Real-time in vivo imaging of stem cells following transgenesis by transposition.
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.
Tolar
Jakub
J
Department of Pediatrics, Division of Hematology, Oncology, Blood and Marrow Transplant Program, University of Minnesota, Minneapolis, MN 55455, USA. tolar003@umn.edu
Osborn
Mark
M
Bell
Scott
S
McElmurry
Ron
R
Xia
Lily
L
Riddle
Megan
M
Panoskaltsis-Mortari
Angela
A
Jiang
Yuehua
Y
McIvor
R Scott
RS
Contag
Christopher H
CH
Yant
Stephen R
SR
Kay
Mark A
MA
Verfaillie
Catherine M
CM
Blazar
Bruce R
BR
D000818:Animals; D002467:Cell Nucleus / Q000235:genetics / Q000378:metabolism; D004251:DNA Transposable Elements; D003952:Diagnostic Imaging; D005434:Flow Cytometry; D018014:Gene Transfer Techniques; D017930:Genes, Reporter; D008156:Luciferases; D008164:Luminescent Proteins; D051379:Mice; D008810:Mice, Inbred C57BL; D039902:Multipotent Stem Cells
D004251:DNA Transposable Elements; D008164:Luminescent Proteins; C413663:fluorescent protein 583; D008156:Luciferases