Grafted human-induced pluripotent stem cells-derived oligodendrocyte progenitor cells combined with human umbilical vein endothelial cells contribute to functional recovery following spinal cord injury.

BACKGROUND: Spinal cord injury (SCI) is a devastating disease that causes extensive damage to oligodendrocytes and neurons leading to demyelination and axonal degeneration. In this study, we co-transplanted cell grafts containing oligodendrocyte progenitor cells (OPCs) derived from human-induced pluripotent stem cells (iPSCs) combined with human umbilical vein endothelial cells (HUVECs), which were reported to promote OPCs survival and migration, into rat contusion models to promote functional recovery after SCI. METHODS: OPCs were derived from iPSCs and identified by immunofluorescence at different time points. Functional assays in vitro were performed to evaluate the effect of HUVECs on the proliferation, migration, and survival of OPCs by co-culture and migration assay, as well as on the neuronal axonal growth. A combination of OPCs and HUVECs was transplanted into the rat contusive model. Upon 8 weeks, immunofluorescence staining was performed to test the safety of transplanted cells and to observe the neuronal repairment, myelination, and neural circuit reconstruction at the injured area; also, the functional recovery was assessed by Basso, Beattie, and Bresnahan open-field scale, Ladder climb, SEP, and MEP. Furthermore, the effect of HUVECs on grafts was also determined in vivo. RESULTS: Data showed that HUVECs promote the proliferation, migration, and survival of OPCs both in vitro and in vivo. Furthermore, 8 weeks upon engraftment, the rats with OPCs and HUVECs co-transplantation noticeably facilitated remyelination, enhanced functional connection between the grafts and the host and promoted functional recovery. In addition, compared with the OPCs-alone transplantation, the co-transplantation generated more sensory neurons at the lesion border and significantly improved the sensory functional recovery. CONCLUSIONS: Our study demonstrates that transplantation of OPCs combined with HUVECs significantly enhances both motor and sensory functional recovery after SCI. No significance was observed between OPCs combined with HUVECs group and OPCs-alone group in motor function recovery, while the sensory function recovery was significantly promoted in OPCs combined with HUVECs groups compared with the other two groups. These findings provide novel insights into the field of SCI research.

A2B5-positive oligodendrocyte precursor cell transplantation improves neurological deficits in rats following spinal cord contusion associated with changes in expression of factors involved in the Notch signaling pathway.

BACKGROUND: Oligodendrocyte precursor cells (OPCs) are myelinated glial cells of the central nervous system (CNS), able to regenerate oligodendrocytes and myelin. This study aimed to elucidate the effect of A2B5-positive (A2B5+) OPC transplantation in rats with spinal cord contusion (SCC) and to investigate changes in expression of various factors involved in the Notch signaling pathway after OPC transplantation. METHODS: OPCs were obtained from induced pluripotent stem cells (iPSCs) originating from mouse embryo fibroblasts (MEFs). After identification of iPSCs and iPSC-derived OPCs, A2B5+ OPCs were transplanted into the injured site of rats with SCC one week after SCC insult. Behavioral tests evaluated motor and sensory function 7 days after OPC transplantation. Real-time quantitative polymerase chain reaction (RT-qPCR) determined the expression of various cytokines related to the Notch signaling pathway after OPC transplantation. RESULTS: IPSC-derived OPCs were successfully generated from MEFs, as indicated by positive immunostaining of A2B5, PDGFα and NG2. Further differentiation of OPCs was identified by immunostaining of Olig2, Sox10, Nkx2.2, O4, MBP and GFAP. Importantly, myelin formation was significantly enhanced in the SCC+ OPC group and SCI-induced motor and sensory dysfunction was largely alleviated by A2B5+ OPC transplantation. Expression of factors involved in the Notch signaling pathway (Notch-1, Numb, SHARP1 and NEDD4) was significantly increased after OPC transplantation. CONCLUSIONS: A2B5+ OPC transplantation attenuates motor and sensory dysfunction in SCC rats by promoting myelin formation, which may be associated with change in expression of factors involved in the Notch signaling pathway.

miR-219 overexpressing oligodendrocyte progenitor cells for treating compression spinal cord injury.

Oligodendrocyte progenitor cells (OPCs) transplantation has been considered a promising treatment for spinal cord injury, according to previous studies. Recent research shed light on the importance of microRNA 219 (miR-219) in oligodendrocyte development, so here miR-219-overexpressing OPCs (miR-219 OPCs) were transplanted in animal models of spinal cord injury to evaluate the impact of miR-219 on oligodendrocyte differentiation and functional recovery in vivo. Our findings demonstrate that transplanted cells were distributed in the tissue sections and contributed to reducing the size of cavity in the injury site. Interestingly, miR-219 promoted OPC differentiation into mature oligodendrocyte expressing MBP in vivo whereas in absence of miR-219, less number of cells differentiated into mature oligodendrocytes. An eight week evaluation using the Basso Beattie Bresnahan (BBB) locomotor test confirmed improvement in functional recovery of hind limbs. Overall, this study demonstrated that miR-219 promoted differentiation and maturation of OPCs after transplantation and can be used in cell therapy of spinal cord injury.

Regionally Specific Human Pre-Oligodendrocyte Progenitor Cells Produce Both Oligodendrocytes and Neurons after Transplantation in a Chronically Injured Spinal Cord Rat Model after Glial Scar Ablation.

Chronic spinal cord injury (SCI) is a devastating medical condition. In the acute phase after injury, there is cell loss resulting in chronic axonal damage and loss of sensory and motor function including loss of oligodendrocytes that results in demyelination of axons and further dysfunction. In the chronic phase, the inhibitory environment within the lesion including the glial scar can arrest axonal growth and regeneration and can also potentially affect transplanted cells. We hypothesized that glial scar ablation (GSA) along with cell transplantation may be required as a combinatorial therapy to achieve functional recovery, and therefore we proposed to examine the survival and fate of human induced pluripotent stem cell (iPSC) derived pre-oligodendrocyte progenitor cells (pre-OPCs) transplanted in a model of chronic SCI, whether this was affected by GSA, and whether this combination of treatments would result in functional recovery. In this study, chronically injured athymic nude (ATN) rats were allocated to one of three treatment groups: GSA only, pre-OPCs only, or GSA+pre-OPCs. We found that human iPSC derived pre-OPCs were multi-potent and retained the ability to differentiate into mainly oligodendrocytes or neurons when transplanted into the chronically injured spinal cords of rats. Twelve weeks after cell transplantation, we observed that more of the transplanted cells differentiated into oligodendrocytes when the glial scar was ablated compared with no GSA. Further, we also observed that a higher percentage of transplanted cells differentiated into V2a interneurons and motor neurons in the pre-OPCs only group when compared with GSA+pre-OPCs. This suggests that the local environment created by ablation of the glial scar may have a significant effect on the fate of cells transplanted into the injury site.

Reactive oligodendrocyte progenitor cells (re-)myelinate the regenerating zebrafish spinal cord.

Spinal cord injury (SCI) results in loss of neurons, oligodendrocytes and myelin sheaths, all of which are not efficiently restored. The scarcity of oligodendrocytes in the lesion site impairs re-myelination of spared fibres, which leaves axons denuded, impedes signal transduction and contributes to permanent functional deficits. In contrast to mammals, zebrafish can functionally regenerate the spinal cord. Yet, little is known about oligodendroglial lineage biology and re-myelination capacity after SCI in a regeneration-permissive context. Here, we report that, in adult zebrafish, SCI results in axonal, oligodendrocyte and myelin sheath loss. We find that OPCs, the oligodendrocyte progenitor cells, survive the injury, enter a reactive state, proliferate and differentiate into oligodendrocytes. Concomitantly, the oligodendrocyte population is re-established to pre-injury levels within 2 weeks. Transcriptional profiling revealed that reactive OPCs upregulate the expression of several myelination-related genes. Interestingly, global reduction of axonal tracts and partial re-myelination, relative to pre-injury levels, persist at later stages of regeneration, yet are sufficient for functional recovery. Taken together, these findings imply that, in the zebrafish spinal cord, OPCs replace lost oligodendrocytes and, thus, re-establish myelination during regeneration.

Oligodendrocyte precursor cells transplantation protects blood-brain barrier in a mouse model of brain ischemia via Wnt/ß-catenin signaling.

Blood-brain barrier damage is a critical pathological feature of ischemic stroke. Oligodendrocyte precursor cells are involved in maintaining blood-brain barrier integrity during the development. However, whether oligodendrocyte precursor cell could sustain blood-brain barrier permeability during ischemic brain injury is unknown. Here, we investigate whether oligodendrocyte precursor cell transplantation protects blood-brain barrier integrity and promotes ischemic stroke recovery. Adult male ICR mice (n = 68) underwent 90 min transient middle cerebral artery occlusion. After ischemic assault, these mice received stereotactic injection of oligodendrocyte precursor cells (6 × 105). Oligodendrocyte precursor cells transplantation alleviated edema and infarct volume, and promoted neurological recovery after ischemic stroke. Oligodendrocyte precursor cells reduced blood-brain barrier leakage via increasing claudin-5, occludin and ß-catenin expression. Administration of ß-catenin inhibitor blocked the beneficial effects of oligodendrocyte precursor cells. Wnt7a protein treatment increased ß-catenin and claudin-5 expression in endothelial cells after oxygen-glucose deprivation, which was similar to the results of the conditioned medium treatment of oligodendrocyte precursor cells on endothelial cells. We demonstrated that oligodendrocyte precursor cells transplantation protected blood-brain barrier in the acute phase of ischemic stroke via activating Wnt/ß-catenin pathway. Our results indicated that oligodendrocyte precursor cells transplantation was a novel approach to the ischemic stroke therapy.

Metformin Restores CNS Remyelination Capacity by Rejuvenating Aged Stem Cells.

The age-related failure to produce oligodendrocytes from oligodendrocyte progenitor cells (OPCs) is associated with irreversible neurodegeneration in multiple sclerosis (MS). Consequently, regenerative approaches have significant potential for treating chronic demyelinating diseases. Here, we show that the differentiation potential of adult rodent OPCs decreases with age. Aged OPCs become unresponsive to pro-differentiation signals, suggesting intrinsic constraints on therapeutic approaches aimed at enhancing OPC differentiation. This decline in functional capacity is associated with hallmarks of cellular aging, including decreased metabolic function and increased DNA damage. Fasting or treatment with metformin can reverse these changes and restore the regenerative capacity of aged OPCs, improving remyelination in aged animals following focal demyelination. Aged OPCs treated with metformin regain responsiveness to pro-differentiation signals, suggesting synergistic effects of rejuvenation and pro-differentiation therapies. These findings provide insight into aging-associated remyelination failure and suggest therapeutic interventions for reversing such declines in chronic disease.

Oligodendrocyte precursors gain Schwann cell-like phenotype and remyelinate axons upon engraftment into peripheral nerves.

The ability to treat large peripheral nerve injuries may be greatly advanced if an accessible source of human myelinating cells is identified, as it overcomes one of the major limitations of acellular or synthetic nerve guides compared with autografts, the gold standard for large defect repair. Methods to derive oligodendrocyte precursor cells (OPCs) from human pluripotent stem cells have advanced to the point where they have been shown capable of myelination and are being evaluated in clinical trials. Here, we test the hypothesis that OPCs can survive and remyelinate axons in the peripheral nervous system during a repair process. Using freshly isolated OPCs from mouse post-natal brains, we engrafted these OPCs into the tibial nerve immediately after it being subjected to cryolesioning. At 1-month postengraftment, we found numerous graft-derived cells that survived in this environment, and many transplanted cells expressed Schwann cell markers such as periaxin and S100ß coexpressed with myelin basic protein, whereas oligodendrocyte markers O4 and Olig2 were virtually absent. Our results demonstrate that OPCs can survive in a peripheral nervous system micro-environment and undergo niche-dependent transdifferentiation into Schwann cell-like cells as has previously been observed in central nervous system focal demyelination models, suggesting that OPCs constitute an accessible source of cells for peripheral nerve cell therapies.

Rapid and Efficient Differentiation of Rodent Neural Stem Cells into Oligodendrocyte Progenitor Cells.

Oligodendrocyte progenitor cells (OPCs) may have beneficial effects in cell replacement therapy of neurodegenerative disease owing to their unique capability to differentiate into myelinogenic oligodendrocytes (OLs) in response to extrinsic signals. Therefore, it is of significance to establish an effective differentiation methodology to generate highly pure OPCs and OLs from some easily accessible stem cell sources. To achieve this goal, in this study, we present a rapid and efficient protocol for oligodendroglial lineage differentiation from mouse neural stem cells (NSCs), rat NSCs, or mouse embryonic stem cell-derived neuroepithelial stem cells. In a defined culture medium containing Smoothened Agonist, basic fibroblast growth factor, and platelet-derived growth factor-AA, OPCs could be generated from the above stem cells over a time course of 4-6 days, achieving a cell purity as high as ∼90%. In particular, these derived OPCs showed high expandability and could further differentiate into myelin basic protein-positive OLs within 3 days or alternatively into glial fibrillary acidic protein-positive astrocytes within 7 days. Furthermore, transplantation of rodent NSC-derived OPCs into injured spinal cord indicated that it is a feasible strategy to treat spinal cord injury. Our results suggest a differentiation strategy for robust production of OPCs and OLs from rodent stem cells, which could provide an abundant OPC source for spinal cord injury.

Immunosuppressive mechanisms for stem cell transplant survival in spinal cord injury.

Spinal cord injury (SCI) has been associated with a dismal prognosis-recovery is not expected, and the most standard interventions have been temporizing measures that do little to mitigate the extent of damage. While advances in surgical and medical techniques have certainly improved this outlook, limitations in functional recovery continue to impede clinically significant improvements. These limitations are dependent on evolving immunological mechanisms that shape the cellular environment at the site of SCI. In this review, we examine these mechanisms, identify relevant cellular components, and discuss emerging treatments in stem cell grafts and adjuvant immunosuppressants that target these pathways. As the field advances, we expect that stem cell grafts and these adjuvant treatments will significantly shift therapeutic approaches to acute SCI with the potential for more promising outcomes.

Stem cell therapies for acute spinal cord injury in humans: a review.

Recent advances in stem cell biology present significant opportunities to advance clinical applications of stem cell-based therapies for spinal cord injury (SCI). In this review, the authors critically analyze the basic science and translational evidence that supports the use of various stem cell sources, including induced pluripotent stem cells, oligodendrocyte precursor cells, and mesenchymal stem cells. They subsequently explore recent advances in stem cell biology and discuss ongoing clinical translation efforts, including combinatorial strategies utilizing scaffolds, biogels, and growth factors to augment stem cell survival, function, and engraftment. Finally, the authors discuss the evolution of stem cell therapies for SCI by providing an overview of completed (n = 18) and ongoing (n = 9) clinical trials.

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, S100ß, 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.

Xeno-free culture for generation of forebrain oligodendrocyte precursor cells from human pluripotent stem cells.

Oligodendrocytes (OLs) show heterogeneous properties that depend on their location in the central nervous system (CNS). In this regard, the investigation of oligodendrocyte precursor cells (OPCs) derived from human pluripotent stem cells (hPSCs) should be reconsidered, particularly in cases of brain-predominant disorders for which brain-derived OPCs are more appropriate than spinal cord-derived OPCs. Furthermore, animal-derived components are responsible for culture variability in the derivation and complicate clinical translation. In the present study, we established a xeno-free system to induce forebrain OPCs from hPSCs. We induced human forebrain neural stem cells (NSCs) on Laminin 511-E8 and directed the differentiation to the developmental pathway for forebrain OLs with SHH and FGF signaling. OPCs were characterized by the expression of OLIG2, NKX2.2, SOX10, and PDGFRA, and subsequent maturation into O4+ cells. In vitro characterization showed that >85% of the forebrain OPCs (O4+ ) underwent maturation into OLs (MBP+ ) 3 weeks after mitogen removal. Upon intracranial transplantation, the OPCs survived, dispersed in the corpus callosum, and matured into (GSTπ+ ) OLs in the host brains 3 months after transplantation. These findings suggest our xeno-free induction of forebrain OPCs from hPSCs could accelerate clinical translation for brain-specific disorders.

Paired Related Homeobox Protein 1 Regulates Quiescence in Human Oligodendrocyte Progenitors.

Human oligodendrocyte progenitor cells (hOPCs) persist into adulthood as an abundant precursor population capable of division and differentiation. The transcriptional mechanisms that regulate hOPC homeostasis remain poorly defined. Herein, we identify paired related homeobox protein 1 (PRRX1) in primary PDGFαR+ hOPCs. We show that enforced PRRX1 expression results in reversible G1/0 arrest. While both PRRX1 splice variants reduce hOPC proliferation, only PRRX1a abrogates migration. hOPC engraftment into hypomyelinated shiverer/rag2 mouse brain is severely impaired by PRRX1a, characterized by reduced cell proliferation and migration. PRRX1 induces a gene expression signature characteristic of stem cell quiescence. Both IFN-γ and BMP signaling upregulate PRRX1 and induce quiescence. PRRX1 knockdown modulates IFN-γ-induced quiescence. In mouse brain, PRRX1 mRNA was detected in non-dividing OPCs and is upregulated in OPCs following demyelination. Together, these data identify PRRX1 as a regulator of quiescence in hOPCs and as a potential regulator of pathological quiescence.

Improvement by Human Oligodendrocyte Progenitor Cells of Neurobehavioral Disorders in an Experimental Model of Neonatal Periventricular Leukomalacia.

The effects of human oligodendrocyte progenitor (F3.olig2) cells on improving neurobehavioral deficits were investigated in an experimental model of periventricular leukomalacia (PVL). Seven-day-old male rats were subjected to hypoxia-ischemia-lipopolysaccharide injection (HIL), and intracerebroventricularly transplanted with F3.olig2 (4 × 105 cells/rat) once at post-natal day (PND) 10 or repeatedly at PND10, 17, 27, and 37. Neurobehavioral disorders were evaluated at PND14, 20, 30, and 40 via cylinder test, locomotor activity, and rotarod performance, and cognitive function was evaluated at PND41-45 through passive avoidance and Morris water-maze performances. F3.olig2 cells recovered the rate of use of the forelimb contralateral to the injured brain, improved locomotor activity, and restored rotarod performance of PVL animals; in addition, marked improvement of learning and memory function was seen. It was confirmed that transplanted F3·olig2 cells migrated to injured areas, matured to oligodendrocytes expressing myelin basic protein (MBP), and markedly attenuated the loss of host MBP in the corpus callosum. The results indicate that the transplanted F3.olig2 cells restored neurobehavioral functions by preventing axonal demyelination, and that human oligodendrocyte progenitor cells could be a candidate for cell therapy of perinatal hypoxic-ischemic and infectious brain injuries including PVL and cerebral palsy.

Efficacy of Oligodendrocyte Progenitor Cell Transplantation in Rat Models with Traumatic Thoracic Spinal Cord Injury: A Systematic Review and Meta-Analysis.

Spinal cord injury (SCI) is a devastating disease that results in severe motor, sensory, and autonomic dysfunction, for which there are currently no available treatments. Subsequent to the primary mechanical damage, progressive secondary damage further exacerbates the functional deficit. Demyelination may play an important role in the pathogenesis of SCI. Oligodendrocyte progenitor cells (OPCs) are considered a candidate cellular treatment approach for SCI attributable to their unique potential. Here, we conducted a systematic review and meta-analysis to assess the efficacy of OPC transplantation in rat models with traumatic thoracic SCI, and 17 studies (20 experiments, 402 rats) were identified. The Basso, Beattie, Bresnahan (BBB) locomotor rating score, latency and amplitude of motor evoked potential (MEP), percentage of myelinated axons, and cavity size were used as outcome measurements. The pooled results of the meta-analysis in different subgroups (based on cell dose or sex) both suggested that the BBB score of the OPC group was significantly improved compared to that of the control group 2, 4, and 8 weeks after transplantation. Whereas the amplitude of MEP was not significantly different, the latency of MEP was significantly shorter compared to that of the control group 4 weeks after transplantation. The percentage of myelinated axons increased significantly in the OPC group compared to that of the control group after OPC transplantation. Area measurements across groups revealed a significant reduction in cavity size in the OPC-treated groups compared to the control group. In conclusion, OPC transplantation provided considerable beneficial effects after traumatic SCI.

[Neuroprotective effects of oligodendrocyte precursor cells on white matter damage in preterm infants].

White matter damage, characterized by demyelination due to the damage of oligodendrocyte precursor cells (OPCs), is the most common type of brain damage in preterm infants. Survivors are often subject to long-term neurodevelopmental sequelae because of the lack of effective treatment. In recent years, it has been found that cell transplantation has the potential for the treatment of white matter damage. OPCs are frequently used cells in cell transplantation therapy. With abilities of migration and myelinization, OPCs are the best seed cells for the treatment of white matter damage. Several studies have found that OPCs may not only replace impaired cells to reconstruct the structure and function of white matter, but also inhibit neuronal apoptosis, promote the proliferation of endogenous neural stem cells, and enhance the repairment of the blood-brain barrier. However, the clinical application of OPC transplantation therapy faces many challenges, such as the effectiveness, risk of tumorigenesis and immune rejection. With reference to these studies, this article reviewed the development of myelination, the obtainment of OPCs, the therapeutic mechanism as well as application research, and analyzed the current challenges of OPC transplantation, in order to provide a new direction for clinical treatment of white matter damage in preterm infants.

Oligodendrocyte precursor cell transplantation promotes functional recovery following contusive spinal cord injury in rats and is associated with altered microRNA expression.

It has been reported that oligodendrocyte precursor cells (OPCs) may be used to treat contusive spinal cord injury (SCC), and may alter microRNA (miRNA/miR) expression following SCC in rats. However, the association between miRNA expression and the treatment of rats with SCC with OPC transplantation remain unclear. The present study transplanted OPCs into the spinal cord of rats with SCC and subsequently used the Basso, Beattie and Bresnahan (BBB) score to assess the functional recovery and pain scores. An miRNA assay was performed to detect differentially expressed miRNAs in the spinal cord of SCC rats transplanted with OPCs, compared with SCC rats transplanted with medium. Quantitative polymerase chain reaction was used to verify significantly altered miRNA expression levels. The results demonstrated that OPC transplantation was able to improve motor recovery and relieve mechanical allodynia in rats with SCC. In addition, through a miRNA assay, 45 differentially expressed miRNAs (40 upregulated miRNAs and 5 downregulated miRNAs) were detected in the spinal cord of rats in the OPC group compared with in the Medium group. Differentially expressed miRNAs were identified according to the following criteria: Fold change >2 and P<0.05. Furthermore, quantitative polymerase chain reaction was used to verify the most highly upregulated (miR­375­3p and miR­1­3p) and downregulated (miR­363­3p, miR­449a­5p and miR­3074) spinal cord miRNAs that were identified in the miRNA assay. In addition, a bioinformatics analysis of these miRNAs indicated that miR­375 and miR­1 may act primarily to inhibit cell proliferation and apoptosis via transcriptional and translational regulation, whereas miR­363, miR­449a and miR­3074 may act primarily to inhibit cell proliferation and neuronal differentiation through transcriptional regulation. These results suggested that OPC transplantation may promote functional recovery of rats with SCC, which may be associated with the expression of various miRNAs in the spinal cord, including miR­375­3p, miR­1­3p, miR­363­3p, miR­449a­5p and miR­3074.