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Multiple sclerosis (MS) is a chronic inflammatory disease of the central nervous system (CNS) resulting in cumulative neurologic deficits associated with progressive myelin loss. We have previously shown that transplantation of neural progenitor cells (NPCs) into mice persistently infected with the JHM strain of mouse hepatitis virus (JHMV) results in enhanced differentiation into oligodendrocyte progenitor cells (OPCs) that is associated with remyelination and axonal sparing. The current study examines the contributions of the transcription factor Olig1 on NPC differentiation and remyelination. Under defined conditions, NPCs preferentially differentiate into oligodendroglia whereas NPCs isolated from Olig1-deficient (Olig1−/−) mice exhibit enhanced differentiation into astrocytes. Transplantation of Olig1−/− and Olig1+/+ NPCs into JHMV-infected mice resulted in similar cell survival, proliferation, and selective migration to areas of demyelination. However, only recipients of wild type NPCs exhibited extensive remyelination compared to mice receiving Olig1−/− NPCs. In vivo characterization of NPCs revealed that Olig1+/+ NPCs preferentially differentiated into NG2-positive OPCs and formed processes expressing myelin basic protein that encircled axons. In contrast, the majority of transplanted Olig1−/− NPCs differentiated into GFAP-positive cells consistent with the astrocyte lineage. These results indicate that exogenous NPCs contribute to improved clinical and histological outcome and this is associated with remyelination by this donor population. Further, these findings reveal that Olig1function is required for the remyelination potential of NPCs after transplant, through specification and/or maintenance of oligodendroglial identity.
An important clinical aspect related to the pathogenesis of the human demyelinating disease multiple sclerosis is the eventual remyelination failure in chronic demeylinated plaques by endogenous neural progenitor cells (NPCs) that give rise to oligodendrocyte precursor cells (OPCs) (Franklin and Ffrench-Constant, 2008). Such failure in myelin regeneration could be due to multiple factors including inflammation, inhibitor molecules present in the lesion or age-related deficits in endogenous OPCs (Fancy, et al., 2010). With this in mind, cell-based therapy using exogenous NPCs or OPCs have emerged as candidate therapies for promoting remyelination (Pluchino, et al., 2009, Sher, et al., 2008). Most studies have utilized either autoimmune models of neuroinflammatory-mediated demyelination, or chemical–induced gliotoxic demyelination to assess the remyelination potential of NPCs. While such models have advantages in capturing certain aspects of MS-like pathogenesis, they do not capture the full range of possible causative factors.
We have focused on a model of persistent viral infection that is correlative with chronic neuroinflammation and demyelination (Hosking and Lane, 2010). Our laboratory has recently demonstrated that transplantation of syngeneic mouse NPCs into mice persistently infected with the neurotropic JHM strain of mouse hepatitis virus (JHMV) is well tolerated and is associated with axonal sparing accompanied by extensive remyelination while not significantly dampening either neuroinflammation or T cell responses (Hardison, et al., 2006, Totoiu, et al., 2004). Evident from this work is i) the ability of engrafted cells to migrate to regions of demyelination by responding to the chemokine ligand CXCL12 (Carbajal, et al., 2010) and ii) preferential differentiation of transplanted NPCs into oligodendrocyte progenitor cells (OPCs) ((Carbajal, et al., 2010, Totoiu, et al., 2004). The experiments described here address the functional contributions of exogenous NPCs to remyelination.
The genes Olig1 and Olig2 encode basic helix-loop helix transcription factors that are expressed in neural progenitor cells (Zhou, et al., 2000) and are required for fundamental processes of CNS development including oligodendrocyte formation (Lu, et al., 2002). Olig1 is especially involved in oligodendrocyte development as well as maturation (Lu, et al., 2002). Previous studies have shown the implication of Olig1 in differentiation and remyelination in toxin induced models of demyelination (Arnett, et al., 2004). However these models do not take into account the potential effects of an inflammatory environment on NPCs. To this end, we have compared the remyelination and differentiation potential of competent and incompetent (Olig1−/−) NPCs post-transplant into JHMV-infected mice.
Five-week-old male C57BL/6 mice were purchased from the National Cancer Institute (Frederick, MD) and Olig1−/− male (Lu et al., 2002) mice (C57Bl/6 background) were bred in the UCI vivarium. For viral infection, mice were anesthetized by intraperitoneal (i.p.) injection of ketamine 80–100mg/kg (MP Biomedicals, OH) and xylazine 5–10 kg/mg (Phoenix Pharmaceutical, MO). Mice were infected i.c. with 200 plaque forming units (PFU) of JHMV (strain 2.2V-1) suspended in 30μl of sterile saline (Fleming, et al., 1986). Timed pregnant C57BL/6 mice (P14) were purchased from the National Cancer Institute and perinatal pups were used for wild type NPC cultures. C57BL/6-Tg (CAG-EGFP) 10sbJ mice (expressing green fluorescent protein, GFP) were purchased from JAX (stock#003291) (Lu, et al., 2002). C57BL/6-Tg (CAG-EGFP) 10sbJ mice were bred with Olig1−/− mice to produce the GFP-positive Olig1−/− colony (confirmed by PCR genotyping). All experiments were reviewed and approved by the University of California, Irvine Institutional Animal Care and Use Committee (IACUC).
Epidermal growth factor (EGF) responsive neurosphere cultures were prepared from either EGFP- Olig1+/+ or EGFP-Olig1−/− mice. Neurosphere cultures were prepared as previously described from the brains of perinatal animals (Ben-Hur, et al., 1998, Totoiu, et al., 2004). Briefly, dissected striata were razor minced and triturated in pre-warmed 0.05% Trypsin (Invitrogen) for 10 minutes. Trypsin digestion was halted with equal volume of 1X anti-trypsin (Invitrogen). Single cells were resuspended in DMEM: F12 (Invitrogen) supplemented with 1X B27 (Invitrogen), 1X Insulin-Transferrin-Selenium-X (Invitrogen), 1X Penicillin-Streptomycin (Invitrogen), 40 ng/ml T3 (Sigma-T67407), and 20 ng/ml human recombinant EGF (Sigma-E9644) and cultured for 5–6 days. Culture supernatant was replaced with fresh media containing EGF on days 1, 3, and 5. After one week, mature neurospheres (100–200μm) were transferred to matrigel (BD Bioscience) coated flasks (thin coat method, 1:30 dilution). Within 24 hours individual cells had spread out from attached spheres and formed a monolayer. Following formation of a monolayer formed, cells were trypsinized and suspended in sterile saline for transplant experiment.
To assess differentiation potential, cells were grown on matrigel coated imaging slides for a total of 4 days, fixed in 4% paraformaldehyde (Fisher Scientific, Fair Lawn, NJ) for 20 min and immunofluorescence staining was performed using standard protocols. Imaging chambers were blocked with 10% normal goat serum (NGS) (Vector Laboratories, Burlingame, CA) for 1 hr at room temperature. Primary antibodies (polyclonal rabbit anti-GalC, Chemicon, 1:50 dilution in 10% NGS; polyclonal rabbit anti-GFAP, Invitrogen, 1:500 dilution in 10% NGS; polyclonal rabbit anti-NG2, Chemicon, 1:200 dilution in 10% NGS or blocking solution (negative control, 10% NGS in PBS) were applied to chambers overnight at 4°C. Slides were rinsed three times with PBS and fluorescent-conjugated secondary antibody (Alexa 594, goat anti-rabbit) was applied and incubated for 1 hour at room temperature. Slides were rinsed three times in PBS and mounted in vectashield (Vector Laboratories) with Dapi to visualize cell nuclei. Cell quantification was conducted using a Nikon Eclipse Ti microscope, 200x magnification. The percentage of immunopositive cells was determined by dividing the total number of immunopositive cells by the total number of Dapi-positive cells in five images, multiplied by 100.
JHMV-infected mice develop demyelination associated with clinical disease 10–14 post-infection (p.i.) (Fleming, et al., 1986, Glass, et al., 2002). Transplant experiments were performed on days 14 p.i., when replicating virus is reduced below detectable levels and there is evidence of demyelinating lesions. Clinical severity was assessed using a previously described 4-point scale (Lane, et al., 2000). Only animals that developed partial-to-complete hind limb paralysis were used for transplantation. For transplant experiments, recipient mice with comparable clinical disease received either GFP-Olig1+/+ or GFP-Olig1−/− NPCs, or vehicle control (sterile saline). Anaesthetized animals received laminectory at T9-T10 to expose the spinal cord. Animals were then transplanted with 2.5μl of NPC (250,000 cells) or 2.5μl of sterile saline using a 10 μl Hamilton syringe (Hamilton) with a silicon-coated pulled glass tip affixed in a stereotactic arm as previously described (Nistor, et al., 2005, Totoiu, et al., 2004).
Transplanted recipient animals were euthanized at 21 and 35 days post-infection (p.i.) (7 and 21 days post-transplant) and tissue was fixed by intracardiac perfusion with 4% paraformaldehyde in PBS (pH 7.4). Intact spinal columns were removed and fixed overnight in 4% paraformaldehyde at 4°C. The bone was removed to expose the fixed spinal cord and the tissue 8 mm anterior and 8mm posterior from the injection site was divided into twelve tissue pieces (1mm). To evaluate GFP+ NPC migration and differentiation in vivo, tissue sections were cryoprotected in 30% sucrose for 7 days and embedded in OCT (Tissue-Tek). Seven-micron thick transverse sections were cut and used for immunofluorescence staining or stained with luxol fast blue (LFB) in combination with H&E (hematoxylin and eosin) to determine the extent of demyelination. To evaluate the remyelination potential of transplanted NPC, even tissue pieces were processed and embedded in resin while odd tissue pieces were processed and embedded in OCT. For resin sectioning, tissue pieces were exposed to 1% Osmium tetroxide (Electron Microscopy Sciences), dehydrated in ascending alcohols, and embedded in Spurr resin (Electron Microscopy Sciences) according to standard protocols. Transverse semi-thin (1 μm) sections were cut from each block, stained with alkaline toluidine blue, cover slipped, and examined by light microscopy using an Olympus BX-60 microscope, 600x magnification. The myelination of axons was determined by assessing the thickness of the myelin sheath in relation to the axons diameter (Guy, et al., 1989, Hildebrand and Hahn, 1978). Demyelinated axons, remyelinated axons and normally myelinated axons were counted within an area equal to 10% of the total area of demyelination. The quantitative assessment of remyelination was conducted throughout the region 8mm caudal and rostral to the transplant site. The number demyelinated axons, remyelinated axons, the total number of axons and the percent remyelinated axons was determined for each of the four regions on each tissue block, averaged, then averaged across animals within each group for each tissue block as previously described (Totoiu, et al., 2004).
The total numbers of GFP-positive cells was determined in each of the twelve sampled locations surrounding the transplant site by counting all GFP-positive cells co-localized with Dapi-positive nuclei. Cell migration was represented in graphs of the number of GFP-positive cells versus distance from transplant site (mm), and the approximate number of GFP-positive NPC 21 days post-transplant was analyzed by area under the curve calculation performed in GraphPad Prism (GraphPad Software) (Behrstock, et al., 2008).
To assess in vivo differentiation of GFP-positive NPC 21 days after transplant, sections were dehydrated, washed in PBS to removed excess OCT, and blocked for 1 h at room temperature with 10% goat serum in PBS. Immunofluorescence staining was performed using standard protocols. The following primary antibodies were added overnight at 4°C: rabbit anti-MBP 1:200 (Chemicon, cat#AB980), rabbit anti-GST-p 1:1,000 (MBL, cat#311), rabbit anti-NG2 1:200 (Chemicon, cat#AB5320), rabbit anti-GFAP 1:1,1000 (Invitrogen, cat#18-0063), rabbit anti-NF-150 (Chemicon, cat#AB1991). Appropriate conjugated goat secondary antibodies were used for visualization (Invitrogen). Slides were mounted in vectashield (Vector Laboratories) with DAPI to visualize cell nuclei and to preserve fluorescence. Cell quantification was conducted using an Olympus BX-60 microscope, 200x magnification. The percentage of immunopositive cells was determined by dividing the number of immunopositive cells by the number of Dapi-positive nuclei, multiplied by 100.
All data is presented as average ± SEM. Statistically significant differences were assessed by one-way ANOVA, and p values less than 0.05 were considered significant.
To investigate the importance of Olig1 in oligodendrocyte lineage commitment, NPCs were cultured from the brains of EGFP-Olig1+/+ (WT) and EGFP-Olig1-deficient mice (Olig1−/− mice). NPCs were cultured on matrigel-coated slides for 5 days to induce differentiation under defined conditions at which point defined cellular antigens were used to identify lineage commitment by immunocytochemical staining. By 5 days post-differentiation, the majority of cells cultured from WT mice expressed antigens NG2 and/or GalC that are markers associated with cells of the oligodendrocyte lineage (Figures 1A and C). Approximately 5% of cultured WT cells expressed the astrocyte-associated marker GFAP (Figures 1D and F). In contrast, genetic ablation of Olig1 in NPCs resulted in reduced expression of both NG2 and GalC (Figures 1B and C) while there was an ~ 2-fold increase in GFAP expression when compared to WT cells (Figures 1E and F). Immunocytochemical staining of differentiated WT and Olig1−/− NPCs revealed GalC-positive cells displaying an arborized morphology consistent with oligodendroglia (Figures 1A and B). GFAP-positive cells derived from either WT or Olig1−/− NPCs exhibited a more flat or stellate morphology (Figures 1D and E).
To avoid toxicity and induction of senescence associated with BrdU labeling of NPC (Ross, et al., 2008), we generated GFP-Olig1−/− mice and used NPC cultured from these mice for subsequent transplantation into mice. NPC cultured from GFP-C57BL/6 mice were used as WT controls. Two weeks prior to transplantation, recipient C57BL/6 mice were infected by i.c. injection of 200 PFU of JHMV, which is a sufficient viral dose to induce immune-mediated demyelination. Transplantation consisted of a single intraspinal injection of 2.5x105 cells (or vehicle control, HBSS) at thoracic vertebrae 9 and recipient animals were sacrificed one week and three weeks post-transplant to evaluate the extent of exogenous cell migration and engraftment. Luxol fast blue (LFB) staining of a representative JHMV-infected mouse 14 days p.i. indicated that at the time of transplantation, white matter demyelination was localized in the ventral white matter in the vicinity of the central canal (Figure 2A). Three weeks post-transplant of either WT or Olig1−/− NPCs, demyelinated lesions were found to extend from the ventral to the lateral white matter (Figure 2A). Quantification of demyelination in experimental mice indicated a similar level of myelin damage in recipients of either WT or Olig1−/− NPCs compared to vehicle control (Figure 2B). Importantly, these findings are consistent with earlier results and indicate that transplantation of NPCs neither exacerbate nor ameliorate JHMV-induced immunopathology and this was associated with no attenuation in neuroinflammation (Hardison, et al., 2006, Totoiu, et al., 2004).
Surgically engrafted NPCs preferentially migrate and accumulate within areas of white matter damage in JHMV-infected mice (Hardison, et al., 2006, Totoiu, et al., 2004). Moreover, we have determined that migration is mediated through CXCR4 expressed on transplanted NPCs responding to CXCL12 that is expressed within demyelinating lesions (Carbajal, et al., 2010). Transplanted WT and Olig1−/− NPCs exhibited similar migration both rostral and caudal to the site of implantation at 1 and 3 weeks post-transplant indicating that Olig1 function does not regulate migration (Figure 3A). In addition, similar numbers of WT and Olig1−/− NPCs were detected in transplanted mice indicating that replication is not negatively affected in the absence of Olig1 (Figure 3B). Importantly, positional migration of transplanted NPCs was not affected in the absence of Olig1 as both populations of NPCs preferentially migrated into ventral and lateral white matter columns of JHMV-infected mice (Figure 3C). Collectively, these findings provide compelling evidence that Olig1 function is not required for NPC migration, proliferation, or preferential accumulation within areas of white matter damage.
We next evaluated the extent of remyelination in JHMV-infected recipients of either WT or Olig1−/− NPCs at 3 weeks post-transplant. Spinal cord sections were used for either tracking migration of GFP-labeled NPCs or measuring remyelination to ensure that assessment of histopathology was performed within areas in which transplanted cells were present (Figure 4A). A representative spinal cord from a non-infected, non-transplanted mouse is shown to demonstrate normal myelin thickness (Figure 4B). Numerous demyelinated axons were present among vacuoles, myelin debris, and activated macrophages in JHMV-infected mice receiving either vehicle control (Figure 4C) or Olig1−/− NPCs (Figure 4D). In contrast, infected mice transplanted with WT NPCs exhibited numerous axons with thin myelin sheaths consistent with remyelination (Figure 4E). Quantification of remyelination in experimental groups of mice revealed an overall increase in the frequency of remyelinated axons in mice transplanted with WT NPCs when compared to either Olig1−/− NPCs or vehicle control (Figure 4F) (Totoiu, et al., 2004). Furthermore, immunofluorescent staining in combination with confocal microscopy revealed GFP signal overlying with MBP staining (Figure 5A) as well as GFP-positive wraps surrounding numerous axons (defined by neurofilament staining) (Figure 5B) in mice receiving WT NPCs yet this was absent in recipients of Olig1−/− NPCs. These findings support and extend previous work from our laboratory indicating that transplantation of NPCs into JHMV-infected mice results in extensive remyelination (Carbajal, et al., 2010, Totoiu, et al., 2004). The overall paucity in remyelinated axons in Olig1−/− NPCs suggests that although these cells are able to migrate and accumulate within areas of pathology they are not able to remyelinate demyelinated axons. Importantly, the demonstration of transplanted NPC-derived myelin wraps surrounding axons in WT NPC recipients provides compelling evidence that transplanted cells directly remyelinate axons.
Evaluation of differentiated WT and Olig1−/− NPCs at 3-weeks post-transplant in JHMV-infected mice was performed by immunofluorescence staining. Consistent with previous findings (Carbajal, et al., 2010), approximately 40% of the engrafted WT NPCs differentiated to an oligodendrocyte lineage as determined by NG2 and GST-π staining (Figure 6A). The majority of WT NPCs had differentiated into NG2-positive OPCs (27.78% ± 3.31) rather than mature oligodendrocytes as evidenced by GST-π staining (11.83% ± 0.72). In marked contrast, very few (<5%) of Olig1−/− NPCs expressed either NG2 (4.11% ± 1.92) or GST-π (2.7%± 0.55) indicating these cells did not preferentially differentiate into a myelin-competent cell (Figure 6A). Indeed, staining for the astrocyte marker GFAP revealed that the majority (>70%) of transplanted Olig1−/− NPCs were GFAP-positive (73.04% ± 3.1) and displayed a morphology consistent with an activated astrocyte (Figure 6B). However, fewer than 20% of transplanted WT NPCs differentiated in GFAP-positive cells (17.05% ± 3.57) (Figure 6B).
There are no effective therapies available for patients with progressive forms of MS. For this and other devastating human white matter disorders, exogenous NPC and OPC transplantation are of great interest as having potential therapeutic roles (Conti, et al., 2006, Fancy, et al., 2010, Kim and de Vellis, 2009, Ligon, et al., 2006, Pluchino and Martino, 2005, Sher, et al., 2008, Tirotta, et al., 2010, Zhao, et al., 2008). The use of a viral model of demyelination to evaluate the remyelination potential of NPCs has unique features that make this a relevant experimental model system. First, the etiology of the human demyelinating disease is enigmatic with both genetic factors and environmental influences considered important in initiation and maintenance of disease. Viral infection has long been viewed as a potential triggering mechanism involved in demyelination and numerous human viral pathogens have been suggested to be involved in eliciting myelin-reactive lymphocytes and/or antibodies that subsequently infiltrate the CNS and damage the myelin sheath (Ascherio and Munger, 2007, Ebers, et al., 1995, Sospedra and Martin, 2005). Therefore, viral models of demyelination are clearly relevant and have provided important insight into mechanisms associated with disease initiation, neuroinflammation and demyelination. Given the possibility of viral infection in initiating demyelination as well as the fact that numerous neurotropic viruses exist that are capable of persisting within the CNS, it is imperative to evaluate the remyelination potential of stem cells in the presence of a persistent viral infection that is correlative with chronic neuroinflammation and demyelination
The present study sought to determine the importance of Olig1 in NPC-mediated remyelination in JHMV-infected mice with established demyelination. To address this, we used NPCs derived from mice deficient in the transcription factor Olig1. The genes Olig1 and Olig2 encode basic helix-loop helix transcription factors that are expressed in neural progenitor cells (Zhou, et al., 2000) and are required for fundamental processes of CNS development including oligodendrocyte formation (Lu, et al., 2002). The Olig1-deficient mice we employed are viable and have mildly delayed myelination during development (Lu, et al., 2002). However, they showed a profound deficiency in remyelination, indicating a nonredundant role for Olig1 in remyelination due to a defect in OPC differentiation to oligodendrocytes after white matter injury (Arnett, et al., 2004). We note that this contrasts findings with a different Olig1-null line that shows a lethal developmental hypomyelination phenotype (Xin, et al., 2005). Thus, Olig1 is critical for normal myelination and remyelination.
We took advantage of this genetic requirement of Olig1 in repair to determine if intraspinal transplantation of Olig1−/− NPCs resulted in remyelination. Our findings reveal that engrafted Olig1−/− NPCs were well tolerated, replicated, and migrated to a similar extent along white matter spinal cord tracts compared to WT NPCs. However, there was a marked reduction in the number of remyelinated axons in comparison to recipients of control cells. Paucity of remyelination from Olig1−/− donor cells was associated with an apparent change in lineage fate commitment, as Olig1-null mice preferentially differentiated into GFAP-positive cells whereas WT cells exhibited commitment to an oligodendrocyte lineage. This was confirmed by our in vitro studies showing the importance of Olig1−/− to enhance oligodendroglial lineage commitment, as the absence of this transcription factor seemed to favor the differentiation into astrocytes by NPCs.
In all recipients, WT NPCs migrated to demyelinated white matter and formed MBP-containing processes that encircle neurofilament positive axons. These findings are consistent with several possibilities. First, Olig1 may be required to regulate cell fate differentiation of NPCs into OPCs; alternatively, Olig1 may be required to maintain OPC fate and in the absence of Olig1 function OPCs will “trans-differentiate” to astrocytes. We favor the former argument as in Olig1−/− mice OPC fate and myelination proceeds to a level comparable to wildtype mice demonstrating Olig1 is not strictly required to maintain OPC fate. Therefore, our findings indicate that Olig1 is not required for migration into lesions but is important in selecting an oligodendrocyte lineage fate. These findings differ from, but are not in conflict, with Arnett et al (Arnett, et al., 2004) where OPCs were recruited into demyelinating lesions induced by lysolethicin treatment but failed to mature to participate in remyelination. Of note, in this study no increased astrogliosis in lesions was observed. Similarly, transplanted Olig1−/− NPCs migrated yet did exhibit differentiation primarily into astrocytes. An important difference in these results may reflect differences in model systems utilized. While lysolethicin induces focal demyelinating lesions in the absence of infiltration of activated T lymphocytes and monocyte/macrophages, JHMV-induced demyelination is characterized by the presence of activated T lymphocytes as well as other inflammatory cells that results in the secretion of numerous proinflammatory cytokines/chemokines (Bergmann, et al., 2006, Hosking and Lane, 2010, Lane and Hosking, 2010). Therefore, one intriguing possibility is that the inflammatory microenvironment may tailor NPCs fate decision through an Olig1-regulated mechanism. In addition, these findings are consistent with other studies highlighting the importance of Olig1 in contributing to myelin repair following experimental demyelination (Burton, 2005, Ligon, et al., 2006, Maire, et al., 2010, Tsiperson, et al., 2010).
Our findings suggest that exogenous NPCs actively participate in remyelination following engraftment. This is supported by counting remyelinated axons in experimental animals as well as the presence of GFP-positive wraps encircling axons following transplantation of wild type NPCs. Our findings are consistent with earlier studies by Cummings et al. (Cummings, et al., 2005) that demonstrated engraftment of human NPCs promoted locomotor recovery in a rodent model of spinal cord injury. Importantly, recovery was abolished by selective ablation of engrafted cells suggesting that the therapeutic benefit was mediated primarily by engrafted cells. However, a recent report indicated that transplanted neural progenitors were shown to enhance proliferation of host OPCs (Einstein, et al., 2009). This was associated with increased remyelination in a model of cuprizone-mediated demyelination indicating that transplanted neural progenitors stimulated endogenous cells to participate in repair. Differences between experimental outcomes most likely reflect differences within the model systems employed as cuprizone represents an acute and focal model of demyelination with a limited role for activated lymphocytes in participating in myelin destruction and this is dramatically different compared to JHMV-induced demyelination. Therefore, the environmental signals encountered by engrafted cells will modulate the ability of the engrafted cell to home, differentiate, and participate in repair. With this in mind, our results reveal insight into the importance of Olig1 within the context of engrafted NPCs into an ongoing immune-mediated demyelinating disease initiated by viral infection: i) Olig1 does not influence NPC positional migration, ii) Olig1 is required for differentiation into OPCs/oligodendrocytes, and iii) preferential differentiation into oligolineage cells is associated with increased remyelination.
Role of the funding source
The funding source had no involvement in study design or in the collection, analysis and interpretation of data as well as in writing of the report and decision of submission for publication.
This work was funded by National Institutes of Health (NIH) Grant R01 NS041249, National Multiple Sclerosis Society (NMSS) Grant RG3857A5, and a Collaborative Center Research Award from NMSS to T.E.L. D.H.R. is an Investigator of the Howard Hughes Medical Institute and is also funded by NIH R01 NS040511. L.M.W. and C.S.S. were supported by California Institute for Regenerative Medicine Training Grant T1-00008.
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