Article

Micro-positron emission tomography imaging of cardiac gene expression in rats using bicistronic adenoviral vector-mediated gene delivery.

Crump Institute for Molecular Imaging, UCLA School of Medicine, Los Angeles, Calif 94305-5427, USA.
Circulation (impact factor: 14.74). 04/2004; 109(11):1415-20. DOI:10.1161/01.CIR.0000121727.59564.5B pp.1415-20
Source: PubMed

ABSTRACT We have previously validated the use of micro-positron emission tomography (microPET) for monitoring the expression of a single PET reporter gene in rat myocardium. We now report the use of a bicistronic adenoviral vector (Ad-CMV-D2R80a-IRES-HSV1-sr39tk) for linking the expression of 2 PET reporter genes, a mutant rat dopamine type 2 receptor (D2R80a) and a mutant herpes simplex virus type 1 thymidine kinase (HSV1-sr39tk), with the aid of an internal ribosomal entry site (IRES).
Rat H9c2 cardiomyoblasts transduced with increasing titers of Ad-CMV-D2R80a-IRES-HSV1-sr39tk (0 to 2.5x10(8) pfu) were assayed 48 hours later for reporter protein activities, which were found to correlate well with viral titer (r2=0.96, P<0.001 for D2R80A; r2=0.98, P<0.001 for HSV1-sr39TK) and each other (r2=0.97; P<0.001). Experimental (n=8) and control (n=6) athymic rats underwent intramyocardial injection of up to 2x10(9) pfu of Ad-CMV-D2R80a-IRES-HSV1-sr39tk and saline, respectively. Forty-eight hours later and weekly thereafter, rats were assessed for D2R80a-dependent myocardial accumulation of 3-(2-[18F]fluoroethyl)spiperone ([18F]-FESP) and HSV1-sr39tk-dependent sequestration of 9-(4-[18F]fluoro-3-hydroxymethylbutyl)guanine ([18F]-FHBG) using microPET. Longitudinal [18F]-FESP and [18F]-FHBG imaging of experimental rats revealed a good correlation between the cardiac expressions of the 2 PET reporter genes (r2=0.73; P<0.001). The location of adenovirus-mediated transgene expression, as inferred from microPET images, was confirmed by ex vivo gamma counting of explanted heart.
The IRES-based bicistronic adenoviral vector can potentially be used in conjunction with PET for indirect imaging of therapeutic gene expression by replacing 1 of the 2 PET reporter genes with a therapeutic gene of choice.

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Keywords

2 PET reporter genes
 
adenovirus-mediated transgene expression
 
bicistronic adenoviral vector
 
D2R80a-dependent myocardial accumulation
 
ex vivo gamma
 
experimental rats
 
explanted heart
 
good correlation
 
HSV1-sr39tk-dependent sequestration
 
internal ribosomal entry site
 
IRES-based bicistronic adenoviral vector
 
Longitudinal [18F]-FESP
 
micro-positron emission tomography
 
microPET images
 
mutant rat dopamine type 2 receptor
 
Rat H9c2 cardiomyoblasts transduced
 
rat myocardium
 
reporter protein activities
 
single PET reporter gene
 
viral titer