Dr S Harmer, Prof J C Hancox
Applications accepted all year round
Self-Funded PhD Students Only
About the Project
Start 23 September 2019
This MSc by Research is a laboratory-based project which offers the exciting opportunity to spend 1 year in one of our State-of-the-Art laboratories (plus 1 year thesis writing) at the University of Bristol and experience life as a researcher. You will be working with one of our cutting-edge research groups where you will learn new skills and techniques, including experimental design and implementation, data analysis and scientific writing.
Introduction: Cardiac arrhythmias are a major cause of morbidity and mortality. Arrhythmias occur when the normal flow of electricity in the heart becomes disordered. In human ventricular and atrial cardiomyocytes (CMs) the action potential (AP) is coordinated by ion channels, exchangers and transporters which act to depolarise and repolarise in a timely manner. The main repolarising currents are the transient outward potassium current (Ito), the rapid and slow delayed-rectifier potassium currents (IKr and IKs), and the inwardly rectifying potassium current (IK1) 1. The sum of these currents acts to provide a ‘repolarisation reserve’ 2. This reserve is important because if one current is diminished then other currents can compensate which prevents excessive AP prolongation and reduces the chance of arrhythmic events. The critical role of these currents in repolarisation is exemplified by the fact that mutations in the pore forming alpha-subunits that compose IKr, IKs and IK1 cause long QT syndrome and short QT syndrome (LQTS and SQTS). In detail, mutations in KCNQ1 and KCNE1, which encode for the alpha and beta subunits of IKs respectively, cause long QT syndrome types 1 and 5 (LQT1 and 5) 3.
Human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) provide a unique model system for investigating disease mechanisms that underlie arrhythmic disorders and have been used to model LQT1 4-6. As a research model, hiPSC-CMs have a number of advantages in that they are human (and patient specific) in origin, provide an alternative to using cardiomyocytes derived from animals, are genetically editable, and can provide a near renewable supply of cells. However, in comparison to adult cardiomyocytes, hiPSC-CMs are structurally immature and exhibit a late feotal phenotype 7. hiPSC-CMs are also electrophysiologically immature and possess greatly reduced levels of the inwardly rectifying potassium current (IK1) that normally underlies the ‘resting potential’ in CMs 8,9. In addition, in contrast to the findings of the first hiPSC-CM based models of LQT1 4-6, we have found that the kinetics and expression of IKs in hiPSC-CMs is highly variable. Our view, and that of others 10, is that the electrophysiological immaturity and diminished repolarisation reserve found in hiPSC-CMs (due to reduced IK1 and IKs) severely hampers their current utility as a model system in which to study the long QT syndrome.
There is a clear consensus that the immature phenotype of hiPSC-CMs hinders their utility in disease modelling. Excitingly, a number of methods have recently been published (for example 11 and 12) that enable significant and accelerated structural and functional maturation of hiPSC-CMs. However, it is not yet known if these maturation protocols act to enhance IKs expression or promote an improved level of repolarisation reserve.
Project Aims and Objectives: To explore whether recently reported maturation protocols, when applied singly or in combination, can act to enhance the expression of IKs in hiPSC-CMs. If robust expression of IKs in hiPSC-CMs can be engineered this will provide an improved model for characterising the disease mechanisms that underlie the long QT syndrome.
Techniques to be used: Work will be conducted in the Harmer and Hancox laboratories and the project will benefit from the combined technical expertise of both groups. Methods to be used will include:
• Culture of hiPSCs and differentiation into hiPSC-CMs.
• Quantitative real-time polymerase chain reaction (qPCR).
• Western blotting.
• Immunostaining and Confocal Laser Scanning Microscopy.
• Patch-clamp electrophysiology- whole-cell patch-clamp (WCPC) and perforated-WCPC.
Funding Notes
This is a one-year, self-funded Masters (MSc) by research. Fees are £4300 (UK/EU Fee) and bench costs are £5000. Applicants should have (or expect to receive) the equivalent of a First or Upper second-class honours degree in a biomedical discipline.
When applying please select the Faculty of Life Sciences, MSc by Research, School of Physiology, Pharmacology and Neuroscience
References
[1] Tamargo, J. et al. Cardiovasc. Res. 62, 9-33 (2004). [2] Roden, D. M. Pacing Clin. Electrophysiol. 21, 1029-1034 (1998). [3] Schwartz, P. J. et al. Circ Arrhythm Electrophysiol 5, 868-877 (2012). [4] Moretti, A. et al. N Engl J Med 363, 1397-1409 (2010). [5] Egashira, T. et al. Cardiovasc Res 95, 419-429 (2012). [6] Zhang, M. et al. Proc Natl Acad Sci U S A 111, E5383-5392 (2014). [7] DeLaughter, D. M. et al. Dev Cell 39, 480-490 (2016). [8] Ma, J. et al. Am J Physiol Heart Circ Physiol 301, H2006-2017 (2011). [9] Doss, M. X. et al. PLoS One 7, e40288 (2012). [10] Christ, T. et al. Proc Natl Acad Sci U S A 112, E1968 (2015). [11] Parikh, S. S. et al. Circ Res 121, 1323-1330 (2017). [12] Hu, D. et al. Circ Res 123, 1066-1079 (2018).
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