Application of double-tagging in monitoring of tumorigenesis and tracking of mesenchymal stem cells in mice

Abstract

To establish murine liver cancer cell line (Hepa1-6) and human mesenchymal stem cells (hMSCs) with dual-modality imaging lentiviral vector transfection and monitor the development of liver tumors and dynamically tracking of hMSCs by bioluminescent imaging in vivo. The cell lines of Hepa 1-6 and hMSCs were constructed abd tranfected with RL-eGFP and FL2-mKate2 lentiviruses respectively. Then the eGFP(+)-Hepa1-6 and mKate2(+)-hMSCs cell lines were selected by fluorescence activated cell sorting (FACS). The bioluminescent imaging technology was employed to evaluate their bioluminescent abilities in vitro and in vivo. And the capacity of mKate2(+)-hMSCs migrating to HCC cell line (eGFP(+)-Hepa1-6) was evaluated by Transwell migration assay. DNA sequencing analysis confirmed that gene sequences of lentiviral vectors were correct without mutation. We successfully constructed the lentivirus vector, produced lentivirus and infected Hepa1-6 and hMSCs respectively. Subsequently eGFP(+)-Hepa1-6 cells and mKate2(+)-hMSCs were obtained by FACS. The expressions of fluorescent protein eGFP and mKate were confirmed by fluorescence microscope. By bioluminescent imaging system, bioluminescence intensity strongly correlated with cell number. The Transwell assay showed that hMSCs migrated toward heptocellular carcinoma (HCC) cells. A lentivirus vector has been successfully constructed for infecting Hepa1-6 and hMSCs and analyzed by fluorescence and bioluminescent imaging. And that makes it technically feasible to further monitor the interaction of hMSCs and liver tumor cells. These results provide rationales for further studying the mechanism of MSCs in the development and progression of HCC.