Bioluminescence Imaging of Olig2-Neural Stem Cells Reveals Improved Engraftment in a Demyelination Mouse Model

Abstract

A major issue in the potential application of neural stem cell (NSC)-based cell replacement therapy for demyelinating diseases is the question of the survival, functional behavior, and stability of implanted NSC-derived oligodendrocyte precursor cells (OPCs) over an extended period. To address this issue, we employed bioluminescence imaging (BLI) as a noninvasive longitudinal in vivo monitoring technique and followed the fate of NSCs isolated from luciferase-green fluorescent protein-actin transgenic mice after stereotactic implantation in the demyelinated corpus callosum of cuprizone-fed mice. We compared normal NSCs with NSCs that were primed to become OPCs by the induction of Olig2 overexpression (Olig2-NSCs). BLI, validated by immunohistochemistry, revealed that, after a steep cell loss after implantation during the first 3 weeks, approximately 10% of the Olig2-NSCs stably survived for 2 months after implantation, in contrast to <1% of the normal NSCs. Immunohistochemistry, at the light and electron microscopic levels, revealed that the majority of the surviving Olig2-NSCs had differentiated into an oligodendrocytic cell lineage and contributed to remyelination of axons in the corpus callosum. The number of axons remyelinated by the implanted cells, however, was a small fraction of the total number of axons remyelinated by endogenous oligodendrocytes. Apparently, most of the implanted NSCs did not survive the transition into an inappropriate non-neurogenic niche, compressed by surrounding host tissue, in hostile, inflammatory conditions created by activated microglia. Only the ones that managed to differentiate rapidly into a mature neural cell type and become functionally integrated survived. STEM CELLS

Differentiation of induced pluripotent stem cells into functional oligodendrocytes

“…On the day of transplantation, cells were harvested with accutase, counted, and resuspended in PBS at concentration of 25,000 cells/lL. Four microliters of cell suspension was injected into corpus callosum of C57BL/6 mice using the following stereotactic co-ordinates (in reference to Bregma point): 10.98 mm (anterioposterior axis), 21.75 mm (lateromedial axis), 22.25 mm (vertical axis) (Copray et al, 2006;Sher et al, 2009), suspensions of 100,000 cells in 4 lL PBS were slowly injected into the corpus callosum of ketamineanesthetized mice using a 10 lL Hamilton injection syringe (22s/2 00 /3) (Hamilton, Reno, Nevada, 80365).…”

“…So far, potential clinically relevant OPCs have been derived from neural stem cells (NSCs) (Ben-Hur et al, 2003;Einstein et al, 2007;Magalon et al, 2007;Pluchino et al, 2003;Sher et al, 2009) or ES cells (Brustle et al, 1999;Liu et al, 2000;Nistor et al, 2005). Both cell types have unacceptable limitations as sources for clinically relevant transplantable OPCs, related to their restricted availability in case of autologous NSCs and heterologous, ethically disputed origin in case of ES cells.…”

Comparison of red-shifted firefly luciferase Ppy RE9 and conventional Luc2 as bioluminescence imaging reporter genes for <italic>in vivo</italic> imaging of stem cells

“…4,5 Introduced expression of luciferase enables researchers to observe luminescence upon injection of its substrates and therefore to monitor the survival, migration, or differentiation of stem cells over time with high sensitivity. Two major directions of improvements have been pursued for optimization of luciferase genes for in vivo imaging.…”

The Myelin Mutants as Models to Study Myelin Repair in the Leukodystrophies

“…This has been demonstrated both ex vivo and in vivo in the md rat and migrations of cells followed longitudinally in the les rat spinal cord [86,87]. Neural stem cells induced to over-express Olig-2, which were derived from a luciferase-GFP-actin transgenic mouse and transplanted into a mouse fed cuprizone, were detected by bioluminescence imaging in the brain of live mice [88].…”