Specific expression of reporter gene FTH1 driven by progression elevated gene-3 promoter in malignant transformation of rat mesenchymal stem cells and its detection by in vitro magnetic resonance imaging

Abstract

Objective To explore the feasibility of using progression elevated gene-3 (PEG3) promoter, a tumor-specific promoter, to drive the specific expression of the reporter gene ferritin heavy chain 1 (FTH1) and the detection of FTH1 expression using in vitro magnetic resonance imaging (MRI) in bone marrow mesenchymal stem cells (MSCs) after malignant transformation. Methods A viral vector carrying the reporter gene FTH1 driven by PEG3 promoter was transfected into rat MSCs to obtain MSCs-PEG3-FTH1 cells, and the MSCs transfected with a viral vector carrying FTH1 driven by the CMV promoter (MSCs-CMV-FTH1 cells) and the MSCs without transfection served as the controls. The non-transfected MSCs, MSCs-PEG3-FTH1 cells, and MSCs-CMV-FTH1 cells were indirectly co-cultured with rat glioma C6 cells for 2 weeks to induce their malignant transformation (TMSCs, TMSCs-PEG3-FTH1 and TMSC-PEG3-FTH1 cells, respectively). The morphology, migration and proliferation, and the cell surface antigens CD29, CD34, CD45 and CD90 of the transformed cells were observed and analyzed, and their capacity of subcutaneous tumor formation was evaluated in nude mice. Western blotting, Prussian blue staining, transmission electron microscopy, and in vitro MRI were used to detect the changes in the expression of FTH1 gene in the cells after the transformation. Results Compared with MSCs, most of TMSCs were small in size with a spindle-like morphology and showed strong proliferation and migration abilities with increased expression of CD34 and CD45, decreased expression of CD90 (P < 0.05), and an ability of subcutaneous tumor formation in nude mice. Western blotting showed that FTH1 protein was not expressed or expressed at low levels in MSCs, MSCs-PEG3-FTH1 cells and TMSCs, while high FTH1 expression was detected in MSCs-CMV-FTH1, TMSCs-PEG3-FTH1 and TMSCs-CMV-FTH1 cells(P < 0.05). Prussian blue staining and transmission electron microscopy identified the presence of iron particles in MSCs-CMV-FTH1, TMSCs-PEG3-FTH1 and TMSCs-CMV-FTH1 cells, but not in MSCs, MSCs-PEG3-FTH1 cells or TMSCs. In vitro MRI showed obviously decreased T2 signal in TMSCs-PEG3-FTH1 cells as compared with MSCs-PEG3-FTH1 cells. Conclusion PEG3 promoter can drive the specific expression of FTH1 that allows MRI detection in rat MSCs after malignant transformation. cell transformation neoplastic

Keywords

• progression elevated gene-3 promoter
• reporter gene ferritin heavy chain 1
• magnetic resonance imaging
• mesenchymal stem cells
• cell transformation neoplastic