Real-time in vivo imaging of stem cells following transgenesis by transposition

Tolar, Jakub; Osborn, Mark; Bell, Scott; McElmurry, Ron; Xia, Lily; Riddle, Megan; Panoskaltsis-Mortari, Angela; Jiang, Yuehua; McIvor, R Scott; Contag, Christopher H; Yant, Stephen R; Kay, Mark A; Verfaillie, Catherine; Blazar, Bruce R

doi:10.1016/j.ymthe.2005.02.023

Real-time in vivo imaging of stem cells following transgenesis by transposition

Author:

Tolar, Jakub

Osborn, Mark ; Bell, Scott ; McElmurry, Ron ; Xia, Lily ; Riddle, Megan ; Panoskaltsis-Mortari, Angela ; Jiang, Yuehua ; McIvor, R Scott ; Contag, Christopher H ; Yant, Stephen R ; Kay, Mark A ; Verfaillie, Catherine ; Blazar, Bruce R

Keywords:

Animals, Cell Nucleus, DNA Transposable Elements, Diagnostic Imaging, Flow Cytometry, Gene Transfer Techniques, Genes, Reporter, Luciferases, Luminescent Proteins, Mice, Mice, Inbred C57BL, Multipotent Stem Cells, Science & Technology, Life Sciences & Biomedicine, Biotechnology & Applied Microbiology, Genetics & Heredity, Medicine, Research & Experimental, Research & Experimental Medicine, multipotent adult progenitor cell, Sleeping Beauty, DsRed2, luciferase, stem cell homing, whole-body imaging, biophotonic imaging, ADULT PROGENITOR CELLS, LONG-TERM EXPRESSION, BONE-MARROW, SLEEPING-BEAUTY, T-CELLS, SYSTEM, TRANSPLANTATION, MOUSE, GENE, DIFFERENTIATION, 06 Biological Sciences, 10 Technology, 11 Medical and Health Sciences, Biotechnology, 3105 Genetics, 3202 Clinical sciences, 3206 Medical biotechnology

Abstract:

Previous studies have identified Sleeping Beauty transposons as efficient vectors for nonviral gene delivery in mammalian cells. However, studies demonstrating the usefulness of transposons as gene delivery vehicles into adult stem cells are lacking. Multipotent adult progenitor cells (MAPC) are nonhematopoietic stem cells with the capacity to form most, if not all, cell types of the body and as such hold great therapeutic potential. The whole-body biodistribution and persistence of MAPC are unknown, and such data would help direct clinical applications. We have nucleofected murine MAPC with two plasmid-based Sleeping Beauty transposons encoding the red fluorescent protein (DsRed2) and firefly luciferase. Transgenic euploid MAPC clones maintained their characteristic multilineage differentiation potential in vitro. DsRed2 and luciferase expression allowed for MAPC detection in vivo and in tissue sections. To confirm that transgenesis occurred by transposition into the genome of MAPC, we mapped Sleeping Beauty transposon integration sites in two MAPC clones using splinkerette PCR. This novel dual-reporter imaging approach based on the transgenesis of MAPC with Sleeping Beauty transposons sheds light on the homing patterns of MAPC and paves the way for quantification of MAPC engraftment in real time in vivo.