387971 Liquid Crystal Elastomers for Culture and Conditioning of Heart Muscle Sheets
In order to develop an effective and mature heart patch, cardiac cells need to be conditioned with pulsatile, unidirectional strain. However, current technologies for applying strain to cell sheet cultures require bulky equipment that limits the culture environment and limits scale up of the system. In this study, we developed a liquid crystal elastomer (LCE) with a thin coating of rigid polystyrene to generate aligned wrinkles for cell patterning and provide cyclic mechanical strain. To date, work has focused primarily on shape-memory polymers, which exhibit a one-way shape response. Here, we demonstrate that electrically conductive liquid crystal elastomer nanocomposites (LCE-NCs) exhibit rapid (response times as fast at 0.6 s), large-amplitude (contraction by up to 30 %), and fully reversible shape changes (stable to over 5000 cycles) under externally applied voltages (5 – 40 V). Cardiomyocytes, when cultured on these surfaces, undergo alignment and a hypertrophic response to generate cell sheets that could be used for cardiac regenerative therapies.
Materials and Methods
LCE-NCs are prepared using a two-step crosslinking method and introducing conductive carbon black nanoparticles both before and after crosslinking. In contrast to other LCE-NCs and shape-responsive materials studied, the electromechanical response is fully reversible (up to 5000 cycles tested) and operable in cell culture media with negligible heating of the surroundings. Depositing a 50nm polystyrene film on the LCE under tension, annealing, and releasing the tension resulted in controlled buckling of the surface, with aligned wrinkles with wavelengths from 1 to 20um, depending on the film thickness. LCEs were coated with 0.05 mg/ml rat tail collagen to enhance cell attachment. Neonatal rat ventricular myocytes (NRVM) were isolated from 3-day-old Sprague-Dawley rats using an isolation kit (Cellutron, Highland Park, NJ) and plated at 80,000 cells/cm2 in a high-serum media. After 3 days of culture, viability was assayed with a Live/Dead stain (Biotium, Hayward, CA) and cells were analyzed for alignment relative to the pattern direction using ImageJ (NIH, Bethesda, MD).
The resulting LCE bilayer composites exhibited rapid (1 Hz, with 0.6s response times) and reversible shape and topography changes in response to voltages of 5 – 40 V Substrate strain was 1-30% and correlated with pulse amplitude. Electromechanical expansion of conductive LCEs was reversible with no hysteresis. NRVM had >70% viability on LCE-carbon black bilayer substrates, and aligned parallel to the wrinkles at low wavelengths (<4.0um) or perpendicular at high wavelengths (>4.0um). Chronic stimulation of cardiomyocytes over 24 hours with cyclic strain resulted in highly aligned cell sheets with elongated myocytes exhibiting a hypertrophic response.
Discussion and Conclusions
These results demonstrate that conductive monodomain LCEs with a film of rigid polystyrene can be manufactured to give aligned wrinkles with controlled wavelength, and cardiomyocytes plated on these surfaces will align either parallel or perpendicular to the surface depending on the wavelength. These LCEs can deform by up to 30% and at 1Hz when a voltage is applied and should be useful in conditioning cardiomyocyte cell sheets.
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