Improving the relaxing effect of terbutaline with phosphodiesterase inhibitors: Studies on pregnant rat uteri in vitro
Introduction
Preterm uterine contractions can develop as a result of several pathological processes, among which intrauterine infection plays a key role (Romero et al., 1988). The inflammatory processes trigger a wide range of uterus-contracting factors, which eventually lead to the development of uterine contractions (Lindström and Bennett, 2005). At the same time, the contractions in the pregnant myometrium are counterbalanced in part by cyclic adenosine-monophosphate (cAMP)-dependent relaxation mechanisms. A plethora of ligands such as β2-adrenergic receptor (β2-AR) agonists can stimulate the generation of cAMP in the myometrium, whereas phosphodiesterase enzymes (PDEs) warrant termination of signalling by the degradation of cAMP to the inactive 5′-AMP (Houslay 2003).
We earlier demonstrated that the in vitro uterus-relaxing potency of the β2-AR agonist terbutaline is enhanced in pregnant rats challenged with bacterial lipopolysaccharide (LPS) to evoke preterm birth. We also found that, in case of local inflammation, terbutaline has an increased cAMP-accumulating potency, which is probably mediated by the sensitization of adenylyl cyclase by tumor necrosis factor-α (TNF-α) (Klukovits et al., 2009). On the other hand, the local PDE activity may as well be an important regulator of cAMP-dependent relaxation processes.
The PDE enzymes are encoded by 11 related gene families and are differentially expressed in human tissues (Lugnier, 2006). Human myometrial cells (Méhats et al., 1999), amniochorionic membranes (Oger et al., 2005), monocytes and neutrophils (Wang et al., 1999) express PDE4, which promotes its role in either smooth muscle or immune functions. At present, PDE4 comprises the largest PDE family, constituted by 4 genes with various alternative mRNA splices encoding at least 35 different PDE4 isoenzymes. Rolipram is the archetypal PDE4 inhibitor (Ki = 0.8 μM); and no selective inhibitor is currently available that discriminates PDE4 isoenzymes (Lugnier, 2006).
The aim of the present study was to test the effects of non-selective PDE or selective PDE4 inhibitors alone or in combination with β2-AR agonists on uterine contractions in vitro. We tested the effects of the non-selective PDE inhibitor theophylline and the specific PDE4 inhibitor rolipram on isolated uterine rings from intact and LPS-treated late-pregnant rats. We also investigated the effects of β2-AR agonist terbutaline with PDE4 blockade in the same experimental model. Uterine cAMP levels were measured by means of enzyme immunoassay (EIA) after stimulation with the direct adenylyl cyclase activator forskolin, in the presence of rolipram. The accumulation of cAMP was also detected in the presence of terbutaline and rolipram alone, and their combination.
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Animals
The animals were treated in accordance with the European Communities Council Directives (86/609/ECC) and the Hungarian Act for the Protection of Animals in Research (XXVIII.tv.32.§). All experiments involving animal subjects were carried out with the approval of the Hungarian Ethical Committee for Animal Research (registration number: IV./01758-2/2008) and under the control of the ISO-9001:2008 Quality Management System.
Sexually mature female Sprague–Dawley rats (body mass: 140-160 g, 50–60 days
In vitro contractility studies
Uterine activity was characterized by the AUCs of KCl-stimulated contractions of uterine rings in vitro. Time control experiments revealed that KCl-stimulated contractions do not decrease significantly through the experiment (40–45 min). Although the non-specific PDE inhibitor theophylline had very limited effect in the uteri of intact rats, it showed significant effect in LPS-treated rats (Fig. 1). In the intact rats, the effect of theophylline varied, depending on the day of pregnancy: the
Discussion
Most of the interventions intended to reduce preterm birth rate have not achieved consistent benefit. Apart from manifest infections in late-pregnancy, when the termination of pregnancy is of utmost importance to save the mother and the infant, the causes of preterm contractions are seldom recognized before the initiation of tocolytic therapy. The administration of uterorelaxant drugs might provide time for obstetrical interventions such as screening for asymptomatic bacteriuria, antenatal
Conclusion
In the light of our results, we can conclude that the uterus-relaxing effect of terbutaline was increased notably in the presence of the PDE4 inhibitor rolipram. The combination of terbutaline + rolipram seems favorable to promote myometrium relaxation, especially in the case of genital inflammation, which condition is liable to trigger preterm birth.
Conflict of interest
No conflicts of interest.
Acknowledgement
This work was supported by the Hungarian OTKA Research Grant (K62707).
References (26)
- et al.
Animal models of preterm birth
Trends Endocrinol Metab
(2004) - et al.
Anti-inflammatory and analgesic effects of the phosphodiesterase 4 inhibitor rolipram in a rat model of arthritis
Eur J Pharmacol
(2000) Phosphodiesterase 4 and tolerance to beta 2-adrenoceptor agonists in asthma
Trends Pharmacol Sci
(1996)- et al.
Different roles of alpha2-adrenoceptor subtypes in non-pregnant and late-pregnant uterine contractility in vitro in the rat
Neurochem Int
(2007) - et al.
Primary, secondary, and tertiary interventions to reduce the morbidity and mortality of preterm birth
Lancet
(2008) Phosphodiesterase-4 inhibitors for asthma and chronic obstructive pulmonary disease
Lancet
(2005)Cyclic nucleotide phosphodiesterase (PDE) superfamily: a new target for the development of specific therapeutic agents
Pharmacol Ther
(2006)- et al.
Effects of rolipram, a selective inhibitor of type 4 phosphodiesterase, on lipopolysaccharide-induced uveitis in rats
Investig Ophthalmol Vis Sci
(2004) - et al.
Acute tocolysis for uterine activity reduction in term labor — a review
Obstet Gynecol Surv
(2008) - et al.
Update on the therapeutic potential of PDE4 inhibitors
Expert Opin Investig Drugs
(2002)
beta-Adrenergic agonists exert their “anti-inflammatory” effects in monocytic cells through the IkappaB/NF-kappaB pathway
Am J Physiol Lung Cell Mol Physiol
Pregnancy-induced decrease in the relaxant effect of terbutaline in the late-pregnant rat myometrium: role of G-protein activation and progesterone
Reproduction
Anti-inflammatory and immunomodulatory potential of the novel PDE4 inhibitor roflumilast in vitro
J Pharmacol Exp Ther
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