An In Vitro and In Vivo Evaluation of a Reporter Gene/Probe System hERL/18F-FES

Abstract

Purpose: To evaluate the feasibility of a reporter gene/probe system, namely the human estrogen receptor ligand binding domain (hERL)/16α-[18F] fluoro-17β-estradiol (18F-FES), for monitoring gene and cell therapy. Methods: The recombinant adenovirus vector Ad5-hERL-IRES-VEGF (Ad-EIV), carrying a reporter gene (hERL) and a therapeutic gene (vascular endothelial growth factor, VEGF165) through an internal ribosome entry site (IRES), was constructed. After transfection of Ad-EIV into bone marrow mesenchymal stem cells (Ad-EIV-MSCs), hERL and VEGF165 mRNA and protein expressions were identified using Real-Time qRT-PCR and immunofluorescence. The uptake of 18F-FES was measured in both Ad-EIV-MSCs and nontransfected MSCs after different incubation time. Micro-PET/CT images were obtained at 1 day after injection of Ad-EIV-MSCs into the left foreleg of the rat. The right foreleg was injected with nontransfected MSCs, which served as self-control. Results: After transfection with Ad-EIV, the mRNA and protein expression of hERL and VEGF165 were successfully detected in MSCs, and correlated well with each other (R2 = 0.9840, P<0.05). This indicated the reporter gene could reflect the therapeutic gene indirectly. Ad-EIV-MSCs uptake of 18F-FES increased with incubation time with a peak value of 9.13%±0.33% at 150 min, which was significantly higher than that of the control group. A far higher level of radioactivity could be seen in the left foreleg on the micro-PET/CT image than in the opposite foreleg. Conclusion: These preliminary in vitro and in vivo studies confirmed that hERL/18F-FES might be used as a novel reporter gene/probe system for monitoring gene and cell therapy. This imaging platform may have broad applications for basic research and clinical studies. © 2013 Qin et al.