IL-1beta-induced changes in intracellular calcium dynamics and TrpC3 expression: delineation of the signal transduction cascade

Lead Research Organisation: King's College London
Department Name: Reproductive Health Endocrinology Dev


Inflammation is a key biological progress that impacts on all aspects of life. It is implicated in fertilisation, embryo development, timing of birth, protection of the living organisms from infection, brain function and ageing. Inappropriate activation of inflammatory processes underlies the development of asthma, arthritis, cardiovascular disease, premature labour and tumour formation. It is essential for scientists, therefore, to fully understand how inflammation influences organisms at the tissue and cellular level. Our project looks how inflammation is involved in priming the pregnant uterus for labour. We seek to understand how an inflammatory agent arriving at the surface of a uterine muscle cell can cause changes in deep within cells in order to switich on specific genes and thereby ultimately make the uterus muscle more capable of contracting powerfully during labour. Knowledge of this process will not only help us understand labour but give new insights into how similar muscles found in blood vessels, airways, gut and the bladder respond to inflammation. This will hopefully inform other researchers attempting to find ways to combat a wide range of diseases and promote human health.

Technical Summary

Activation of a proinflammatory response is essential to numerous physiological processes including embryo implantation, parturition, apoptosis, proliferation, leucocyte recruitment and anticoagulation and pain modulation. Proinflammatory conditions and immune response patterns also appear to serve as a common pathway in a number of diseases including arthritis, asthma, ageing, premature labour, tumour formation and cardiovascular disease. The signal transduction pathways mediating these responses have not been clearly defined. There is, therefore, a need to fully elucidate the cellular processes activated by cytokines in the early and late phases of an inflammatory response and relate this to functional outputs. In particular, it is important to determine the impact of inflammatory mediators on intracellular calcium signals as these modulate gene expression and a wide range of cellular events and functions. Our specific interest concerns the mechanisms by which inflammation primes the uterus to contract prior to labour onset. We have previously reported the novel observation that the proinflammatory cytokine interleukin-1beta induces changes in uterine smooth muscle calcium signalling and enhanced expression of TrpC3, a protein associated with cation channel formation and calcium entry pathways activated by agonist stimulation of PtdIns(4,5)P(2) hydrolysis. The current project focuses on elucidating key signal transduction events between interleukin-1beta binding with its plasma membrane receptor and down stream activation of calcium entry pathways and enhancement of TrpC3 protein expression.
Description Inflammation is a key biological progress that impacts on all aspects of life. The goal of this project was to gain further insight into how inflammation contributes to the priming of the pregnant uterus for labour. We sought to understand how inflammatory cytokines arriving at the surface of a uterine muscle cell could enhance uterine smooth muscle excitability (particularly via intracellular calcium signalling) in order to ensure the uterus contracts powerfully enough in labour.

As a result of this grant, we generated:

i) Novel evidence for STIM and Orai isoforms (proteins associated with calcium signalling) in human uterine smooth muscle and cells. We also identified one of these proteins as alternative candidate for cytokine (IL-1ß)- mediated changes in calcium signalling. STIM and Orai isoforms were found to be modulated in labour and by experimental cell culture conditions. Orai1 was induced by the pro-inflammatory cytokine IL-1ß, but not via MAPkinase or PI3K/Akt pathways.

ii) Increased knowledge of transcription factor activation in human uterine smooth muscle cells. IL-1ß activation of transcription factors appears to be patient (heterogeneous) specific, in contrast to studies in cells maintained in long term culture. This indicates that cell culture studies may not be suitable for this type of work if we really want to understand the processes underpinning labour and delivery in women. Use of fresh human tissue for research is challenging but our group has successfully demonstrated that this is possible.

iii) First evidence for NFATc isoforms in human myometrial tissue and cells. Again, we discovered differences between expression in myometrium tissue taken form women at the time of caesarean section compared to cells extracted from tissues and maintained in culture. Novel demonstration that NFATc2 is differentially modulated by intracellular calcium and labour associated stimuli (mechanical strain and IL-1ß). This indicates that NFATc2 may be important in mediating the changes in uterine function required for labour.
Exploitation Route The technique for effective knockdown of membrane proteins in human myometrium primary cells in short term culture has been shared with researchers in our field (education) and is being utilized for an ongoing MRC project. Whilst 3Rs agenda was not a primary focus of our project, our research strongly promotes the use of primary human uterine smooth muscle cells as an alternative to animal tissues for pregnancy research. Use of primary cells in short term culture is avoided for molecular based studies because of difficulty in obtaining human tissue and the limited quantities of primary cells. However, our work clearly demonstrates that perseverance with human uterine smooth muscle is merited and essential if important biological questions are to be answered. The development of reliable protocols for gene silencing in primary cells makes this cell type an alternative to using transgenic animals.
Sectors Education

Description This grant provided novel data that was presented at international conferences and a recently published paper on the work has been accessed already 913 times online by interested researchers/academics/students across the world (USA, UK, China, India, Russia, Japan, Canada, Switzerland, Germany and Netherlands). A second paper has similarly been accessed (1,586 views and downloads). The data have had impact with regard the 3Rs agenda. A PhD student associated with this project gained her PhD award in 2010 and is now employed in the Financial Services sector.
Sector Education,Financial Services, and Management Consultancy
Impact Types Societal

Description PhD studentship
Amount £77,000 (GBP)
Organisation Biotechnology and Biological Sciences Research Council (BBSRC) 
Sector Public
Country United Kingdom
Start 09/2009 
End 10/2013
Description Project grant
Amount £112,000 (GBP)
Organisation Dr Hadwen Trust (DHT) 
Sector Charity/Non Profit
Country United Kingdom
Start 12/2007 
End 03/2011
Description RO1
Amount $198,564 (USD)
Organisation National Institutes of Health (NIH) 
Sector Public
Country United States
Start 03/2018 
End 03/2023