RESUMO
Embryos devoid of autonomic innervation suffer sudden cardiac death. However, whether autonomic neurons have a role in heart development is poorly understood. To investigate if sympathetic neurons impact cardiomyocyte maturation, we co-cultured phenotypically immature cardiomyocytes derived from human induced pluripotent stem cells with mouse sympathetic ganglion neurons. We found that 1) multiple cardiac structure and ion channel genes related to cardiomyocyte maturation were up-regulated when co-cultured with sympathetic neurons; 2) sarcomere organization and connexin-43 gap junctions increased; 3) calcium imaging showed greater transient amplitudes. However, sarcomere spacing, relaxation time, and level of sarcoplasmic reticulum calcium did not show matured phenotypes. We further found that addition of endothelial and epicardial support cells did not enhance maturation to a greater extent beyond sympathetic neurons, while administration of isoproterenol alone was insufficient to induce changes in gene expression. These results demonstrate that sympathetic neurons have a significant and complex role in regulating cardiomyocyte development.
RESUMO
Understanding the role of mechanophenotype in competitive adherence of cells to other cells versus underlying substrates can inform such processes as tissue development, cancer progression, and wound healing. This study investigated how mechanophenotype, defined by whole-cell, elastic/viscoelastic properties for the perinuclear region, and cellular assembly are intertwined through the mechanosensing process. Atomic force microscopy was used to characterize the temporal elastic/viscoelastic properties of individual and assembled fibroblasts grown on substrates with elastic moduli above, below, or similar to whole-cell mechanophenotypes measured for three, genetically modified cell lines. All cells were at their most compliant immediately after plating but transitioned to distinct, stiffer mechanophenotypes by Day 1 after acclimation. This mechanical state, and cellular assembly/morphology, did not change significantly over the following three days of testing, regardless of substrate compliance or cellular organization (multi-cell nodules/plaques or single cells). Interestingly, cells formed 3D nodules when attached to substrates with elastic moduli less than their own but spread readily on substrates with moduli equal to or greater than their own, suggesting a preference to adhere to the stiffest surface sensed (substrate or cell). This suggests that inherent mechanophenotype plays a role as a competing surface during microenvironment mechanosensing and subsequent cell-cell-substrate organization.
