Diffraction phase microscopy applied to characterize cardiomyocyte morphology and motion

Cardiomyocytes which are derived from human induced pluripotent stem cells (hiPSC-CMs) have broad applications such as drug testing, personalized medicine, and eventually repairing damaged heart tissue. However, the quality of these cells needs to be determined for such applications, and hiPSC-CMs are known to be highly heterogenous and have not matured to the level of human adult primary cardiomyocytes. Methods to analyze these cells of often either destructive, invasive, or require a fluorescent label that alters the cells performance. In this thesis, I describe a non-invasive image-based method for analyzing and classifying cardiomyocytes based on their morphology and contractile properties using diffraction phase microscopy (DPM). DPM yields optical thickness measurements which enables analysis of cell morphology and beating characteristics. DPM is a single-shot, low noise implementation of quantitative phase microscopy, and is ideal for this application of measuring fast dynamics of contracting cardiomyocytes with excellent sensitivity. In this thesis, the optimization of the microscope is described for analyzing cardiomyocytes, and the system noise / sensitivity is characterized. The system is capable of measuring changes of only a few milliradians at speeds over 100 frames per second (fps), which is fast and sensitive enough to capture beating of single hiPSC-CMs. Then hiPSC-CMs are measured and analyzed using the optimized DPM system for their beating characteristics such as frequency, regularity, and timing of contraction and relaxation. These methods can be applied for high-throughput, non-invasive drug testing and classification of hiPSC-CMs. Because the action potential drives the cellular motion, these methods have the potential to replace the traditional electrophysiological characterization, such as slow and tedious patch clamping. Population-level statistics on beating characteristics as well as for morphological characteristics such as the area, shape and optical volume of of hiPSC-CMs show vast heterogeneity and continuous distributions of all parameters analyzed. These results highlight the need for more robust classification and quality control of these cells in order to get reliable results in drug testing and research settings, and in order to generate safe and effective cells for regenerative medicine applications such as repairing damaged heart tissue. Finally, DPM and these analysis methods are applied to compare populations of hiPSC-CMs with controlled (patterned) and uncontrolled (unpatterned) shape. The key findings obtained by applying these methods were: 1) patterning effectively controls the shape of the cells, while cells with the desired mature shape rarely appear in the unpatterned population, 2) patterned cells are more likely to beat with consistent and lower beat frequency compared to unpatterned cells, and 3) the patterns tend to select for larger (more mature) cells. Finally, there appears to be a cutoff point under which cells of a certain volume or dry mass don't attach to the patterns. These results indicate that controlling the shape of hiPSC-CMs improves their quality and has the potential to yield more consistent research results and homogenous populations of cells for clinical applications.

Keywords: Electrical Engineering.
Advisor: Solgaard Olav 
Abilez Oscar
Bowden Audrey