2D and 3D-Organized Cardiac Cells Shows Differences in Cellular Morphology, Adhesion Junctions, Presence of Myofibrils and Protein Expression

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.

Osteocyte differentiation and the formation of an interconnected cellular network in vitro

“…The MC3T3-E1 osteoblast cell line represents a uniformly defined cell population, which have similar traits to primary cells and can be readily expanded (Czekanska et al, 2012) and differentiated (Chatterjee et al, 2010;Keogh et al, 2010;Krishnan et al, 2010;Mullen et al, 2013;Partap et al, 2010;Przybylowski et al, 2012;St-Pierre et al, 2005;Uchihashi et al, 2013), making the cell line a good representative of pre-osteoblasts (Grigoriadis et al, 1985;Quarles et al, 1992;Sudo et al, 1983;Wang et al, 1999). It should be noted that ALP activity was only assessed from extracellular ALP in the media at specific time-points and thus might not be a reflection of the total ALP activity.…”

“…In vivo, osteoblasts and osteocytes primarily exist within a complex three dimensional (3D) environment (Boukhechba and Balaguer, 2009), and it is known that 3D environment has a significant effect on cell morphology and geometry, as shown in NIH 3T3 fibroblast (Legant et al, 2010), cardiac cells (Soares et al, 2012) and MC3T3-E1 osteoblasts (Murshid et al, 2007). The process of osteocyte dendrite formation within a 3D environment is highly dynamic, as the embedding cells repeatedly extend and retract their dendrites.…”

“…as, cardiac cells, human embryonic stem cells and HepG2 liver cells (Lan et al, 2010;Soares et al, 2012;Tian et al, 2008).…”

Model Systems for Cardiovascular Regenerative Biology

“…Cell phenotype can be dramatically affected by the culture geometry; for example, chick CMs grown in 3D cultures have more mitochondria, display more cell -cell junctions, show more spontaneous contractions, and express a greater abundance of proteins found in mature CMs (e.g., desmin, a-actin, and cadherin) when compared with CMs grown in 2D cultures (Soares et al 2012).…”

A Non-woven Path: Electrospun Poly(lactic acid) Scaffolds for Kidney Tissue Engineering

“…It is well documented that a 3D structure affects many different cell types [17][18][19][20] and could quite possibly be a crucial element of kidney tissue engineering. Decellularised tissue is currently the predominant vehicle for 3D kidney cell culture with impressive advances made [21][22][23][24].…”