, THG113.31a, is effective in delaying RU486-induced preterm labour and delivery in a sheep infused for 9 h (the length of the infusion) beyond the time an untreated ewe delivered, indicating that FP is involved in generating uterine contractile activity at preterm labour (Hirst, Grigsby, Peri and Olson, unpublished) (Figure 4).

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term and preterm birth in several species. Determining the level of FP or other PG receptor expression in infectionassociated preterm deliveries or in births c30 weeks or >30 weeks’ gestational age may provide an indication of the relative importance of these receptors under these conditions. Antagonists to FP action may be effective in delaying preterm birth and should be developed and tested. Elucidating the intracellular mechanisms of FP mRNA stimulation may reveal convergence of mechanisms that regulate expression of other uterine activation proteins and thereby other potential tocolytic targets.

FUTURE DIRECTIONS The data from our laboratory and several others clearly shows that increases in FP mRNA or protein occur in the uterus at ACKNOWLEDGEMENTS The authors are funded by the Canadian Institutes of Health Research, the Alberta Heritage Foundation for Medical Research, and the National Health and Medical Research Council of Australia. Dr Krishna Peri of Theratechnologies kindly provided THG113.31a. Ms Sheila McManus provided valuable assistance with the preparation of the manuscript.

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