Cardiac Organogenesisin Vitro: Reestablishment of Three-Dimensional Tissue Architecture by Dissociated Neonatal Rat Ventricular Cells

Abstract

The mammalian heart does not regenerate in vivo. The heart is, therefore, an excellent candidate for tissue engineering approaches and for the use of biosynthetic devices in the replacement or augmentation of defective tissue. Unfortunately, little is known about the capacity of isolated heart cells to re-establish tissue architectures in vitro. In this study, we examined the possibility that cardiac cells possess a latent organizational potential that is unrealized within the mechanically active tissue but that can be accessed in quiescent environments in culture. In the series of experiments presented here, total cell populations were isolated from neonatal rat ventricles and recombined in rotating bioreactors containing a serum-free medium and surfaces for cell attachment. The extent to which tissue-like structure and contractile function were established was assessed using a combination of morphological, physiological, and biochemical techniques. We found that mixed populations of ventricular cells formed extensive three-dimensional aggregates that were spontaneously and rhythmically contractile and that large aggregates of structurally-organized cells contracted in unison. The cells were differentially distributed in these aggregates and formed architectures that were indistinguishable from those of intact tissue. These architectures arose in the absence of three-dimensional cues from the matrix, and the formation of organotypic structures was apparently driven by the cells themselves. Our observations suggest that cardiac cells possess an innate capacity to re-establish complex, three-dimensional, cardiac organization in vitro. Understanding the basis of this capacity, and harnessing the organizational potential of heart cells, will be critical in the development of tissue homologues for use in basic research and in the engineering of biosynthetic implants for the treatment of cardiac disease.

Cardiac tissue engineering using perfusion bioreactor systems

“…Three-dimensional cardiac tissue constructs that express structural and physiological features characteristic of native cardiac muscle have been engineered in vitro using fetal or neonatal rat cardiac myocytes (CMs) on collagen fibers 2 , fibrous polyglycolic acid scaffolds [3][4][5][6][7] and porous collagen scaffolds [8][9][10] .…”

“…In early studies, cells were seeded onto scaffolds and cultivated in dishes 4,7,8,11 , in spinner flasks 3,4,7 or in rotating vessels 2,4,7 . Inallof these systems, oxygen dissolved in culture medium was transported to the cells by molecular diffusion.…”

“…All calculations are based on the scaffold volume experiencing the interstitial flow (i.e., based on the inner diameter of the silicone gasket) and the total surface area of the pores in that volume. The average fluid velocity, U, though each pore can be determined according to the following equation: (2) where Q is volumetric flow rate through the cardiac patch, V is the scaffold volume and ε is the void fraction. The following equation, based on Poiseuille flow, can be used to estimate the shear stress:…”

Endothelial Cells Promote Cardiac Myocyte Survival and Spatial Reorganization

“…11 Cell-seeded grafts have been used instead of isolated cells for in vitro cardiac tissue growth or in vivo transplantation. [12][13][14][15][16][17][18] These grafts can develop a high degree of myocyte spatial organization, differentiation, and spontaneous and coordinated contractions. On implantation in vivo, cardiac grafts can integrate into the host tissue and neovascularization can develop.…”

Formation of three‐dimensional fetal myocardial tissue cultures from rat for long‐term cultivation

“…Three-dimensional cardiomyocyte cultures in combination with recent molecular biology techniques may offer new possibilities for the examination of the growth potential, analysis of time-specific gene expression, and cellular arrangement in an organotypical environment (Eschenhagen et al, 1997;Akins et al, 1999;Zimmermann et al, 2002;Baar et al, 2005). Numerous different tissue-like culture models are established by using various culture techniques, biological and nonbiological scaffolds, culture media, and cell sources (Zimmermann et al, 2000;Shimizu et al, 2003;Shin et al, 2003;Kelm et al, 2004;Kofidis et al, 2004).…”