Biomedical Engineering - Theses

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    Decoding calcium signalling crosstalk in cardiac hypertrophy
    Bass, Gregory ( 2017)
    The concentration of calcium ions (Ca2+) within a cell is important for governing many different processes across a range of cell types. In heart muscle cells, proper calcium handling is critical for maintaining the rhythmic cycle of contraction and relaxation. Chronic stresses can drive changes in calcium signalling which trigger heart cells to grow in size. This process is called hypertrophy and is a common cause of heart failure. It was unknown how a cell could distinguish calcium signals leading to cell growth from those leading to contraction. The central hypothesis explored in this thesis is that calcium can simultaneously yet specifically effect two distinct responses in a cardiomyocyte by altering the shape of the cellular calcium transient, but how this might be achieved without disrupting contractile function was unknown. New line-scan data shows that IP3R-mediated Ca2+ release widens the cellular transient following RyR-mediated Ca2+-induced Ca2+-release events. The data supports the hypothesis that RyR and IP3R systems interact by inducing a global yet transient elevation in Ca2+. A mathematical model of the whole-cell adult rat ventricular myocyte Ca2+ transient was developed by combining existing models of RyR and IP3R and fitting to the line-scan data. The model includes the two major compartments that interpret the calcium signalling and provide spatial separation -- the cytosol which achieves the calcium-dependent contractile response, and the nucleus which achieves calcium-dependent gene expression. The compartmental model was found to reproduce the observed shape of the Ca2+ transient, but only if IP3Rs exhibit a refractory response. This difference in time-course kinetics could underlie the signal separation between cell contraction and cell growth. Advanced immunofluorescence imaging and statistical methods were used to map the spatial positioning of RyRs and IP3Rs in adult rat heart cells. A statistical tool was developed to simulate physiologically-realistic protein distributions on images of the cell architecture. A spatial model of the Ca2+ transient based on realistic RyR distributions showed that both cellular architecture and the distance between RyR clusters could affect local signalling events. For the first time, super-resolution data was used to establish the relationship between RyRs and IP3Rs. Data analysis indicated that cardiomyocyte-specific IP3R cluster distribution reduces the effective spatial distance between RyR clusters which may promote Ca2+ signal propagation or which may enhance Ca2+ signal strength and longevity. This research combining mathematical modelling and advanced imaging has shown that RyR and IP3R proteins co-exist within the same areas of the heart cell and can modify the normal contractile signal without disrupting it. The modification of the Ca2+ signal may subsequently be interpreted by the cell as a signal to alter calcium-dependent gene expression. This finding is important because it reveals how the cell growth signal might be encoded in the heart cell, and this encoding mechanism may be extensible to many other signaling processes in different cells.