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Formation of three‐dimensional fetal myocardial tissue cultures from rat for long‐term cultivation
Author(s) -
Just Lothar,
Kürsten Anne,
BorthBruhns Thomas,
Lindenmaier Werner,
Rohde Manfred,
Dittmar Kurt,
Bader Augustinus
Publication year - 2006
Publication title -
developmental dynamics
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.634
H-Index - 141
eISSN - 1097-0177
pISSN - 1058-8388
DOI - 10.1002/dvdy.20871
Subject(s) - biology , microbiology and biotechnology , cell , myocyte , embryonic stem cell , cell culture , anatomy , biochemistry , gene , genetics
Three‐dimensional cardiomyocyte cultures offer new possibilities for the analysis of cardiac cell differentiation, spatial cellular arrangement, and time‐specific gene expression in a tissue‐like environment. We present a new method for generating homogenous and robust cardiomyocyte tissue cultures with good long‐term viability. Ventricular heart cells prepared from fetal rats at embryonic day 13 were cultured in a scaffold‐free two‐step process. To optimize the cell culture model, several digestion protocols and culture conditions were tested. After digestion of fetal cardiac ventricles, the resultant cell suspension of isolated cardiocytes was shaken to initialize cell aggregate formation. In the second step, these three‐dimensional cell aggregates were transferred onto a microporous membrane to allow further microstructure formation. Autonomously beating cultures possessed more than 25 cell layers and a homogenous distribution of cardiomyocytes without central necrosis after 8 weeks in vitro. The cardiomyocytes showed contractile elements, desmosomes, and gap junctions analyzed by immunohistochemistry and electron microscopy. The beat frequency could be modulated by adrenergic agonist and antagonist. Adenoviral green fluorescent protein transfer into cardiomyocytes was possible and highly effective. This three‐dimensional tissue model proved to be useful for studying cell–cell interactions and cell differentiation processes in a three‐dimensional cell arrangement. Developmental Dynamics 235:2200–2209, 2006. © 2006 Wiley‐Liss, Inc.