Abstract
Cardiac cells are organized in vivo in a complex tridimensional structural organization that is crucial for heart function. While in vitro studies can reveal details about cardiac cell biology, usually cells are grown on simplified two-dimensional (2D) environments. To address these differences, we established a cardiac cell culture composed of both 2D and three-dimensional (3D)-organized cells. Our results shows significant differences between the two culture contexts in relation to the overall morphology of the cells, contraction ability, proliferation rate, presence of intercellular adhesion structures, organization of myofibrils, mitochondria morphology, endoplasmic reticulum contents, cytoskeletal filaments and extracellular matrix distribution, and expression of markers of cardiac differentiation. Cardiac cells grown in 2D-context displayed a flattened and well spread shape, were mostly isolated and their cytoplasm was filled with a large network of microfilaments and microtubules. In contrast, 3D-cells were smaller in size, were always in close contact with each other with several cellular junctions, and displayed a less conspicuous cytoskeletal network. 3D-cells had more mitochondria and myofibrils and these cells contract spontaneously more often than 2D-cells. On the other hand, endoplasmic reticulum membranes were present in higher amounts in 2D-cells when compared to 3D-cells. The expression of desmin, cadherin and alpha-actinin was higher in 3D-aggregates compared to 2D-spread cells. These findings indicate that the tridimensional environment in which the cardiac cells are grown influence several aspects of cardiac differentiation, including cell adhesion, cell shape, myofibril assembly, mitochondria contents and protein expression. We suggest that the use of this cardiac culture model, with 2D and 3D-context cells, could be useful for studies on the effects of different drugs, or growth factors, giving valuable information on the biological response of cells grown in different spatial organizations.
Highlights
The heart is composed of contractile muscle cells and non-muscle cell types including fibroblasts and blood vessel cells
These cell types are organized in a complex tridimensional structural organization that is crucial for cardiac function and depends on autocrine, paracrine, and cell-cell interactions
Any attempt to study cardiac cells must consider that a complex and intricate system of myofibrils connected to intercellular adhesion structures at the subsarcolemmal region of each cardiomyocyte is indispensable for cardiac function
Summary
The heart is composed of contractile muscle cells and non-muscle cell types including fibroblasts and blood vessel cells. In this culture system we were able to find significant differences between 2D- and 3D-cells in several parameters, including cell morphology, contraction ability, presence of adhesion structures, organization of myofibrils, mitochondria shape and contents, cytoskeletal filaments and extracellular matrix distribution and expression of proteins that are markers of cardiac differentiation. We decided to study and compare directly 2D- and 3D-cardiac cells cultured together in many aspects, including their overall morphology, sub-cellular organization, ability to contract and the expression of markers of cardiac differentiation.
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