Cardiomyocyte Shape Control Improves Contractility Tests in Drug Screening Applications
Shen. Trong 1 ; Kim, Gyu-Tae 2 ; Shafieyan, Yousef 2 ; Hinz, Boris 1, 2
1. Institute of Biomaterials and Biomedical Engineering, University of Toronto; 2. Faculty of Dentistry, University of Toronto
Background: Unexpected cardiovascular side-effects of drugs put patients at risk and require withdrawal from the market with billions of dollars associated costs. The drug development industry is in critical need for human cardiomyocyte (CM)-based preclinical assays with high throughput drug screening (HTS) capability. Current HTS tests cannot directly assess contraction force as a central CM function. We have developed a novel HTS device that is able to measure changes in CM contractile force and frequency by analyzing visible deformations (wrinkles) exerted by CMs to silicone-based culture substrates. The device has been successfully benchmarked against existing technologies using a panel of drugs. While our device was highly sensitive in detecting drug effects, cell population heterogeneity contributed to high variance of baseline CM beating behavior.
Objective: To standardize CM behavior for automated analysis in drug screens by patterning adhesion islands on the silicone surface of our contraction measurement device.
Hypothesis: Geometrically confining CMs into their ‘physiological’ shape on compliant silicone substrate improves force production and beating frequency by guiding CM sarcomere organization and avoiding cell aggregate formation.
Materials and Methods: To achieve CM shape patterning we developed microcontact printing to very soft recipient surfaces. A master wafer containing a negative of the desired patterns was created by photolithography and microfabrication techniques. Polydimethylsiloxane was poured and cured on the master to create a positive stamp. Soluble fluorescently tagged proteins were then used to ink the stamp and protein was transferred to the soft silicone substrate of our device by gentle pressure. 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. MATLAB-based computational analysis was developed to automate data analysis.
Results: Distinct protein and cell patterns were achieved on silicone substrates with improved microcontact printing. We identified protein patterns that supported homogenous CM beating with high amplitudes. Data analysis was successfully automated in MATLAB, which greatly reduced analysis time over existing procedures.
Significance/Impact: Cell shape patterning enhances our HTS-capable device by supporting homogenous beating and increasing force of CM contractions. Together with the heart-soft biomimetic culture surface, CM shape and position control further increases the efficacy of our device to evaluating cardiovascular side-effects in preclinical HTS tests.