Cardiomyocyte Shape Control Improves Contractility Tests In Drug Screening Applications
Shen, Trong 1, 2 ; Kim, Gyu-tae 1 ; Shafieyan, Yousef 1 ; Hinz, Boris 1, 2
1. Laboratory of Tissue Repair and Regeneration, Faculty of Dentistry, University of Toronto; 2. Institute of Biomaterials and Biomedical Engineering, University of Toronto
Background: Pharmaceutical companies must perform pharmacology and toxicology pre-clinical trials to commercialize drugs. Over 90% of drugs that are effective in preclinical assessment later fail in clinical trials because of poor efficacy or adverse effects. Frequent causes of drug failure and/or market withdrawal are adverse cardiovascular effects. To address this gap, human induced stem-cell-derived cardiomyocytes (CMs) are used in cell-based high throughput drug screening (HTS). However, contraction force -a central CM function- is not measured in current HTS. We have developed a novel HTS device that measures changes in CM contractile force and frequency by quantifying visible deformations in silicone-based culture substrates. The device has been successfully benchmarked against existing technologies using a panel of drugs. However, the heterogeneity of stem-cell derived CM populations introduces data variability.
Objective/Hypothesis: We hypothesize that geometrically confining CMs on compliant silicone substrates into their physiological shape improves force production and beating frequency. By patterning the silicone surface of our contraction measurement device with arrays of adhesive islands with the footprint of CMs, we aim to standardize CM behavior for automated analysis in drug screens.
Methods: It is a major challenge to transfer adhesive proteins onto soft surfaces. To achieve patterns that determine CM shape on very soft recipient surfaces, we developed microcontact printing with fluorescently tagged proteins. Non-printed areas of the silicone substrate were passivated to prevent cell attachment, followed by seeding of CMs. To evaluate protein patterning success and effect on CM beating behavior, we used live imaging and fluorescence microscopy. Computational scripts were developed to automate data analysis.
Results: With improved microcontact printing, we achieved protein patterns of 120×20 µm rectangles on soft silicone substrates. CMs adhered and were geometrically confined to the island shape and exhibited visible contractility. Automated image data analysis was successfully implemented and demonstrated to greatly reduce analysis time.
Conclusion: The improved microcontact printing improved our CM-based HTS platform by achieving distinct CMs patterns without compromising CM contractility.
Significance/Impact: Controlling CM shape and position with protein patterning increases the potential of our HTS platform. We are currently analyzing whether CM cell shape confinement generates homogenous CM maturation states, regular beating, and increases CM contraction forces.