MYH7 mutations induce changes in single cell mechanobiology.

Abstract

Hypertrophic Cardiomyopathy (HCM) is characterized by thickening of the left ventricular wall and hypercontractility and has been linked to mutations in the sarcomere motor protein β-myosin (MYH7). Using single cell mechanobiology studies, we examined how the effects of single point mutations propagate to change the contractile dynamics and cellular morphology (sarcomere spacing, spread area, myofibril alignment) of human induced pluripotent stem cell derived cardiomyocytes (hiPSC-CMs). We micropattern islands of adhesive protein to constraining the spreading and alignment of hiPSC-CM on hydrogel substrates containing fluorescent microbeads as fiducial markers for traction force microscopy (TFM). We deployed substrate stiffnesses ranging from physiological (10 kPa) to heavily diseased/fibrotic (100 kPa) to test the role of increased “afterload” in functional phenotypes. We use image and video analysis to assess the contractile dynamics of the hiPSC-CM in terms of force, power, and velocities of relaxation and contraction. For example, we assessed multiple MYH7 mutations edited into the WTC line along with isogenic controls. Some lines carried an endogenously labeled alpha-actinin GFP reporter of sarcomere structure to enable visualization of sarcomere structure and dynamics. We assessed the magnitude and dynamics of contractile force output from TFM video analysis and observed increased the contractile force when compared to the control hiPSC-CMs. We also measured significantly different dynamics in the relaxation or contraction velocities compared to control hiPSC-CMs. Interestingly, not all HCM mutant lines presented a significant increase in cell spread area, a proxy for hypertrophy, and this correlated with culture conditions, such as the size of the protein pattern constraining the cells or stiffness of the substrate. Taken together, these results suggest a role for MYH7 mutations driving remodeling of structure and function at a cell-intrinsic level via changes in mechanosignaling.