Skip Navigation

Cardiovascular Research 2003 59(4):874-882; doi:10.1016/S0008-6363(03)00508-X
© 2003 by European Society of Cardiology
This Article
Right arrow Full Text Freely available
Right arrow FREE Full Text (PDF) Freely available
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Add to My Personal Archive
Right arrow Download to citation manager
Right arrowRequest Permissions
Google Scholar
Right arrow Articles by Weikert, C.
Right arrow Articles by Eppenberger, H. M
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Weikert, C.
Right arrow Articles by Eppenberger, H. M
Social Bookmarking
Add to CiteULike   Add to Connotea   Add to Del.icio.us  
What's this?

Copyright © 2003, European Society of Cardiology

Cellular engineering of ventricular adult rat cardiomyocytes

Christian Weikert, Monika Eppenberger-Eberhardt and Hans M Eppenberger*

Institute of Cell Biology, ETH-Hoenggerberg Swiss Federal Institute of Technology, CH-8093 Zurich, Switzerland

hme{at}cell.biol.ethz.ch

* Corresponding author. Tel.: +41-1-633-3357; fax: +41-1-633-1152.

Objective: Preparation of viable cultured adult cardiomyocytes (vARCs) is a prerequisite for cell-based transplantation and tissue engineering. Ectopic gene expression is important in this context. Here, we present an in vitro cell replating strategy using AccutaseTM for cultured vARCs, allowing ectopic gene expression. Methods: Cultured vARCs from 6- to 8-week-old rats were used. Transfections with EGFP (enhanced green fluorescent protein) constructs, Mlc-3f-EGFP or {alpha}-actinin-EGFP were performed using adenovirus-enhanced transferrin-mediated infection (AVET). AccutaseTM (PAA Laboratories, Linz, Austria) was used for the detachment of cultured cells. Immunohistochemical analysis, together with confocal laser microscopy was used for structural analysis of the cells. Results: Cultured vARCs could be detached with a high yield (40 to 60%) from primary cultures using AccutaseTM. The cultivation period plays an important role in the yield of viable cells. Resultant replated vARCs (rep-vARCs) rapidly (1–2 h) acquired a rounded up shape without degradation of their contractile apparatus, which is in contrast to the rod-shaped freshly isolated vARCs (fi-vARCs). The detached cells survived passage through a narrow syringe needle. After seeding, detached cells rapidly attached to various substrates, increased their content of the contractile apparatus, and formed cell–cell contacts within 3 days after reseeding. The detached cells survived passage through a narrow syringe needle. The high recovery of cells after replating enabled the use of the AVET system for gene delivery. AVET is free of infectious particles and does not lead to expression of viral proteins. Transfection of vARCs prior to detachment had a small effect on cell recovery and ectopically synthesized proteins were properly localized after replating. Conclusions: Detachment of cultured vARCs using AccutaseTM is well compatible with ectopic gene expression and yields a viable transgenic population of vARCs that eventually may be suitable as transgenic cardiomyocyte grafts.

KEYWORDS Experimental; Heart; Cellular; Cell culture; Transplantation; Contractile function; Gene therapy; Hypertrophy


Add to CiteULike CiteULike   Add to Connotea Connotea   Add to Del.icio.us Del.icio.us    What's this?




Disclaimer:
Please note that abstracts for content published before 1996 were created through digital scanning and may therefore not exactly replicate the text of the original print issues. All efforts have been made to ensure accuracy, but the Publisher will not be held responsible for any remaining inaccuracies. If you require any further clarification, please contact our Customer Services Department.