Two Novel Reporter Gene HERL And Tyrosinase Molecular Imaging Research

Reporter gene is an important tool for basic research in molecular imaging, which can be used to monitor gene expression in vivo non-invasively. Many reporter genes have been reported, but none of them has good clinical value. Therefore, looking for new reporter genes with great potential applications is very important. In this thesis, two new human reporter genes have been studied.The first part studied the human estrogen receptor ligand binding domain (hERL). hERL/18F-FES was used as a reporter gene system, bone marrow mesenchymal stem cells (MSCs) and vascular endothelial growth factor (VEGF) gene were the therapeutic cells and gene. MSCs were transfected with recombinant vector containing hERL and VEGF gene. In vitro, ER and VEGF expressions were detected, and cellular uptakes of18F-FES and125I-E2were also studied.In vivo, rat muscle model and myocardial infarction (MI) models were prepared, transfected cells were applied to treat MI,18F-FES PET imaging was the monitoring method. We assessed the feasibility of using hERL/18F-FES reporter gene system for monitoring stem cell/gene therapy of ischemic heart disease from both in vitro and in vivo. A new treatment for ischemic heart disease with clinical value and a safety and non-invasive monitoring method of treatment were found.The second part studied human tyrosinase (TYR). TYR was used as a single reporter gene; its final product melanin was the target for photoacoustic/MRI/PET tri-modality molecular imaging. Breast cancer MCF-7cells were transfected with a plasmid encoding TYR and stable expressing cell line was selected. In vitro, tyrosinase and melanin expressions were detected, and cell imaging with three modalities was studied. In vivo, nude mice tumor models were prepared and three imaging modalities were performed non-invasively. We proved its feasibility from both in vitro and in vivo. This is a new method of reporter gene multi-modality imaging. Compared with traditional ones, this strategy has several advantages, such as easy construction of vector, single product without interference of other proteins, signal amplification in imaging using the product melanin, no corresponding substrate or probe required for MRI and photoacoustic imaging, no immune response since it’s a human-derived gene. These three imaging modalities complemented with each other, PET and photoacoustic imaging has high sensitivity, MRI is compensating for the shortcomings of low spatial resolution, while producing a high T1signal. In short, this is the first time tyrosinase was reported as a single reporter gene for photoacoustic/MRI/PET multi-modality imaging.The good results of these two novel reporter genes in this thesis will play an important influence on the development of novel reporter genes, and may be converted to clinical in the future, so it worth further studies.