Parturition, oxytocin, inflammation, myocyte damage and obesity; A study of myometrium and haematological parameters in human pregnancy and labour at term

Abstract

The process of parturition resulting in the delivery of a newborn is a fundamental event ensuring survival of the species. In humans, the main clinical problems of parturition include activation of the process too early or too late resulting in the delivery of pre-term and post-term infants, both with their own implications for future health for the mother and baby. Additionally, where parturition systems are not activated correctly, dysfunctional labour with the resulting need for caesarean delivery (CS), in addition to atonic post-partum haemorrhage can also ensue. Overall, in the UK up to 40% of pregnancies are affected by one of these problems. However, the exact processes involved in the initiation and maintenance of parturition in the human are not fully understood. With such an important event, influences are most likely to be multi-factorial, with hormonal, mechanical, inflammatory, biochemical and maternal environmental factors playing a part. The aims of this thesis were to investigate influences on parturition in human pregnancy. Firstly, the myometrial transcriptional effects of long term exposure to the uterotonic oxytocin (OT) were examined. Further investigation of the myometrial and maternal peripheral response to uterine contractions in-vitro and in-vivo was also made with particular reference to the role of inflammation and myocyte damage. Additionally, the influence of maternal factors, particularly obesity, on the myometrial in-vitro contractile function and response to OT was studied. Initially, 150 gene arrays were produced using the Illumina platform. The samples were derived from myometrium taken at pre-labour CS which subsequently underwent functional contractility experiments in an organ bath. Five drug environments were studied, namely OT, acetic acid (OT vehicle), ML7(a tocolytic acting via inhibition of myosin light chain kinase), ML7 & OT and finally DMSO (ML7 vehicle). Additionally, five time-points of 0, 1, 2, 4, and 6 hours after drug addition were used, resulting in 5 samples for each drug and time combination. The results indicated that despite a clear enhancement of myometrial contractile activity by OT, this functional response does not appear to be mediated by cellular transcription. However, there was a clear contraction and time dependent transcriptional wave, with overrepresentation of genes associated with inflammation and cellular damage/apoptosis, and down-regulation of pathways concerning cellular metabolism. These findings were confirmed by QPCR on further myometrial samples undergoing additional in-vitro functional studies. In addition to the temporal and contractile association with the inflammatory response, our data suggest inflammation occurs in response to myocyte cellular damage regardless of mode of damage e.g. contractile or chemically induced. This was demonstrated by inflammatory upregulation in myometrium exposed to the tocolytic agents nifedipine and ritodrine, which is not seen in response to ML7. Additionally, the myometrial inflammatory response was enhanced by the infective agent LPS. However, contrary to other proposals, the enhanced inflammatory response of the myometrium did not alter or promote the in-vitro contractile ability of the myometrium or its response to OT. This myometrial transcriptional data therefore suggests that the inflammatory response of labour is associated with contraction, chemical or infection induced myometrial cellular damage, but would not be considered necessary for a contractile response. Our in-vivo study of peripheral changes in the maternal circulation again supported our in-vitro myometrial data. Data showed that the effect of pregnancy at term was limited to increased white cell count driven by a neutrophilia, with no suggestion of leukocyte priming prior to labour. Additionally, term pregnancy is associated with an increase in CRP, an increase in GCSF (corresponding with the neutrophilia) in addition to suppression of the chemokines CCL11 and CCL22. Subsequently, we found that repeated blood samples taken at 2 hourly intervals during term labour induced dramatic changes in inflammatory cells and inflammatory mediators in the maternal circulation. Importantly, these changes occur in a co-ordinated time and contraction dependent manner, with the degree of inflammation associated with the length of time in labour and the degree of myocyte damage as measured by circulating CK and Mb. Our study of the influence of maternal factors on myometrial contractile ability and response to OT examined in-vitro myometrial contractility of 609 myometrial strips from 85 women. We demonstrated that maternal obesity does not impair spontaneous or OT induced myometrial contractions in-vitro. Furthermore, maternal age, ethnicity, parity, previous caesarean delivery,gestation at delivery and birthweight do not influence in-vitro myometrial spontaneous or OT induced contractile activity. This therefore suggests that the observed implication of these maternal and infant factors on parturition in-vivo (high rates of induction of labour, high rates of intrapartum caesarean delivery and post partum haemorrhage) cannot be explained by an effect on myometrial contraction per se. This therefore merits further investigation as to alternative mechanisms to ultimately promote and effective, uncomplicated and safe labour and vaginal delivery for at risk mothers. In summary, this thesis provides evidence that the myometrial contractions of human labour, whether spontaneous or OT induced are capable of inducing a temporal wave of transcriptional changes associated with the processes of inflammation, cellular damage/apoptosis with inhibition of cellular metabolic processes. In addition, maternal peripheral circulating factors mirror the myometrial transcriptional changes. These changes are highly comparable with those seen in response to exercising skeletal muscle, and in this model have been shown to play an important role in muscle repair and remodelling after exercise. Therefore, we would suggest that the inflammatory reaction typically associated with human labour occurs as a non-specific response to contraction induced cellular damage and may play a role in postpartum repair and remodelling of the uterus