In vitro analysis of SERCA2 gene regulation in hypertrophic cardiomyocytes and increasing transfection efficiency by gene-gun biolistics.
The transcriptional downregulation of the SERCA2 gene is studied using neonatal rat cardiomyocytes stimulated with endothelin-1 to induce hypertrophy. Liposome-based transfection of cells with a 1.9 kb SERCA2 promoter fragment directed expression of a reporter gene identical to the downregulation of genomic SERCA2 expression by endothelin-1. Results of a new gene gun technology for transient transfection of cardiomyocytes with a RSV-beta-galactosidase construct are reported. This new method for propelling DNA-coated gold beads into cardiomyocytes is extremely suitable for directly testing promoter/reporter gene DNA constructs since the transfection efficiency (approximately 10%) appears to be higher than traditional transfection methods.
Department of Biochemistry, Erasmus University Rotterdam, Netherlands.
K
K
Eizema
H A
HA
Van Heugten
K
K
Bezstarosti
M C
MC
Van Setten
J M
JM
Lamers
D000818:Animals; D019470:Biolistics / Q000592:standards*; D000252:Calcium-Transporting ATPases / Q000235:genetics*; D006332:Cardiomegaly / Q000235:genetics* / Q000473:pathology; D002478:Cells, Cultured; D019332:Endothelin-1 / Q000494:pharmacology; D015870:Gene Expression / Q000187:drug effects; D005786:Gene Expression Regulation / Q000502:physiology*; D009206:Myocardium / Q000166:cytology; D051381:Rats; D012519:Sarcoplasmic Reticulum / Q000201:enzymology*; D014158:Transcription, Genetic / Q000502:physiology; D014162:Transfection / Q000502:physiology*
D019332:Endothelin-1; D000252:Calcium-Transporting ATPases