Towards an active matter perspective on sarcomere dynamics in cardiomyocytes
by Daniel Härtter
Date of Examination:2023-05-23
Date of issue:2023-04-19
Advisor:Prof. Dr. Christoph F. Schmidt
Referee:Prof. Dr. Christoph F. Schmidt
Referee:Prof. Dr. Jörg Enderlein
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EnglishThe contraction of cardiac muscles emerges from the collective dynamics of myosin motor proteins organized in sarcomeres, 2 μm-sized contractile units. Inside individual cardiomyocytes, dozens of sarcomeres in series form long myofibrils enabling rapid, strong and anisotropic cell-level contractions. While the molecular basis of sarcomere contraction is well studied, and the macroscopic motion of cardiac muscles is phenomenologically well described, far less is known about the contraction and expansion of individual sarcomeres and their competitive interactions inside myofibrils during cardiac beating. I developed a set of novel experimental and computational tools to study sarcomere dynamics in-vitro in hiPSC-derived cardiomyocytes. I cultured cardiomyocytes differentiated from a custom CRISPR-generated ACTN2-YFP cell line with endogenously labeled sarcomere z-bands on micropatterned polyacrylamide gels, and recorded high-speed movies of single beating cells. To automatically quantify the structure and function of sarcomeres, I developed SarcAsM (Sarcomere Analysis Multi-tool), an AI-based computational tool for the multi-parametric analysis of sarcomere structure and for the tracking of sarcomere dynamics. Using this high-throughput pipeline, I collected a large data set from 1,300 cells adherent to six different substrates, varying in stiffness, to study the effects of mechanical constraints on the dynamics and interactions of sarcomeres in myofibrils. I found that stiff substrates (>20 kPa) inhibited overall cellular contraction, but not motions of individual sarcomeres. Instead, sarcomeres were forced into a tug-of-war- like competition and moved heterogeneously, exhibiting rich dynamic phenomena such as rapid oscillatory motion and sarcomere popping. I could further show that the observed heterogeneity is in part stochastic, meaning that the type of motion of a sarcomere is not predetermined by static non-uniformities, but varies stochastically from beat to beat. To better understand the dynamic rules underlying these complex phenomena, I modeled the myofibril as a series of dynamic contractile elements with non-monotonous force-velocity relations, as predicted in theoretical models for the collective dynamics of motor proteins. Although it makes highly simplifying assumptions for the passive mechanics of sarcomeres, surprisingly, this model was able to reproduce various crucial aspects of our experimental data, such as the response to mechanical constraints, stochastic heterogeneity and sarcomere popping. This means that stochastic fluctuations at the sarcomere level are entirely sufficient to induce heterogeneous motion even in the absence of static non-uniformities between sarcomeres. Our model also gives insight into how additional static non-uniformities of sarcomeres are, to a certain extent, compensated by the stochastic fluctuations on the sarcomere level. These findings suggest that dynamic properties of cardiac muscle are not sarcomere-autonomous as widely assumed. Instead, they emerge due to the interplay of a heterogeneous ensemble of sarcomeres governed by (1) dynamic instabilities and (2) stochastic fluctuations, two previously overlooked aspects of sarcomere dynamics. The methodology I developed and the novel non-equilibrium active matter perspective on sarcomere dynamics in cardiac myofibrils open new opportunities to better understand cardiomuscular diseases and will aid the development of new therapies.
Keywords: sarcomeres; cardiomyocytes; myofibrils; biophysics; stochastic heterogeneity; cardiac muscle