University PhD Dissertation Defense
Microfabricated tools for functional assessment of developing cardiomyocytes
Rebecca Taylor
Advisors: Prof. Beth L. Pruitt and Prof. Ellen Kuhl
Department of Mechanical Engineering, Mechanics and Computation Division, Stanford University
Thursday, November 8th, 2012 at 8:00am (Refreshments at 7:45am)
Location: Mitchell Earth Sciences Building, Hartley Conference Room 130
Abstract:
Ischemic damage following myocardial infarction often leads to heart failure, contributing to cardiovascular disease's status as the number one killer in developed countries. This year an estimated 785,000 people in the United States alone will have their first heart attack. This underscores the critical need for cardiac therapies to actively repair damaged tissue. These therapies will be predicated upon knowledge of the mechanisms of cardiac growth and disease, including the development of contractile function and electrophysiological properties in maturing heart cells.
Two major barriers to this work involve the lack of tools for direct functional assessment for developing cardiomyocytes: (1) Axial force generation can not be assessed using current platforms and imaging techniques. (2) While cardiomyocyte phenotype and twitch power are improved when these cells are cultured on soft, tissue-like substrates in the 10-15 kPa range, standard in vitromicroelectrode arrays can not be used to study electrophysiology with cells cultured on soft, stretchable substrates. To address these issues, two different classes of device were microfabricated to perform direct functional assessment of developing cardiomyocytes.
A sacrificial layer technique was developed to suspend immature cardiomyocytes across pairs of widely-separated elastomer microposts. By prescribing a physiological cell shape and two-point loading, purely axial measurements of force generation were made during cardiomyocyte contraction. This force post technique achieved unmatched accuracy and precision, because microposts were directly calibrated using piezoresistive cantilevers of known stiffness, and in this measurement uncertainty was shown to be less than biological variation, a critical achievement for an elastomer technique. Forces of up to 146 nanoNewtons were measured. These forces were much smaller than the microNewton-scale forces reported from adult cardiomyocytes, suggesting that force generation capacity may increase with cardiomyocyte development. This technique provides a window into the development biology of healthy cardiomyocytes and a means to study the cardiac disease progression that was previously impossible.
In addition, to address the need for electrophysiological characterization of cardiomyocytes grown on soft, stretchable substrates, two different approaches were used to fabricate stretchable microelectrode arrays (SMEAs). A microfluidic platform filled with conductive ink and a flex circuit-based SMEA with a novel geometry were created. Both SMEAs maintain planarity and electrical properties throughout cyclic strains of up to 15%, and enable electrophysiological study of heart cells grown in biomimetic, soft and stretching environments.
Microfabrication has been used to develop devices for directly assessing cardiomyocyte function. These platforms overcome critical challenges to the handling, manipulation, and culture of immature cardiomyocytes, and are relevant for translational research as well as basic developmental and physiological investigations of stem cell-derived cardiomyocytes.
Rebecca Taylor
Bio-X and DARE Fellow
Graduate Research Assistant
Microsystems & Biomechanical Computation Groups
Mechanical Engineering
Stanford University
Bio-X and DARE Fellow
Graduate Research Assistant
Microsystems & Biomechanical Computation Groups
Mechanical Engineering
Stanford University
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