Abstract Body (Do not enter title and authors here): Introduction: Cardiomyocyte alignment increases over time during normal heart development. This alignment is associated with aligned mechanical tension and plays a key role in electromechanical function of the tissue. Myocardial disarray, in which the alignment is disrupted, is a hallmark of cardiomyopathy. This structural disarray is often accompanied by electrophysiological disfunction. Prior work, including ours, suggests that sodium channel function in particular is enhanced in aligned 3D tissues engineered from human induced pluripotent stem cells (iPSC). We hypothesized that cardiomyocyte alignment in 3D tissue could lead to enhanced sodium channel function by improving sodium channel expression.
Methods: To test our hypothesis, we developed a micro-engineered heart tissue (μEHT) platform in which total mechanical tension could be decoupled from cardiomyocyte alignment. The μEHTs were cultured for at least 7 days in either a 4-post configuration to induce high uniaxial tension (expected to cause cardiomyocyte alignment), or 8-post configuration to induce multiaxial tension (expected to cause cardiomyocyte disarray; Fig. 1a). Optocardiography was carried out to study calcium dynamics (GCaMP6f reporter) and action potential waveforms (voltage-sensitive dye, BeRST-1), in heart muscle engineered from induced pluripotent stem cells (iPSC) derived cardiomyocytes. A sodium channel blocker, Saxitoxin, was added for drug response study. Sodium channel expression was assessed with Western blot and immunofluorescence analysis.
Results: 4-post mEHTs had a lower action potential excitation threshold and faster phase-0 in action potential duration, which suggests improved electrophysiology compared to 8-post EHT. Calcium dynamics data, however, did not show significant difference. Additionally, 4-post EHTs exhibited a more pronounced response to Saxitoxin. Immunofluorescence shows a trend of higher colocalization of NaV1.5 and N-cadherin in the area with high predicted uniaxial tension and cardiomyocyte alignment. Western analysis suggested that aligned mechanical tension led to higher overall expression of Na_V1.5, the cardiac sodium channel (paired t-test n=3, p=0.0315).