Shiverer Mice Research Articles

Multisite Injections of Canine Glial-Restricted Progenitors Promote Brain Myelination and Extend the Survival of Dysmyelinated Mice

Glial cell dysfunction results in myelin loss and leads to subsequent motor and cognitive deficits throughout the demyelinating disease course.Therefore, in various therapeutic approaches, significant attention has been directed toward glial-restricted progenitor (GRP) transplantation for myelin repair and remyelination, and numerous studies using exogenous GRP injection in rodent models of hypomyelinating diseases have been performed. Previously, we proposed the transplantation of canine glial-restricted progenitors (cGRPs) into the double-mutant immunodeficient, demyelinated neonatal shiverer mice (shiverer/Rag2−/−). The results of our previous study revealed the myelination of axons within the corpus callosum of transplanted animals; however, the extent of myelination and lifespan prolongation depended on the transplantation site (anterior vs. posterior). The goal of our present study was to optimize the therapeutic effect of cGRP transplantation by using a multisite injection protocol to achieve a broader dispersal of donor cells in the host and obtain better therapeutic results. Experimental analysis of cGRP graft recipients revealed a marked elevation in myelin basic protein (MBP) expression and prominent axonal myelination across the brains of shiverer mice. Interestingly, the proportion of galactosyl ceramidase (GalC) positive cells was similar between the brains of cGRP recipients and control mice, implying a natural propensity of exogenous cGRPs to generate mature, myelinating oligodendrocytes. Moreover, multisite injection of cGRPs improved mice survival as compared to non-transplanted animals.

Quantitative Analysis of Wild Type (WT) and Shiverer (SHR) Mouse Spinal Cords using Ultrahigh-b DWI (UHb-DWI) (P3-4.010)

Quantitative Analysis of Wild Type (WT) and Shiverer (SHR) Mouse Spinal Cords using Ultrahigh-b DWI (UHb-DWI) (P3-4.010)

Inhomogeneous magnetization transfer MRI of white matter structures in the hypomyelinated shiverer mousebrain.

Inhomogeneous magnetization transfer (ihMT) MRI is uniquely sensitive to myelin with lipids as a primary source of its contrast. In this study, we investigated whether ihMT can detect white matter structures in the hypomyelinated shiverer mouse brain, a model of dysmyelination. Conventional MT and ihMT images were acquired from ex vivo Rag2-/- control and shiverer mouse brains at 7T using previously reported optimized saturation parameters. ihMT ratio (ihMTR) maps revealed hypomyelinated corpus callosum in the shiverer mouse brain, whereas conventional MT ratio (MTR) maps showed no clear contrast. The ihMTR values of the corpus callosum in the shiverer mice were reduced by approximately 40% compared to controls, but remained significantly higher than the ihMTR values of the cortex. The finding further confirms ihMT's high myelin specificity and suggests its use as a marker to detect early myelination or myelin repair.

OCT4-induced oligodendrocyte progenitor cells promote remyelination and ameliorate disease

The generation of human oligodendrocyte progenitor cells (OPCs) may be therapeutically valuable for human demyelinating diseases such as multiple sclerosis. Here, we report the direct reprogramming of human somatic cells into expandable induced OPCs (iOPCs) using a combination of OCT4 and a small molecule cocktail. This method enables generation of A2B5+ (an early marker for OPCs) iOPCs within 2 weeks retaining the ability to differentiate into MBP-positive mature oligodendrocytes. RNA-seq analysis revealed that the transcriptome of O4+ iOPCs was similar to that of O4+ OPCs and ChIP-seq analysis revealed that putative OCT4-binding regions were detected in the regulatory elements of CNS development-related genes. Notably, engrafted iOPCs remyelinated the brains of adult shiverer mice and experimental autoimmune encephalomyelitis mice with MOG-induced 14 weeks after transplantation. In conclusion, our study may contribute to the development of therapeutic approaches for neurological disorders, as well as facilitate the understanding of the molecular mechanisms underlying glial development.

Derivation of Oligodendrocyte Precursors from Adult Bone Marrow Stromal Cells for Remyelination Therapy.

Transplantation of oligodendrocyte precursors (OPs) is potentially therapeutic for myelin disorders but a safe and accessible cell source remains to be identified. Here we report a two-step protocol for derivation of highly enriched populations of OPs from bone marrow stromal cells of young adult rats (aMSCs). Neural progenitors among the aMSCs were expanded in non-adherent sphere-forming cultures and subsequently directed along the OP lineage with the use of glial-inducing growth factors. Immunocytochemical and flow cytometric analyses of these cells confirmed OP-like expression of Olig2, PDGFRα, NG2, and Sox10. OPs so derived formed compact myelin both in vitro, as in co-culture with purified neurons, and in vivo, following transplantation into the corpus callosum of neonatal shiverer mice. Not only did the density of myelinated axons in the corpus callosum of recipient shiverer mice reach levels comparable to those in age-matched wild-type mice, but the mean lifespan of recipient shiverer mice also far exceeded those of non-recipient shiverer mice. Our results thus promise progress in harnessing the OP-generating potential of aMSCs towards cell therapy for myelin disorders.

Ultrahigh-b diffusion-weighted imaging for quantitative evaluation of myelination in shiverer mouse spinal cord.

To perform a quantitative evaluation of myelination on WT and myelin-deficient (shiverer) mouse spinal cords using ultrahigh-b diffusion-weighted imaging (UHb-DWI). UHb-DWI of ex vivo on spinal cord specimens of two shiverer (C3HeB/FeJ-shiverer, homozygous genotype for MbPshi ) and six WT (Black Six, C3HeB/FeJ) mice were acquired using 3D multishot diffusion-weighted stimulated-echo EPI, a homemade RF coil, and a small-bore 7T MRI system. Imaging was performed in transaxial plane with 75 × 75 μm2 in-plane resolution, 1-mm-slice thickness, and radial DWI using bmax = 42,890 s/mm2 . Histological evaluation was performed on upper thoracic sections using optical and transmission electron microscopy. Numerical Monte Carlo simulations (MCSs) of water diffusion were performed to facilitate interpretation of UHb-DWI signal-b curves. The white matter ultrahigh-b radial DWI (UHb-rDWI) signal-b curves of WT mouse cords behaved biexponentially with high-b diffusion coefficient DH < 0.020 × 10-3 mm2 /s. However, as expected with less myelination, the signal-b of shiverer mouse cords behaved monoexponentially with significantly greater DH = 0.162 × 10-3 , 0.142 × 10-3 , and 0.164 × 10-3 mm2 /s at anterodorsal, posterodorsal, and lateral columns, respectively. The axial DWI signals of all mouse cords behaved monoexponentially with D = (0.718-1.124) × 10-3 mm2 /s. MCS suggests that these elevated DH are mainly induced by increased water exchange at the myelin sheath. Microscopic results were consistent with the UHb-rDWI findings. UHb-DWI provides quantitative differences in myelination of spinal cords from myelin-deficit shiverer and WT mice. UHb-DWI may become a powerful tool to evaluate myelination in demyelinating disease models that may translate to human diseases, including multiple sclerosis.

Oligodendrocyte progenitor cell maturation is dependent on dual function of MCT8 in the transport of thyroid hormone across brain barriers and the plasma membrane.

Inactivating mutations in the thyroid hormone (TH) transporter monocarboxylate transporter 8 (MCT8) causes a rare and debilitating form of X-linked psychomotor disability known as Allan Herndon Dudley syndrome (AHDS). One of the most prominent pathophysiological symptoms of MCT8-deficiency is hypomyelination. Here, patient-derived induced pluripotent stem cells (iPSCs) were used to study the role of MCT8 and TH on the maturation of oligodendrocytes. Interestingly, neither MCT8 mutations nor reduced TH affected the in vitro differentiation of control or MCT8-deficient iPSCs into oligodendrocytes. To assess whether patient-derived iPSC-derived oligodendrocyte progenitor cells (iOPCs) could provide myelinating oligodendrocytes in vivo, cells were transplanted into the shiverer mouse corpus callosum where they survived, migrated, and matured into myelinating oligodendrocytes, though the myelination efficiency was reduced compared with control cells. When MCT8-deficient and healthy control iOPCs were transplanted into a novel hypothyroid immunodeficient triple knockout mouse (tKO, mct8-/- ; oatp1c1-/- ; rag2-/- ), they failed to provide behavioral recovery and did not mature into oligodendrocytes in the hypothyroid corpus callosum, demonstrating the critical role of TH transport across brain barriers in oligodendrocyte maturation. We conclude that MCT8 plays a cell autonomous role in oligodendrocyte maturation and that functional TH transport into the central nervous system will be required for developing an effective treatment for MCT8-deficient patients.

Traumatic brain injury does not disrupt costimulatory blockade-induced immunological tolerance to glial-restricted progenitor allografts

BackgroundCell transplantation-based treatments for neurological disease are promising, yet graft rejection remains a major barrier to successful regenerative therapies. Our group and others have shown that long-lasting tolerance of transplanted stem cells can be achieved in the brain with systemic application of monoclonal antibodies blocking co-stimulation signaling. However, it is unknown if subsequent injury and the blood-brain barrier breach could expose the transplanted cells to systemic immune system spurring fulminant rejection and fatal encephalitis. Therefore, we investigated whether delayed traumatic brain injury (TBI) could trigger graft rejection.MethodsGlial-restricted precursor cells (GRPs) were intracerebroventricularly transplanted in immunocompetent neonatal mice and co-stimulation blockade (CoB) was applied 0, 2, 4, and 6 days post-grafting. Bioluminescence imaging (BLI) was performed to monitor the grafted cell survival. Mice were subjected to TBI 12 weeks post-transplantation. MRI and open-field test were performed to assess the brain damage and behavioral change, respectively. The animals were decapitated at week 16 post-transplantation, and the brains were harvested. The survival and distribution of grafted cells were verified from brain sections. Hematoxylin and eosin staining (HE) was performed to observe TBI-induced brain legion, and neuroinflammation was evaluated immunohistochemically.ResultsBLI showed that grafted GRPs were rejected within 4 weeks after transplantation without CoB, while CoB administration resulted in long-term survival of allografts. BLI signal had a steep rise following TBI and subsequently declined but remained higher than the preinjury level. Open-field test showed TBI-induced anxiety for all animals but neither CoB nor GRP transplantation intensified the symptom. HE and MRI demonstrated a reduction in TBI-induced lesion volume in GRP-transplanted mice compared with non-transplanted mice. Brain sections further validated the survival of grafted GRPs and showed more GRPs surrounding the injured tissue. Furthermore, the brains of post-TBI shiverer mice had increased activation of microglia and astrocytes compared to post-TBI wildtype mice, but infiltration of CD45+ leukocytes remained low.ConclusionsCoB induces sustained immunological tolerance towards allografted cerebral GRPs which is not disrupted following TBI, and unexpectedly TBI may enhance GRPs engraftment and contribute to post-injury brain tissue repair.

Study on the Safety of Human Oligodendrocyte Precursor Cell Transplantation in Young Animals and Its Efficacy on Myelination.

Oligodendrocyte precursor cells (OPCs) can differentiate into myelinating oligodendrocytes during embryonic development, thereby representing an important potential source for myelin repair or regeneration. To the best of our knowledge, there are very few OPCs from human sources (human-derived OPCs [hOPCs]). In this study, we aimed to evaluate the safety and remyelination capacity of hOPCs developed in our laboratory, transplanted into the lateral ventricles of young animals. Several acute and chronic toxicity experiments were conducted in which different doses of hOPCs were transplanted into the lateral ventricles of Sprague–Dawley rats of different ages. The toxicity, biodistribution, and tumor formation ability of the injected hOPCs were examined by evaluating the rats' vital signs, developmental indicators, neural reflexes, as well as by hematology, immunology, and pathology. In addition, the hOPCs were transplanted into the corpus callosum of the shiverer mouse to verify cell myelination efficacy. Overall, our results show that transplanted hOPCs into young mice are nontoxic to their organ function or immune system. The transplanted cells engrafted in the brain and did not appear in other organs, nor did they cause tissue proliferation or tumor formation. In terms of efficacy, the transplanted hOPCs were able to form myelin in the corpus callosum, alleviate the trembling phenotype of shiverer mice, and promote normal development. The transplantation of hOPCs is safe; they can effectively form myelin in the brain, thereby providing a theoretical basis for the future clinical transplantation of hOPCs.

The Absence of Myelin Basic Protein Reduces Non-Amyloidogenic Processing of Amyloid Precursor Protein.

The accumulation of amyloid β-protein (Aβ) in the brain is a pathological feature of Alzheimer's disease (AD). Aβ peptides originate from amyloid precursor protein (APP). APP can be proteolytically cleaved through amyloidogenic or non-amyloidogenic pathways. The molecular effects on APP metabolism/processing may be influenced by myelin and the breakdown of myelin basic protein (MBP) in AD patients and mouse models of AD pathology. We directly tested whether MBP can alter influence APP processing in MBP-/- mice, known as Shiverer (shi/shi) mice, in which no functional MBP is produced due to gene breakage from the middle of MBP exon ll. A significant reduction of the cerebral sAPPα level in Shiverer (shi/shi) mice was found, although the levels of both total APP and sAPPβ remain unchanged. The reduction of sAPPα was considered to be due to the changes in the expression levels of a disintegrin and metalloproteinase-9 (ADAM9) catalysis and non-amyloid genic processing of APP in the absence of MBP because it binds to ADAM9. MBP -/- mice exhibited increased Aβ oligomer production. These findings suggest that in the absence of MBP, there is a marked reduction of nonamyloidogenic APP processing to sAPPα, and targeting myelin of oligodendrocytes may be a novel therapy for the prevention and treatment of AD.

Sensitivity to myelin using model-free analysis of the water resonance line-shape in postmortem mouse brain.

Dysmyelinating diseases are characterized by abnormal myelin formation and function. Such microstructural abnormalities in myelin have been demonstrated to produce measurable effects on the MR signal. This work examines these effects on measurements of voxel-wise, high-resolution water spectra acquired using a 3D echo-planar spectroscopic imaging (EPSI) pulse sequence from both postmortem fixed control mouse brains and a dysmyelination mouse brain model. Perfusion fixed, resected control (n = 5) and shiverer (n = 4) mouse brains were imaged using 3D-EPSI with 100 µm isotropic resolution. The free induction decay (FID) was sampled every 2.74 ms over 192 echoes, for a total sampling duration of 526.08 ms. Voxel-wise FIDs were Fourier transformed to produce water spectra with 1.9 Hz resolution. Spectral asymmetry was computed and compared between the two tissue types. The water resonance is more asymmetrically broadened in the white matter of control mouse brain compared with dysmyelinated white matter. In control brain, this is modulated by and consistent with previously reported orientationally dependent effects of white matter relative to B0 . Similar sensitivity to orientation is observed in dysmyelinated white matter as well; however, the magnitude of the resonance asymmetry is much lower across all directions. Results demonstrate that components of the spectra are specifically differentially affected by myelin concentration. This suggests that water proton spectra may be sensitive to the presence of myelin, and as such, could serve as a MRI-based biomarker of dysmyelinating disease, free of mathematical models.

The Median Eminence, A New Oligodendrogenic Niche in the Adult Mouse Brain.

SummaryThe subventricular zone (SVZ) of the lateral ventricles and the subgranular zone (SGZ) of the dentate gyrus in the hippocampus are known as neurogenic niches. We show that the median eminence (ME) of the hypothalamus comprises BrdU+ newly proliferating cells co-expressing NG2 (oligodendrocyte progenitors) and RIP (pre-myelinating oligodendrocytes), suggesting their differentiation toward mature oligodendrocytes (OLs). ME cells can generate neurospheres (NS) in vitro, which differentiate mostly to OLs compared with SVZ-NS that typically generate neurons. Interestingly, this population of oligodendrocyte progenitors is increased in the ME from experimental autoimmune encephalomyelitis (EAE)-affected mice. Notably, the thrombospondin 1 (TSP1) expressed by astrocytes, acts as negative regulator of oligodendrogenesis in vitro and is downregulated in the ME of EAE mice. Importantly, transplanted ME-NS preferentially differentiate to MBP+ OLs compared with SVZ-NS in Shiverer mice. Hence, discovering the ME as a new site for myelin-producing cells has a great importance for advising future therapy for demyelinating diseases and spinal cord injury.

Human Glial Progenitor Cells Effectively Remyelinate the Demyelinated Adult Brain

Neonatally transplanted human glial progenitor cells (hGPCs) can myelinate the brains of myelin-deficient shiverer mice, rescuing their phenotype and survival. Yet, it has been unclear whether implanted hGPCs are similarly able to remyelinate the diffusely demyelinated adult CNS. We, therefore, ask if hGPCs could remyelinate both congenitally hypomyelinated adult shiverers and normal adult mice after cuprizone demyelination. In adult shiverers, hGPCs broadly disperse and differentiate as myelinating oligodendrocytes after subcortical injection, improving both host callosal conduction and ambulation. Implanted hGPCs similarly remyelinate denuded axons after cuprizone demyelination, whether delivered before or after demyelination. RNA sequencing (RNA-seq) of hGPCs back from cuprizone-demyelinated brains reveals their transcriptional activation of oligodendrocyte differentiation programs, while distinguishing them from hGPCs not previously exposed to demyelination. These data indicate the ability of transplanted hGPCs to disperse throughout the adult CNS, to broadly myelinate regions of dysmyelination, and also to be recruited as myelinogenic oligodendrocytes later in life, upon demyelination-associated demand.

Transplanted t(1;11) patient-derived OPCs form shorter myelin internodes in the hypomyelinated shiverer mice

Transplanted t(1;11) patient-derived OPCs form shorter myelin internodes in the hypomyelinated shiverer mice

Inappropriate Intrusion of an Axonal Mitochondrial Anchor into Dendrites Causes Neurodegeneration

SUMMARYSyntaphilin (SNPH) is a major mitochondrial anchoring protein targeted to axons and excluded from dendrites. In this study, we provide in vivo evidence that this spatial specificity is lost in Shiverer (Shi) mice, a model for progressive multiple sclerosis (MS), resulting in inappropriate intrusion of SNPH into dendrites of cerebellar Purkinje cells with neurodegenerative consequences. Thus, reconstituting dendritic SNPH intrusion in SNPH-KO mice by viral transduction greatly sensitizes Purkinje cells to excitotoxicity when the glutamatergic climbing fibers are stimulated. Finally, we demonstrate in vitro that overexpression of SNPH in dendrites compromises neuronal viability by inducing N-methyl-D-aspartate (NMDA) excitotoxicity, reducing mitochondrial calcium uptake, and interfering with quality control of mitochondria by blocking somal mitophagy. Collectively, we propose that inappropriate immobilization of dendritic mitochondria by SNPH intrusion produces excitotoxicity and suggest that interception of dendritic SNPH intrusion is a therapeutic strategy to combat neurodegeneration.

Development of glial restricted human neural stem cells for oligodendrocyte differentiation in vitro and in vivo

In this study, we have developed highly expandable neural stem cells (NSCs) from HESCs and iPSCs that artificially express the oligodendrocyte (OL) specific transcription factor gene Zfp488. This is enough to restrict them to an exclusive oligodendrocyte progenitor cell (OPC) fate during differentiation in vitro and in vivo. During CNS development, Zfp488 is induced during the early stages of OL generation, and then again during terminal differentiation of OLs. Interestingly, the human ortholog Znf488, crucial for OL development in human, has been recently identified to function as a dorsoventral pattering regulator in the ventral spinal cord for the generation of P1, P2/pMN, and P2 neural progenitor domains. Forced expression of Zfp488 gene in human NSCs led to the robust generation of OLs and suppression of neuronal and astrocyte fate in vitro and in vivo. Zfp488 expressing NSC derived oligodendrocytes are functional and can myelinate rat dorsal root ganglion neurons in vitro, and form myelin in Shiverer mice brain in vivo. After transplantation near a site of demyelination, Zfp488 expressing hNSCs migrated to the lesion and differentiated into premyelinating OLs. A certain fraction also homed in the subventricular zone (SVZ). Zfp488-ZsGreen1-hNSC derived OLs formed compact myelin in Shiverer mice brain seen under the electron microscope. Transplanted human neural stem cells (NSC) that have the potential to differentiate into functional oligodendrocytes in response to remyelinating signals can be a powerful therapeutic intervention for disorders where oligodendrocyte (OL) replacement is beneficial.

Generation of Oligodendrocyte Progenitor Cells From Mouse Bone Marrow Cells.

Oligodendrocyte progenitor cells (OPCs) are a subtype of glial cells responsible for myelin regeneration. Oligodendrocytes (OLGs) originate from OPCs and are the myelinating cells in the central nervous system (CNS). OLGs play an important role in the context of lesions in which myelin loss occurs. Even though many protocols for isolating OPCs have been published, their cellular yield remains a limit for clinical application. The protocol proposed here is novel and has practical value; in fact, OPCs can be generated from a source of autologous cells without gene manipulation. Our method represents a rapid, and high-efficiency differentiation protocol for generating mouse OLGs from bone marrow-derived cells using growth-factor defined media. With this protocol, it is possible to obtain mature OLGs in 7–8 weeks. Within 2–3 weeks from bone marrow (BM) isolation, after neurospheres formed, the cells differentiate into Nestin+ Sox2+ neural stem cells (NSCs), around 30 days. OPCs specific markers start to be expressed around day 38, followed by RIP+O4+ around day 42. CNPase+ mature OLGs are finally obtained around 7–8 weeks. Further, bone marrow-derived OPCs exhibited therapeutic effect in shiverer (Shi) mice, promoting myelin regeneration and reducing the tremor. Here, we propose a method by which OLGs can be generated starting from BM cells and have similar abilities to subventricular zone (SVZ)-derived cells. This protocol significantly decreases the timing and costs of the OLGs differentiation within 2 months of culture.

Generation of Anterior Hindbrain-Specific, Glial-Restricted Progenitor-Like Cells from Human Pluripotent Stem Cells.

Engraftment of oligodendrocyte progenitor cells (OPCs), which form myelinating oligodendrocytes, has the potential to treat demyelinating diseases such as multiple sclerosis. However, conventional strategies for generating oligodendrocytes have mainly focused on direct differentiation into forebrain- or spinal cord-restricted oligodendrocytes without establishing or amplifying stem/progenitor cells. Taking advantage of a recently established culture system, we generated expandable EN1- and GBX2-positive glial-restricted progenitor-like cells (GPLCs) near the anterior hindbrain. These cells expressed PDGFRα, CD9, S100β, and SOX10 and mostly differentiated into GFAP-positive astrocytes and MBP-positive oligodendrocytes. RNA-seq analysis revealed that the transcriptome of GPLCs was similar to that of O4-positive OPCs, but distinct from that of rosette-type neural stem cells. Notably, engrafted GPLCs not only differentiated into GFAP-positive astrocytes but also myelinated the brains of adult shiverer mice 8 weeks after transplantation. Our strategy for establishing anterior hindbrain-specific GPLCs with gliogenic potency will facilitate their use in the treatment of demyelinating diseases and studies of the molecular mechanisms underlying glial development in the hindbrain.

CRISPR/Cas9‐Mediated Rescue of Shiverer Mice (MBPShi) via AAV9 Delivery across the Blood‐Brain Barrier

The shiverer mouse is a genetically modified strain of M. musculus which exhibits ataxia, tremor, and seizure, among other symptoms, due to impaired CNS function. The strain is generated through deletion of a large portion of the Myelin Basic Protein (MBP) gene. Due to lack of MBP, the shiverer mouse presents with hypomyelination and the corresponding “shivering” phenotype. To date, shiverer mouse rescue has been accomplished through embryological stem‐cell implant, or gene delivery via zygote microinjection. However, these methods are limited to rescue of the subsequent generation and cannot be applied to a mouse of the current generation. Another obstacle to consider when exploring pathologies affecting the CNS is the ability of a therapeutic candidate to cross the blood‐brain barrier (BBB). Here we propose a novel method for the genetic rescue of current‐generation shiverer mice. This method involves CRISPR‐mediated insertion of a functional MBP transgene via the delivery of an AAV9 delivery mechanism. This involves a simple procedure of non‐specific intravenous injection of the mouse, relying on viral delivery across the BBB and bypassing the need for invasive surgical intervention.Support or Funding InformationD'Youville CollegeThis abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Combination Therapy With Fingolimod and Neural Stem Cells Promotes Functional Myelination in vivo Through a Non-immunomodulatory Mechanism.

Myelination, which occurs predominantly postnatally and continues throughout life, is important for proper neurologic function of the mammalian central nervous system (CNS). We have previously demonstrated that the combination therapy of fingolimod (FTY720) and transplanted neural stem cells (NSCs) had a significantly enhanced therapeutic effect on the chronic stage of experimental autoimmune encephalomyelitis, an animal model of CNS autoimmunity, compared to using either one of them alone. However, reduced disease severity may be secondary to the immunomodulatory effects of FTY720 and NSCs, while whether this therapy directly affects myelinogenesis remains unknown. To investigate this important question, we used three myelination models under minimal or non-inflammatory microenvironments. Our results showed that FTY720 drives NSCs to differentiate into oligodendrocytes and promotes myelination in an ex vivo brain slice culture model, and in the developing CNS of healthy postnatal mice in vivo. Elevated levels of neurotrophic factors, e.g., brain-derived neurotrophic factor and glial cell line-derived neurotrophic factor, were observed in the CNS of the treated infant mice. Further, FTY720 and NSCs efficiently prolonged the survival and improved sensorimotor function of shiverer mice. Together, these data demonstrate a direct effect of FTY720, beyond its known immunomodulatory capacity, in NSC differentiation and myelin development as a novel mechanism underlying its therapeutic effect in demyelinating diseases.