Uptake of reporter probe 125I-FIAU in neonatal cardiac myocytes after adenoviral transfer of herpes simplex virus type I thymidine kinase reporter gene

Cardiac myocytes were obtained from neonatal rat heart and cultured. HSV1-tk, chosen as the reporter gene, was inserted into adenovirus vector (Ad5-tk), and the recombinant adenovirus infected neonatal cardiac myocytes with multiplicity of infection (MOI) of 20, 40 and 80. MTT assay was used to detect the viability of infected cardiac myocytes, and semi-quantitative reverse transcriptase polymerase chain reaction (RT-PCR) was applied to determine HSV1-tk mRNA expression in the infected ceils. The accumulation of 125I-FIAU into infected cardiac myocytes was detected as uptake rate. The survival rate of infected ceils had negative correlation with time, and it decreased with higher viral multiplicity of infection (MOI). After transferred for 24 h, the cardiac myocytes showed significant expression of HSV1-tk mRNA, while the control groups showed negative. Moreover, the exogenous gene expression in the infected cells at MOI 40 was significantly higher than in the ones at MOI 20 (P<0.01). The cardiac myocytes infected with increasing titer of Ad5-tk showed positive correlation between uptake rate and time, and the uptake rate of 125I-FIAU in the MOI 40 group was significantly higher than all the other groups (P<0.01). The HSV1-tk reporter gene could be transferred into the neonatal cardiac myocytes successfully with adenovirus vector, and encoded for HSV1-TK catalyzing phosphorylation so as to accumulate the marker substrates 125I-FIAU. The uptake rates, correlated to the uptake time, were the highest with transferred titer 40 at all the time points. This provides some cellular evidence for cardiac imaging of reporter gene in vivo. (authors)