Adenovirus-mediated gene transfer to adult mouse cardiomyocytes is selectively influenced by culture medium.
BACKGROUND
As development of cardiac gene therapies progresses, virally mediated genetic manipulations in cultured cardiomyocytes has become an important experimental approach. While adenovirus (Ad)-mediated gene transfer to neonatal and adult rat cardiomyocytes is well established, viral transduction of cultured adult mouse cardiomyocytes (AMCM) has been more difficult. This study was designed to test the hypothesis that culture medium is a critical determinant of efficient gene transfer in AMCM.


METHODS
AMCM from 8-week-old C57BL/6 mice were cultured in either minimum essential medium (MEM) or medium M199 and then infected with an Ad beta-galactosidase and transduction efficiency was quantified by cytochemistry and beta-galactosidase activity assay. Coxsackie-adenovirus receptor (CAR) levels and Ad binding were evaluated by immunocytochemistry in M199- vs. MEM-cultured AMCM.


RESULTS
Our results demonstrated dramatic differences in efficiency of Ad-mediated gene transfer in AMCM cultured in MEM (90 +/- 8%) vs. M199 (5 +/- 1.2%). This difference was specific to AMCM, and was not observed in a number of other cells including neonatal rat cardiomyocytes. The enhanced transduction in MEM was associated with increased levels of CAR and Ad binding in AMCM.


CONCLUSIONS
Culture medium has a profound effect on the efficiency of Ad-mediated gene transfer in AMCM, perhaps via differential effects on CAR expression. These findings have important implications for increasing numbers of studies that employ viral gene transfer in adult cardiomyocytes derived from mouse models of cardiac diseases.
Li
Zhenbo
Z
Department of Anatomy and Cell Biology, Roy J. and Lucille A. Carver College of Medicine, The University of Iowa, Iowa City 52242, USA.
Sharma
Ram V
RV
Duan
Dongsheng
D
Davisson
Robin L
RL
D000256:Adenoviridae / Q000235:genetics* / Q000378:metabolism; D000818:Animals; D062727:Coxsackie and Adenovirus Receptor-Like Membrane Protein; D003470:Culture Media; D005822:Genetic Vectors; D051379:Mice; D032383:Myocytes, Cardiac / Q000737:chemistry / Q000378:metabolism*; D011991:Receptors, Virus / Q000235:genetics / Q000378:metabolism; D014161:Transduction, Genetic / Q000379:methods*
C568772:CLMP protein, mouse; C568773:Clmp protein, rat; D062727:Coxsackie and Adenovirus Receptor-Like Membrane Protein; D003470:Culture Media; D011991:Receptors, Virus