Objective: Radionuclide imaging of reporter gene expression holds promise for noninvasive monitoring of gene therapy. Herpes simplex virus 1-thymidine kinase (HSV1-tk) has been successfully applied to the tumor tissue.We explored the feasibility of the expression imaging of HSV1-tk reporter gene in rat myocardium by using SPECT reporter probe (131)I-2'-fluoro-2'-deoxy-1-beta-D-arabinofuranosyl-5-iodouracil ((131)I-FIAU) and autoradiography (ARG).
Methods: The recombinant Ad5-tk carrying HSV1-tk gene and adenovirus (Ad5-null) as vector were constructed and intramyocardially injected into SD rats. Experiment was grouped for different aims as follows: (1) Influence of time on the imaging after transfection reporter gene: rats were injected with (131)I-FIAU at day 1, 2, 3, 5 and 7 after transfection of 1 x 10(8)pfu Ad5-tk; (2) Influence of various titers on the imaging: rats underwent intramyocardial injection with various titers of Ad5-tk (5 x 10(8), 1 x 10(8), 5 x 10(7), 1 x 10(7)pfu). After 2 days, rats were injected with (131)I-FIAU in tail vein. Equal volume Ad-nulls was intramyocardially injected to control rats. Rats were killed 24 h after injection of (131)I-FIAU and the hearts were rapidly dissected for gamma counts measurement. The total myocardial (131)I-FIAU accumulation was quantified in percent of injected dose per gram myocardium (%ID/g). The myocardial reporter gene expression was semi-quantitatively determined by ARG and RT-PCR.
Results: ARG and RT-PCR showed that the local expression of reporter gene increased in proportion with increasing titer and decreased in proportion with time post injection. The semi-quantitative assay showed there were significant correlations among %ID/g, RT-PCR and ARG: r(2) = 0.963, P < 0.05 for RT-PCR and ARG; r(2) = 0.996, P < 0.01 for %ID/g and ARG in rats received various reporter gene titers at identical time point post injection; r(2) = 0.950, P < 0.05 for RT-PCR and ARG; r(2) = 0.980, P < 0.01 for %ID/g and ARG for rats received identical reporter gene titer on various time points post injection.
Conclusions: The present study showed that cardiac reporter gene imaging with HSV1-tk as reporter gene and (131)I-FIAU as reporter probe was feasible in rats. The optimal Ad5-tk titer is 1 x 10(8) pfu and the optimal imaging time is 24 h to 48 h post gene transfer. HSV1-tk/FIAU may be used for the noninvasive monitoring of cardiac gene therapy.