Exploring the potential role of CRTH2 agonists in the prevention of inflammation induced preterm labour

Abstract

Introduction: Preterm labour occurs in 76,000 women in England and Wales each year and accounts for 75% of neonatal mortality. Infection and inflammation are key triggers of preterm labour. The transcription factor NF-ĸB plays a key regulatory role in the expression of the labour associated genes such as COX-2, prostaglandins and MMPs, as well as pro-inflammatory cytokines such as IL-8, IL-1β, IFN-γ and TNF-α. The anti-inflammatory prostaglandin 15-deoxyΔ-12,14 Prostaglandin J2, a CRTH2 agonist can inhibit NF-ĸB and delay preterm labour in a murine model of inflammation induced preterm labour. It is well recognised that the immunology of pregnancy is biased towards a Th2 rather than Th1 response. CRTH2 agonists induce the production of the anti-inflammatory cytokines IL-4 and IL-10 in T-helper 2 cells in vitro, thus have potential to further alter the Th1:Th2 ratio in favour of the Th2 bias. Aims: This thesis examines the role of the CRTH2 receptor and its agonists in pregnancy and parturition, and its potential as a therapeutic target for the prevention of preterm labour. Methods: PCR, western analysis and flow cytometry were used for CRTH2 expression studies. To examine the effect of CRTH2 agonists on NF-ĸB activation, western analysis of p65 and phospho-p65 was performed. The effect of the agonists on interleukin profiles was demonstrated by flow cytometry. Myometrial contractility response to agonists was measured using a myograph. To determine the in vivo effect of a small molecule CRTH2 agonist on inflammation induced preterm labour, CD1 mice were given an intrauterine injection of LPS and the CRTH2 agonist. Delivery outcome and the effect on labour associated genes and interleukins were determined by PCR, western analysis and ELISA. Results: Amniocytes and myocytes show low CRTH2 mRNA levels however do not express the protein at detectable levels. The small molecule CRTH2 agonist Pyl A does not inhibit NF-ĸB in amniocytes and myocytes, indicating that the inhibition of NF-ĸB by 15dPGJ2 is independent of CRTH2. Although 15dPGJ2 was able to inhibit the production of pro-inflammatory cytokines in peripheral blood mononuclear cells, Pyl A was unable to alter the Th2 cytokine profile. Pyl A inhibits myometrial contractility, although in a mechanism independent of CRTH2. In the murine model of inflammation induced preterm labour fetal viability was significantly increased by Pyl A, however, preterm labour was augmented rather than prevented. Conclusion: 15dPGJ2 inhibits NF-ĸB and inflammation induced preterm labour in the mouse in a mechanism independent of CRTH2. Contrary to anticipated, Pyl A does not modulate the Th1:Th2 bias or delay preterm labour in vivo. However, 15dPGJ2 suppresses the Th1 response in vitro, thus represents a potential anti-inflammatory therapeutic target for both the prevention of preterm labour and inflammation induced brain injury.