Genetic susceptibility and causal pathway analysis of eye disorders coexisting in multiple sclerosis.

Introduction: The comorbidity of optic neuritis with multiple sclerosis has been well recognized. However, the causal association between multiple sclerosis and optic neuritis, as well as other eye disorders, remains incompletely understood. To address these gaps, we investigated the genetically relationship between multiple sclerosis and eye disorders, and explored potential drugs. Methods: In order to elucidate the genetic susceptibility and causal links between multiple sclerosis and eye disorders, we performed two-sample Mendelian randomization analyses to examine the causality between multiple sclerosis and eye disorders. Additionally, causal single-nucleotide polymorphisms were annotated and searched for expression quantitative trait loci data. Pathway enrichment analysis was performed to identify the possible mechanisms responsible for the eye disorders coexisting with multiple sclerosis. Potential therapeutic chemicals were also explored using the Cytoscape. Results: Mendelian randomization analysis revealed that multiple sclerosis increased the incidence of optic neuritis while reducing the likelihood of concurrent of cataract and macular degeneration. Gene Ontology enrichment analysis implicated that lymphocyte proliferation, activation and antigen processing as potential contributors to the pathogenesis of eye disorders coexisting with multiple sclerosis. Furthermore, pharmaceutical agents traditionally employed for allograft rejection exhibited promising therapeutic potential for the eye disorders coexisting with multiple sclerosis. Discussion: Multiple sclerosis genetically contributes to the development of optic neuritis while mitigating the concurrent occurrence of cataract and macular degeneration. Further research is needed to validate these findings and explore additional mechanisms underlying the comorbidity of multiple sclerosis and eye disorders.

Single-cell RNA-sequencing analysis reveals enhanced non-canonical neurotrophic factor signaling in the subacute phase of traumatic brain injury.

BACKGROUND: Traumatic brain injury (TBI) is a leading cause of long-term disability in young adults and induces complex neuropathological processes. Cellular autonomous and intercellular changes during the subacute phase contribute substantially to the neuropathology of TBI. However, the underlying mechanisms remain elusive. In this study, we explored the dysregulated cellular signaling during the subacute phase of TBI. METHODS: Single-cell RNA-sequencing data (GSE160763) of TBI were analyzed to explore the cell-cell communication in the subacute phase of TBI. Upregulated neurotrophic factor signaling was validated in a mouse model of TBI. Primary cell cultures and cell lines were used as in vitro models to examine the potential mechanisms affecting signaling. RESULTS: Single-cell RNA-sequencing analysis revealed that microglia and astrocytes were the most affected cells during the subacute phase of TBI. Cell-cell communication analysis demonstrated that signaling mediated by the non-canonical neurotrophic factors midkine (MDK), pleiotrophin (PTN), and prosaposin (PSAP) in the microglia/astrocytes was upregulated in the subacute phase of TBI. Time-course profiling showed that MDK, PTN, and PSAP expression was primarily upregulated in the subacute phase of TBI, and astrocytes were the major source of MDK and PTN after TBI. In vitro studies revealed that the expression of MDK, PTN, and PSAP in astrocytes was enhanced by activated microglia. Moreover, MDK and PTN promoted the proliferation of neural progenitors derived from human-induced pluripotent stem cells (iPSCs) and neurite growth in iPSC-derived neurons, whereas PSAP exclusively stimulated neurite growth. CONCLUSION: The non-canonical neurotrophic factors MDK, PTN, and PSAP were upregulated in the subacute phase of TBI and played a crucial role in neuroregeneration.

Stem Cell Factor and Granulocyte Colony-Stimulating Factor Promote Remyelination in the Chronic Phase of Severe Traumatic Brain Injury.

Severe traumatic brain injury (TBI) causes long-term disability and death in young adults. White matter is vulnerable to TBI damage. Demyelination is a major pathological change of white matter injury after TBI. Demyelination, which is characterized by myelin sheath disruption and oligodendrocyte cell death, leads to long-term neurological function deficits. Stem cell factor (SCF) and granulocyte colony-stimulating factor (G-CSF) treatments have shown neuroprotective and neurorestorative effects in the subacute and chronic phases of experimental TBI. Our previous study has revealed that combined SCF and G-CSF treatment (SCF + G-CSF) enhances myelin repair in the chronic phase of TBI. However, the long-term effect and mechanism of SCF + G-CSF-enhanced myelin repair remain unclear. In this study, we uncovered persistent and progressive myelin loss in the chronic phase of severe TBI. SCF + G-CSF treatment in the chronic phase of severe TBI enhanced remyelination in the ipsilateral external capsule and striatum. The SCF + G-CSF-enhanced myelin repair is positively correlated with the proliferation of oligodendrocyte progenitor cells in the subventricular zone. These findings reveal the therapeutic potential of SCF + G-CSF in myelin repair in the chronic phase of severe TBI and shed light on the mechanism underlying SCF + G-CSF-enhanced remyelination in chronic TBI.

Stem cell factor and granulocyte colony-stimulating factor promote remyelination in the chronic phase of severe traumatic brain injury.

Severe traumatic brain injury (TBI) causes long-term disability and death in young adults. White matter is vulnerable to TBI damage. Demyelination is a major pathological change of white matter injury after TBI. Demyelination which is characterized by myelin sheath disruption and oligodendrocyte cell death leads to long-term neurological function deficits. Stem cell factor (SCF) and granulocyte colonyâ€"stimulating factor (G-CSF) treatments have shown neuroprotective and neurorestorative effects in the subacute and chronic phases of experimental TBI. Our previous study has revealed that combined SCF and G-CSF treatment (SCF+G-CSF) enhances myelin repair in the chronic phase of TBI. However, the long-term effect and mechanism of SCF+G-CSF-enhanced myelin repair remain unclear. In this study, we uncovered persistent and progressive myelin loss in the chronic phase of severe TBI. SCF+G-CSF treatment in the chronic phase of severe TBI enhanced remyelination in the ipsilateral external capsule and striatum. The SCF+G-CSF-enhanced myelin repair is positively correlated with the proliferation of oligodendrocyte progenitor cells in the subventricular zone. These findings reveal the therapeutic potential of SCF+G-CSF in myelin repair in the chronic phase of severe TBI and shed light on the mechanism underlying SCF+G-CSF-enhanced remyelination in chronic TBI.

SCF + G-CSF treatment in the chronic phase of severe TBI enhances axonal sprouting in the spinal cord and synaptic pruning in the hippocampus.

Traumatic brain injury (TBI) is a major cause of long-term disability in young adults. An evidence-based treatment for TBI recovery, especially in the chronic phase, is not yet available. Using a severe TBI mouse model, we demonstrate that the neurorestorative efficacy of repeated treatments with stem cell factor (SCF) and granulocyte colony-stimulating factor (G-CSF) (SCF + G-CSF) in the chronic phase is superior to SCF + G-CSF single treatment. SCF + G-CSF treatment initiated at 3 months post-TBI enhances contralesional corticospinal tract sprouting into the denervated side of the cervical spinal cord and re-balances the TBI-induced overgrown synapses in the hippocampus by enhancing microglial function of synaptic pruning. These neurorestorative changes are associated with SCF + G-CSF-improved somatosensory-motor function and spatial learning. In the chronic phase of TBI, severe TBI-caused microglial degeneration in the cortex and hippocampus is ameliorated by SCF + G-CSF treatment. These findings reveal the therapeutic potential and possible mechanism of SCF + G-CSF treatment in brain repair during the chronic phase of severe TBI.

Brain-derived CCR5 Contributes to Neuroprotection and Brain Repair after Experimental Stroke.

Chemokine (C-C motif) receptor 5 (CCR5) is expressed not only in the immune cells but also in cerebral cells such as neurons, glia, and vascular cells. Stroke triggers high expression of CCR5 in the brain. However, the role of CCR5 in stroke remains unclear. In this study, using bone marrow chimeras we have determined the involvement of brain-derived or bone marrow-derived CCR5 in neuroprotection and brain repair after experimental stroke. CCR5-/- mice that received either wild-type (WT) or CCR5-/- bone marrow transplantation showed larger infarction sizes than the WT mice that received either WT or CCR5-/- bone marrow transplantation in both the acute (48h) and subacute (2 months) phases after cerebral cortical ischemia, suggesting that the lack of CCR5 in the brain leads to severe brain damage after stroke. However, the lack of CCR5 in the bone marrow-derived cells did not affect infarction size. The impairments of somatosensory-motor function and motor coordination were exacerbated in the mice lacking CCR5 in the brain. At 2 months post-stroke, increased degenerative neurons, decreased dendrites and synapses, decreased Iba1+ microglia/ macrophages, reduced myelination and CNPase+ oligodendrocytes in the peri-infarct cortex were observed in the mice lacking CCR5 in the brain. These pathological changes are significantly correlated with the increased infarction size and exacerbated neurological deficits. These data suggest that brain-derived CCR5 plays a key role in neuroprotection and brain repair in the subacute phase of stroke. This study reveals a novel role of CCR5 in stroke, which sheds new light on post-stroke pathomechanism.

The contribution of stem cell factor and granulocyte colony-stimulating factor in reducing neurodegeneration and promoting neurostructure network reorganization after traumatic brain injury.

Traumatic brain injury (TBI) is a major cause of death and disability in young adults worldwide. TBI-induced long-term cognitive deficits represent a growing clinical problem. Stem cell factor (SCF) and granulocyte colony-stimulating factor (G-CSF) are involved in neuroprotection and neuronal plasticity. However, the knowledge concerning reparative efficacy of SCF + G-CSF treatment in post-acute TBI recovery remains incomplete. This study aims to determine the efficacy of SCF + G-CSF on post-acute TBI recovery in young adult mice. The controlled cortical impact model of TBI was used for inducing a severe damage in the motor cortex of the right hemisphere in 8-week-old male C57BL mice. SCF + G-CSF treatment was initiated 3 weeks after induction of TBI. Severe TBI led to persistent motor functional deficits (Rota-Rod test) and impaired spatial learning function (water maze test). SCF + G-CSF treatment significantly improved the severe TBI-impaired spatial learning function 6 weeks after treatment. TBI also caused significant increases of Fluoro-Jade C positive degenerating neurons in bilateral frontal cortex, striatum and hippocampus, and significant reductions in MAP2+ apical dendrites and overgrowth of SMI312+ axons in peri-TBI cavity frontal cortex and in the ipsilateral hippocampal CA1 at 24 weeks post-TBI. SCF + G-CSF treatment significantly reduced TBI-induced neurodegeneration in the contralateral frontal cortex and hippocampal CA1, increased MAP2+ apical dendrites in the peri-TBI cavity frontal cortex, and prevented TBI-induced axonal overgrowth in both the peri-TBI cavity frontal cortex and ipsilateral hippocampal CA1.These findings reveal a novel pathology of axonal overgrowth after severe TBI and demonstrate a therapeutic potential of SCF + G-CSF in ameliorating severe TBI-induced long-term neuronal pathology, neurostructural network malformation, and impairments in spatial learning.

Long-term beneficial effects of hematopoietic growth factors on brain repair in the chronic phase of severe traumatic brain injury.

Severe traumatic brain injury (TBI) is the major cause of long-term, even life-long disability and cognitive impairments in young adults. The lack of therapeutic approaches to improve recovery in the chronic phase of severe TBI is a big challenge to the medical research field. Using a single severe TBI model in young adult mice, this study examined the restorative efficacy of two hematopoietic growth factors, stem cell factor (SCF) and granulocyte colony-stimulating factor (G-CSF), on brain repair in the chronic phase of TBI. SCF and G-CSF alone or combination (SCF + G-CSF) treatment was administered at 3 months post-TBI. Functional recovery was evaluated by neurobehavioral tests during the period of 21 weeks after treatment. Neuropathology was examined 22 weeks after treatment. We observed that severe TBI caused persistent impairments in spatial learning/memory and somatosensory-motor function, long-term and widespread neuropathology, including dendritic reduction, decrease and overgrowth of axons, over-generated excitatory synapses, and demyelination in the cortex, hippocampus and striatum. SCF, G-CSF, and SCF + G-CSF treatments ameliorated severe TBI-induced widespread neuropathology. SCF + G-CSF treatment showed superior efficacy in improving long-term functional outcome, enhancing neural plasticity, rebalancing neural structure networks disturbed by severe TBI, and promoting remyelination. These novel findings demonstrate the therapeutic potential of SCF and G-CSF in enhancing recovery in the chronic phase of severe TBI .

Stem Cell Factor in Combination With Granulocyte Colony-Stimulating Factor Protects the Brain From Capillary Thrombosis-Induced Ischemic Neuron Loss in a Mouse Model of CADASIL.

Cerebral autosomal dominant arteriopathy with subcortical infarct and leukoencephalopathy (CADASIL) is a Notch3 mutation-induced cerebral small vessel disease, leading to recurrent ischemic stroke and vascular dementia. There is currently no treatment that can stop or delay CADASIL progression. We have demonstrated the efficacy of treatment with combined stem cell factor (SCF) and granulocyte colony-stimulating factor (G-CSF) (SCF+G-CSF) in reducing cerebral small vessel thrombosis in a TgNotch3R90C mouse model of CADASIL. However, it remains unknown whether SCF+G-CSF treatment protects neurons from microvascular thrombosis-induced ischemic damage. Using bone marrow transplantation to track thrombosis, we observed that capillary thrombosis was widely distributed in the cortex, striatum and hippocampus of 22-month-old TgNotch3R90C mice. However, the capillary thrombosis mainly occurred in the cortex. Neuron loss was seen in the area next to the thrombotic capillaries, and severe neuron loss was found in the areas adjacent to the thrombotic capillaries with bifurcations. SCF+G-CSF repeated treatment significantly attenuated neuron loss in the areas next to the thrombotic capillaries in the cortex of the 22-month-old TgNotch3R90C mice. Neuron loss caused by capillary thrombosis in the cerebral cortex may play a crucial role in the pathogenesis of CADASIL. SCF+G-CSF treatment ameliorates the capillary thrombosis-induced ischemic neuron loss in TgNotch3R90C mice. This study provides new insight into the understanding of CADASIL progression and therapeutic potential of SCF+G-CSF in neuroprotection under microvascular ischemia in CADASIL.

S100 Calcium-Binding Protein A9 Knockout Contributes to Neuroprotection and Functional Improvement after Traumatic Brain Injury.

S100 calcium-binding protein A9 (S100a9), a proinflammatory protein, has been shown to be involved in the development of neuroinflammatory disorders and neurodegenerative diseases. Upregulation of S100a9 in the brain during acute brain injury has been proposed to be associated with acute neuroinflammation. However, it remains unclear whether eliminating S100a9 expression will show beneficial outcomes after traumatic brain injury (TBI). Using S100a9 knockout mice, this study has demonstrated that S100a9 deletion ameliorates post-TBI anxiety, improves TBI-impaired motor and cognitive function, reduces lesion size, prevents perilesional neuron loss and neurodegeneration, diminishes neuroinflammation and TBI-induced neurogenesis, and enhances perilesional expression of neuroplasticity protein. These findings suggest that S100a9 plays a detrimental role in TBI. Genetic deletion of S100a9 enhances neuroprotection and improves functional outcome after TBI. This study sheds light on the pathological involvement of S100a9 in TBI, which would provide a new therapeutic target to minimize TBI-induced brain damage.

Stem cell factor and granulocyte colony-stimulating factor promote brain repair and improve cognitive function through VEGF-A in a mouse model of CADASIL.

Cerebral autosomal dominant arteriopathy with subcortical infarct and leukoencephalopathy (CADASIL) is a cerebral small vascular disease caused by NOTCH3 gene mutation in vascular smooth muscle cells (VSMCs), leading to ischemic stroke and vascular dementia. To date, the pathogenesis of CADASIL remains poorly understood, and there is no treatment that can slow the progression of CADASIL. Using a transgenic mouse model of CADASIL (TgNotch3R90C), this study reveals novel findings for understanding CADASIL pathogenesis that decreased cerebral vascular endothelial growth factor (VEGF/VEGF-A) is linked to reduced cerebral blood vessel density. Reduced endothelial cell (EC) proliferation and angiogenesis are seen in TgNotch3R90C mouse brain-isolated ECs. Decreased dendrites, axons, and synapses in the somatosensory and motor cortex layer 2/3 and in the hippocampal CA1, and reduced neurogenesis in both the subventricular zone and subgranular zone occur in 15-month-old TgNotch3R90C mice. These reductions in neuron structures, synapses, and neurogenesis are significantly correlated to decreased cerebral vasculature in the corresponding areas. Impaired spatial learning and memory in TgNotch3R90C mice are significantly correlated with the reduced cerebral vasculature, neuron structures, and synapses. Repeated treatment of stem cell factor and granulocyte colony-stimulating factor (SCF+G-CSF) at 9 and 10 months of age improves cognitive function, increases cerebral VEGF/VEGF-A, restores cerebral vasculature, and enhances regeneration of neuronal structures, synaptogenesis and neurogenesis in TgNotch3R90C mice. Pretreatment with Avastin, an angiogenesis inhibitor by neutralizing VEGF-A, completely eliminates the SCF+G-CSF-enhanced cognitive function, vascular and neuronal structure regeneration, synaptogenesis and neurogenesis in TgNotch3R90C mice. SCF+G-CSF-enhanced EC proliferation and angiogenesis in TgNotch3R90C mouse brain-isolated ECs are also blocked by Avastin pretreatment. These data suggest that SCF+G-CSF treatment may repair Notch3R90C mutation-damaged brain through the VEGF-A-mediated angiogenesis. This study provides novel insight into the involvement of VEGF/VEGF-A in the pathogenesis of CADASIL and sheds light on the mechanism underlying the SCF+G-CSF-enhanced brain repair in CADASIL.

Electroacupuncture Facilitates the Integration of Neural Stem Cell-Derived Neural Network with Transected Rat Spinal Cord.

The hostile environment of an injured spinal cord makes it challenging to achieve higher viability in a grafted tissue-engineered neural network used to reconstruct the spinal cord circuit. Here, we investigate whether cell survival and synaptic transmission within an NT-3 and TRKC gene-overexpressing neural stem cell-derived neural network scaffold (NN) transplanted into transected spinal cord could be promoted by electroacupuncture (EA) through improving the microenvironment. Our results showed that EA facilitated the cell survival, neuronal differentiation, and synapse formation of a transplanted NN. Pseudorabies virus tracing demonstrated that EA strengthened synaptic integration of the transplanted NN with the host neural circuit. The combination therapy also promoted axonal regeneration, spinal conductivity, and functional recovery. The findings highlight EA as a potential and safe supplementary therapeutic strategy to reinforce the survival and synaptogenesis of a transplanted NN as a neuronal relay to bridge the two severed ends of an injured spinal cord.

Stem Cell Factor in Combination with Granulocyte Colony-Stimulating Factor reduces Cerebral Capillary Thrombosis in a Mouse Model of CADASIL.

Cerebral autosomal dominant arteriopathy with subcortical infarcts and leucoencephalopathy (CADASIL) is a cerebral small vascular disease caused by NOTCH3 mutation-induced vascular smooth muscle cell (VSMC) degeneration, leading to ischemic stroke and vascular dementia. Our previous study has demonstrated that repeated treatment with a combination of stem cell factor (SCF) and granulocyte colony-stimulating factor (G-CSF) reduces VSMC degeneration and cerebral endothelial cell (EC) damage and improves cognitive function in a mouse model of CADASIL (TgNotch3R90C). This study aimed to determine whether cerebral thrombosis occurs in TgNotch3R90C mice and whether repeated SCF+G-CSF treatment reduces cerebral thrombosis in TgNotch3R90C mice. Using the approaches of bone marrow transplantation to track bone marrow-derived cells and confocal imaging, we observed bone marrow-derived blood cell occlusion in cerebral small vessels and capillaries (thrombosis). Most thrombosis occurred in the cerebral capillaries (93% of total occluded vessels), and the thrombosis showed an increased frequency in the regions of capillary bifurcation. Degenerated capillary ECs were seen inside and surrounding the thrombosis, and the bone marrow-derived ECs were also found next to the thrombosis. IgG extravasation was seen in and next to the areas of thrombosis. SCF+G-CSF treatment significantly reduced cerebral capillary thrombosis and IgG extravasation. These data suggest that the EC damage is associated with thrombosis and blood-brain barrier leakage in the cerebral capillaries under the CADASIL-like condition, whereas SCF+G-CSF treatment diminishes these pathological alterations. This study provides new insight into the involvement of cerebral capillary thrombosis in the development of CADASIL and potential approaches to reduce the thrombosis, which may restrict the pathological progression of CADASIL.

Neurotrophin-3 released from implant of tissue-engineered fibroin scaffolds inhibits inflammation, enhances nerve fiber regeneration, and improves motor function in canine spinal cord injury.

Spinal cord injury (SCI) normally results in cell death, scarring, cavitation, inhibitory molecules release, etc., which are regarded as a huge obstacle to reconnect the injured neuronal circuits because of the lack of effective stimulus. In this study, a functional gelatin sponge scaffold was used to inhibit local inflammation, enhance nerve fiber regeneration, and improve neural conduction in the canine. This scaffold had good porosity and modified with neurotrophin-3 (NT-3)/fibroin complex, which showed sustained release in vitro. After the scaffold was transplanted into canine spinal cord hemisection model, hindlimb movement, and neural conduction were improved evidently. Migrating host cells, newly formed neurons with associated synaptic structures together with functional blood vessels with intact endothelium in the regenerating tissue were identified. Taken together, the results demonstrated that using bioactive scaffold could establish effective microenvironment stimuli for endogenous regeneration, providing a potential and practical strategy for treatment of spinal cord injury. © 2018 The Authors Journal of Biomedical Materials Research Part A Published by Wiley Periodicals, Inc. J Biomed Mater Res Part A: 106A: 2158-2170, 2018.

Perineurium-like sheath derived from long-term surviving mesenchymal stem cells confers nerve protection to the injured spinal cord.

The functional multipotency enables mesenchymal stem cells (MSCs) promising translational potentials in treating spinal cord injury (SCI). Yet the fate of MSCs grafted into the injured spinal cord has not been fully elucidated even in preclinical studies, rendering concerns of their safety and genuine efficacy. Here we used a rat spinal cord transection model to evaluate the cell fate of allograft bone marrow derived MSCs. With the application of immunosuppressant, donor cells, delivered by biocompatible scaffold, survived up to 8 weeks post-grafting. Discernible tubes formed by MSCs were observed beginning 2 weeks after transplantation and they dominated the morphological features of implanted MSCs at 8 weeks post-grafting. The results of immunocytochemistry and transmission electron microscopy displayed the formation of perineurium-like sheath by donor cells, which, in a manner comparable to the perineurium in peripheral nerve, enwrapped host myelins and axons. The MSC-derived perineurium-like sheath secreted a group of trophic factors and permissive extracellular matrix, and served as a physical and chemical barrier to insulate the inner nerve fibers from ambient oxidative insults by the secretion of soluble antioxidant, superoxide dismutase-3 (SOD3). As a result, many intact regenerating axons were preserved in the injury/graft site following the forming of perineurium-like sheath. A parallel study utilizing a good manufacturing practice (GMP) grade human umbilical cord-derived MSCs or allogenic MSCs in an acute contusive/compressive SCI model exhibited a similar perineurium-like sheath formed by surviving donor cells in rat spinal cord at 3 weeks post-grafting. The present study for the first time provides an unambiguous morphological evidence of perineurium-like sheath formed by transplanted MSCs and a novel therapeutic mechanism of MSCs in treating SCI.

Traumatic Brain Injury: Current Treatment Strategies and Future Endeavors.

Traumatic brain injury (TBI) presents in various forms ranging from mild alterations of consciousness to an unrelenting comatose state and death. In the most severe form of TBI, the entirety of the brain is affected by a diffuse type of injury and swelling. Treatment modalities vary extensively based on the severity of the injury and range from daily cognitive therapy sessions to radical surgery such as bilateral decompressive craniectomies. Guidelines have been set forth regarding the optimal management of TBI, but they must be taken in context of the situation and cannot be used in every individual circumstance. In this review article, we have summarized the current status of treatment for TBI in both clinical practice and basic research. We have put forth a brief overview of the various subtypes of traumatic injuries, optimal medical management, and both the noninvasive and invasive monitoring modalities, in addition to the surgical interventions necessary in particular instances. We have overviewed the main achievements in searching for therapeutic strategies of TBI in basic science. We have also discussed the future direction for developing TBI treatment from an experimental perspective.

Autocrine fibronectin from differentiating mesenchymal stem cells induces the neurite elongation in vitro and promotes nerve fiber regeneration in transected spinal cord injury.

Extracellular matrix (ECM) expression is temporally and spatially regulated during the development of stem cells. We reported previously that fibronectin (FN) secreted by bone marrow mesenchymal stem cells (MSCs) was deposited on the surface of gelatin sponge (GS) soon after culture. In this study, we aimed to assess the function of accumulated FN on neuronal differentiating MSCs as induced by Schwann cells (SCs) in three dimensional transwell co-culture system. The expression pattern and amount of FN of differentiating MSCs was examined by immunofluorescence, Western blot and immunoelectron microscopy. The results showed that FN accumulated inside GS scaffold, although its mRNA expression in MSCs was progressively decreased during neural induction. MSC-derived neuron-like cells showed spindle-shaped cell body and long extending processes on FN-decorated scaffold surface. However, after blocking of FN function by application of monoclonal antibodies, neuron-like cells showed flattened cell body with short and thick neurites, together with decreased expression of integrin ß1. In vivo transplantation study revealed that autocrine FN significantly facilitated endogenous nerve fiber regeneration in spinal cord transection model. Taken together, the present results showed that FN secreted by MSCs in the early stage accumulated on the GS scaffold and promoted the neurite elongation of neuronal differentiating MSCs as well as nerve fiber regeneration after spinal cord injury. This suggests that autocrine FN has a dynamic influence on MSCs in a three dimensional culture system and its potential application for treatment of traumatic spinal cord injury. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 104A: 1902-1911, 2016.

Donor mesenchymal stem cell-derived neural-like cells transdifferentiate into myelin-forming cells and promote axon regeneration in rat spinal cord transection.

INTRODUCTION: Severe spinal cord injury often causes temporary or permanent damages in strength, sensation, or autonomic functions below the site of the injury. So far, there is still no effective treatment for spinal cord injury. Mesenchymal stem cells (MSCs) have been used to repair injured spinal cord as an effective strategy. However, the low neural differentiation frequency of MSCs has limited its application. The present study attempted to explore whether the grafted MSC-derived neural-like cells in a gelatin sponge (GS) scaffold could maintain neural features or transdifferentiate into myelin-forming cells in the transected spinal cord. METHODS: We constructed an engineered tissue by co-seeding of MSCs with genetically enhanced expression of neurotrophin-3 (NT-3) and its high-affinity receptor tropomyosin receptor kinase C (TrkC) separately into a three-dimensional GS scaffold to promote the MSCs differentiating into neural-like cells and transplanted it into the gap of a completely transected rat spinal cord. The rats received extensive post-operation care, including cyclosporin A administrated once daily for 2 months. RESULTS: MSCs modified genetically could differentiate into neural-like cells in the MN + MT (NT-3-MSCs + TrKC-MSCs) group 14 days after culture in the GS scaffold. However, after the MSC-derived neural-like cells were transplanted into the injury site of spinal cord, some of them appeared to lose the neural phenotypes and instead transdifferentiated into myelin-forming cells at 8 weeks. In the latter, the MSC-derived myelin-forming cells established myelin sheaths associated with the host regenerating axons. And the injured host neurons were rescued, and axon regeneration was induced by grafted MSCs modified genetically. In addition, the cortical motor evoked potential and hindlimb locomotion were significantly ameliorated in the rat spinal cord transected in the MN + MT group compared with the GS and MSC groups. CONCLUSION: Grafted MSC-derived neural-like cells in the GS scaffold can transdifferentiate into myelin-forming cells in the completely transected rat spinal cord.

Integration of donor mesenchymal stem cell-derived neuron-like cells into host neural network after rat spinal cord transection.

Functional deficits following spinal cord injury (SCI) primarily attribute to loss of neural connectivity. We therefore tested if novel tissue engineering approaches could enable neural network repair that facilitates functional recovery after spinal cord transection (SCT). Rat bone marrow-derived mesenchymal stem cells (MSCs), genetically engineered to overexpress TrkC, receptor of neurotrophin-3 (NT-3), were pre-differentiated into cells carrying neuronal features via co-culture with NT-3 overproducing Schwann cells in 3-dimensional gelatin sponge (GS) scaffold for 14 days in vitro. Intra-GS formation of MSC assemblies emulating neural network (MSC-GS) were verified morphologically via electron microscopy (EM) and functionally by whole-cell patch clamp recording of spontaneous post-synaptic currents. The differentiated MSCs still partially maintained prototypic property with the expression of some mesodermal cytokines. MSC-GS or GS was then grafted acutely into a 2 mm-wide transection gap in the T9-T10 spinal cord segments of adult rats. Eight weeks later, hindlimb function of the MSC-GS-treated SCT rats was significantly improved relative to controls receiving the GS or lesion only as indicated by BBB score. The MSC-GS transplantation also significantly recovered cortical motor evoked potential (CMEP). Histologically, MSC-derived neuron-like cells maintained their synapse-like structures in vivo; they additionally formed similar connections with host neurites (i.e., mostly serotonergic fibers plus a few corticospinal axons; validated by double-labeled immuno-EM). Moreover, motor cortex electrical stimulation triggered c-fos expression in the grafted and lumbar spinal cord cells of the treated rats only. Our data suggest that MSC-derived neuron-like cells resulting from NT-3-TrkC-induced differentiation can partially integrate into transected spinal cord and this strategy should be further investigated for reconstructing disrupted neural circuits.