The effect of microgrooved culture substrates on calcium cycling of cardiac myocytes derived from human induced pluripotent stem cells

Abstract

Induced pluripotent stem cell-derived cardiomyocytes (iPSC-CM) have been widely proposed as in vitro models of myocardial physiology and disease. A significant obstacle, however, is their immature phenotype. We hypothesised that Ca2+ cycling of iPSC-CM is influenced by culture conditions and can be manipulated to obtain a more mature cellular behaviour. To test this hypothesis we seeded iPSC-CM onto fibronectin coated microgrooved polydimethylsiloxane (PDMS) scaffolds fabricated using photolithography, or onto unstructured PDMS membrane. After two weeks in culture, the structure and function of iPSC-CM were studied. PDMS microgrooved culture substrates brought about cellular alignment (p < 0.0001) and more organised sarcomere. The Ca2+ cycling properties of iPSC-CM cultured on these substrates were significantly altered with a shorter time to peak amplitude (p = 0.0002 at 1 Hz), and more organised sarcoplasmic reticulum (SR) Ca2+ release in response to caffeine (p < 0.0001), suggesting improved SR Ca2+ cycling. These changes were not associated with modifications in gene expression. Whilst structured tissue culture may make iPSC-CM more representative of adult myocardium, further construct development and characterisation is required to optimise iPSC-CM as a model of adult myocardium.

Surface Chemistry and Microtopography of Parylene C Films Control the Morphology and Microtubule Density of Cardiac Myocytes

“…This particular pattern was adopted based on previous experiments with different pattern layouts, because it induced the greatest cellular alignment and nuclear elongation. 1 We plasma oxidized both groups under the same conditions, thus creating identical microtopography in both cases with grooves *0.5 mm deep. Films from both groups were deposited on top of glass coverslips, thus we consider that the films have roughly the same stiffness.…”

“…Among different approaches, cell patterning induces the alignment and elongation of immature cardiac myocytes, improving many morphological and physiological properties of cardiac tissue, such as sarcomere organization, 1 Ca 2+ handling, [1][2][3] gap junction protein (connexin43) expression, 4 and directionality of action potential propagation. [5][6][7][8][9] Several different methodologies have been developed for cardiac tissue engineering using neonatal rat ventricular myocytes (NRVM) 2,[5][6][7][8][9] and induced pluripotent stem cellderived cardiomyocytes.…”

“…[5][6][7][8][9] Several different methodologies have been developed for cardiac tissue engineering using neonatal rat ventricular myocytes (NRVM) 2,[5][6][7][8][9] and induced pluripotent stem cellderived cardiomyocytes. 1 These techniques include microcontact printing of substrates with extracellular matrix (ECM) compounds, 5 deposition of ECM proteins and cells through microfluidic channels, 10 microgrooved substrates, 1,5,6,8,9 nanotextured hydrogels, 11 and lithographically patterned substrates. 2,7 Structured cardiomyocyte cultures have also been realized in three dimensions with the use of naturally or synthetically produced ECMs produced through electrospinning of aligned polymer nanofibers 12 and production of macroporous preformed scaffolds with complex biomimetic pore structures.…”

Capillary Force Lithography for Cardiac Tissue Engineering

“…Various other methods, such as microcontact printing, electrospinning, and microtopography, have previously been employed to control the structure of engineered cardiac tissue on the microscale [39][40][41] . While these techniques have proven successful in gaining cellular alignment, it is likely that the structure and function of in vivo cardiac tissue is governed by the much smaller, nanotopographical cues of the ECM and thus our NP substrate should prove advantageous.…”

Characterization of human iPS cell-derived cardiomyocyte sheets as a model to detect drug-induced conduction disturbance

“…It was reported that expression levels of genes of KCNH2, KCNE2 and KCNQ1 were higher in hiPSC-CMs than those in the human adult heart, whereas that of KCNE1 was similar between them (Uesugi et al, 2014). In addition, the extent of expression of a gene SLC8A1 encoding Na + / Ca 2+ exchanger 1 (NCX1) has been reported to be similar between adult heart tissue and structured hiPSC-CMs (Rao et al, 2013). Meanwhile, inward rectifier K + current (I K1 ) was shown to be hardly expressed in the hiP-SC-CMs (Ma et al, 2011;Doss et al, 2012;Meijer van Putten et al, 2015;Uesugi et al, 2014).…”