|Ph.D Thesis||Department of Medicine|
|Supervisor:||Prof. Gepstein Lior|
|Full Thesis text|
Traditional antiarrhythmic pharmacological therapies are limited by their global cardiac action, low efficacy, and significant proarrhythmic effects. In this work, a novel approach for the modification of the myocardial electrophysiological substrate is presented. This approach is based on utilizing cell grafts, genetically engineered to express specific ionic channels, to modulate the excitable properties of host cardiac tissue through electrotonic interactions. To test the aforementioned concept, the work included ex vivo, in vivo, and computer simulation studies to determine the ability of fibroblasts, transfected to express various voltage-sensitive potassium channels (Kv1.3, Kir 2.1 and Kv1.3H401W), to modify the local myocardial excitable properties. Initially, dye transfer experiments were performed in fibroblast-cardiomyocyte co-cultures. These studies demonstrated the formation of structural coupling (formation of functional gap junctions) between the two cell types. Next, whole cell recordings in these co-cultures demonstrated a significant alteration in the cardiomyocyte action potential morphology as a consequence of electrotonic current induced by the coupled transfected fibroblast.
In vivo grafting of the transfected fibroblasts in the rat ventricular myocardium significantly prolonged the local effective refractory period from an initial value of 84±8 ms (cycle length, 200 ms) to 154±13 ms (P<0.01) in the case of Kv 1.3 and even further for the other ion channel types. Margatoxin (a specific, Kv1.3 channel blocker) partially reversed this effect (effective refractory period, 117±8 ms; P<0.01). In contrast, the effective refractory period did not change in non-transplanted sites (86±7 ms) and was only mildly increased in the animals injected with wild-type fibroblasts (73±5 to 88±4 ms; P<0.05). Similar prolongation of the effective refractory period was also found during slower pacing drives (cycle length, 350 to 500 ms) after transplantation of the potassium channels expressing fibroblasts (Kv1.3 and Kir2.1) in pigs. Optical mapping studies in the Langendorff - perfused rat heart preparation demonstrated significant rate-dependent functional conduction blocks at the sites of transplantation of the K channel-expressing fibroblasts. In contrast, grafting of non-transfected fibroblasts did not alter conduction significantly. Finally, computer modeling studies confirmed the in vivo
results, by demonstrating the prolongation of the refractory period that was due to post-hyperpolarization electrotonic interactions between the engineered-fibroblasts and the coupling myocyte. In summary, this work has shown that genetically engineered cell grafts, transfected to express potassium channels, can couple with host cardiomyocytes and alter the local myocardial electrophysiological properties by reducing cardiac automaticity and prolonging refractoriness, providing a possible novel future therapeutic approach for the treatment of cardiac arrhythmias.