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Tissue engineering is a promising strategy to repair spinal cord injury (SCI). However, a bioscaffold with mechanical properties that match those of the pathological spinal cord tissue and a pro-regenerative matrix that allows robust neurogenesis for overcoming post-SCI scar formation has yet to be developed. Here, we report that a mechanically enhanced decellularized spinal cord (DSC) scaffold with a thin poly (lactic-co-glycolic acid) (PLGA) outer shell may fulfill the requirements for effective in situ neuroengineering after SCI. Using chemical extraction and electrospinning methods, we successfully constructed PLGA thin shell-ensheathed DSC scaffolds (PLGA-DSC scaffolds) in a way that removed major inhibitory components while preserving the permissive matrix. The DSCs exhibited good cytocompatibility with neural stem cells (NSCs) and significantly enhanced their differentiation toward neurons in vitro. Due to the mechanical reinforcement, the implanted PLGA-DSC scaffolds showed markedly increased resilience to infiltration by myofibroblasts and the deposition of dense collagen matrix, thereby creating a neurogenic niche favorable for the targeted migration, residence and neuronal differentiation of endogenous NSCs after SCI. Furthermore, PLGA-DSC presented a mild immunogenic property but prominent ability to polarize macrophages from the M1 phenotype to the M2 phenotype, leading to significant tissue regeneration and functional restoration after SCI. Taken together, the results demonstrate that the mechanically matched PLGA-DSC scaffolds show promise for effective tissue repair after SCI., (Copyright © 2021. Published by Elsevier Ltd.)

The inability of adult mammals to recover function lost after severe spinal cord injury (SCI) has been known for millennia and is mainly attributed to a failure of brain-derived nerve fiber regeneration across the lesion. Potential approaches to re-establishing locomotor function rely on neuronal relays to reconnect the segregated neural networks of the spinal cord. Intense research over the past 30 years has focused on endogenous and exogenous neuronal relays, but progress has been slow and the results often controversial. Treatments with stem cell-derived neuronal relays alone or together with functional electrical stimulation offer the possibility of improved repair of neuronal networks. In this review, we focus on approaches to recovery of motor function in paralyzed patients after severe SCI based on novel therapies such as implantation of stem cell-derived neuronal relays and functional electrical stimulation. Recent research progress offers hope that SCI patients will one day be able to recover motor function and sensory perception., (Copyright © 2021 The Authors. Published by Elsevier Ltd.. All rights reserved.)

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., (Copyright © 2018 The Authors. Published by Elsevier Inc. All rights reserved.)

Severe spinal cord injury (SCI) causes loss of neural connectivity and permanent functional deficits. Re-establishment of new neuronal relay circuits after SCI is therefore of paramount importance. The present study tested our hypothesis if co-culture of neurotrophin-3 (NT-3) gene-modified Schwann cells (SCs, NT-3-SCs) and TrkC (NT-3 receptor) gene-modified neural stem cells (NSCs, TrkC-NSCs) in a gelatin sponge scaffold could construct a tissue engineering neural network for re-establishing an anatomical neuronal relay after rat spinal cord transection. Eight weeks after transplantation, the neural network created a favorable microenvironment for axonal regeneration and for survival and synaptogenesis of NSC-derived neurons. Biotin conjugates of cholera toxin B subunit (b-CTB, a transneuronal tracer) was injected into the crushed sciatic nerve to label spinal cord neurons. Remarkably, not only ascending and descending nerve fibers, but also propriospinal neurons, made contacts with b-CTB positive NSC-derived neurons. Moreover, b-CTB positive NSC-derived neurons extended their axons making contacts with the motor neurons located in areas caudal to the injury/graft site of spinal cord. Further study showed that NT-3/TrkC interactions activated the PI3K/AKT/mTOR pathway and PI3K/AKT/CREB pathway affecting synaptogenesis of NSC-derived neurons. Together, our findings suggest that NT-3-mediated TrkC signaling plays an essential role in constructing a tissue engineering neural network thus representing a promising avenue for effective exogenous neuronal relay-based treatment for SCI., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

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.

Biological materials combined with genetically-modified neural stem cells (NSCs) are candidate therapy targeting spinal cord injury (SCI). Based on our previous studies, here we performed gelatin sponge (GS) scaffold seeded with neurotrophin-3 (NT-3) and its receptor TrkC gene modifying NSCs for repairing SCI. Eight weeks later, compared with other groups, neurofilament-200 and 5-hydroxytryptamine positive nerve fibers were more in the injury site of the N+T-NSCs group. Immunofluorescence staining showed the grafted NSCs could differentiate into microtubule associated protein (Map2), postsynaptic density (PSD95), and mouse oligodendrocyte special protein (MOSP) positive cells. The percentage of the Map2, PSD95, and MOSP positive cells in the N+T-NSCs group was higher than the other groups. Immuno-electron microscopy showed the grafted NSCs making contact with each other in the injury site. Behavioral analysis indicated the recovery of hindlimbs locomotion was better in the groups receiving cell transplant, the best recovery was found in the N+T-NSCs group. Electrophysiology revealed the amplitude of cortical motor evoked potentials was increased significantly in the N+T-NSCs group, but the latency remained long. These findings suggest the GS scaffold containing genetically-modified NSCs may bridge the injury site, promote axon regeneration and partial functional recovery in SCI rats., (© 2014 Wiley Periodicals, Inc.)

Remyelination remains a challenging issue in spinal cord injury (SCI). In the present study, we cocultured Schwann cells (SCs) and neural stem cells (NSCs) with overexpression of neurotrophin-3 (NT-3) and its high affinity receptor tyrosine kinase receptor type 3 (TrkC), respectively, in a gelatin sponge (GS) scaffold. This was aimed to generate a tissue-engineered neural scaffold and to investigate whether it could enhance myelination after a complete T10 spinal cord transection in adult rats. Indeed, many NT-3 overexpressing SCs (NT-3-SCs) in the GS scaffold assumed the formation of myelin. More strikingly, a higher incidence of NSCs overexpressing TrkC differentiating toward myelinating cells was induced by NT-3-SCs. By transmission electron microscopy, the myelin sheath showed distinct multilayered lamellae formed by the seeded cells. Eighth week after the scaffold was transplanted, some myelin basic protein (MBP)-positive processes were observed within the transplantation area. Remarkably, certain segments of myelin derived from NSC-derived myelinating cells and NT-3-SCs were found to ensheath axons. In conclusion, we show here that transplantation of the GS scaffold promotes exogenous NSC-derived myelinating cells and SCs to form myelins in the injury/transplantation area of spinal cord. These findings thus provide a neurohistological basis for the future application or transplantation using GS neural scaffold to repair SCI.

Rebuilding structures that can bridge the injury gap and enable signal connection remains a challenging issue in spinal cord injury. We sought to determine if genetically enhanced expression of TrkC in neural stem cells (NSCs) and neurotrophin-3 in Schwann cells (SCs) co-cultured in a gelatin sponge scaffold could constitute a neural network, and whether it could act as a relay to rebuilt signal connection after spinal cord transection. Indeed, many NSCs in the scaffold assumed neuronal features including formation of synapses. By whole-cell patch clamp, the synapses associated with NSC-derived neurons were excitable. Grafting of the scaffold with differentiating NSCs + SCs into rats with a segment of the spinal cord removed had resulted in a significant functional recovery of the paralyzed hind-limbs. Remarkably, the NSC-derived neurons formed new synaptic contacts suggesting that the scaffold can form a relay for conduction of signals through the injury gap of spinal cord., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

Effectively bridging the lesion gap is still an unmet demand for spinal cord repair. In the present study, we tested our hypothesis if cograft of Schwann cells (SCs) and neural stem cells (NSCs) with genetically enhanced expression of neurotrophin-3 (NT-3) and its high affinity receptor TrkC, respectively, could strengthen neural repair through increased NSC survival and neuronal differentiation at the epicenter after complete T10 spinal cord transection in adult rats. Transplantation of NT-3-SCs + TrkC-NSCs in Gelfoam (1 × 10(6)/implant/rat; n = 10) into the lesion gap immediately following injury results in significantly improved relay of the cortical motor evoked potential (CMEP) and cortical somatosensory evoked potential (CSEP) as well as ameliorated hindlimb deficits, relative to controls (treated with LacZ-SCs + LacZ-NSCs, NT-3-SCs + NSCs, NSCs alone, or lesion only; n = 10/group). Further analyses demonstrate that NT-3-SCs + TrkC-NSCs cografting augments levels of neuronal differentiation of NSCs, synaptogenesis (including inhibitory/type II-like synapses) and myelin formation of SCs, in addition to neuroprotection and outgrowth of serotonergic fibers in the lesioned spinal cord. Compared with controls, the treated spinal cords also show elevated expression of laminin, a pro-neurogenic factor, and decreased presence of chondroitin sulfate proteoglycans, major inhibitors of axonal growth and neuroplasticity. Together, our data suggests that coimplantation of neurologically compatible cells with compensatorily overexpressed therapeutic genes may constitute a valuable approach to study, and/or develop therapies for spinal cord injury (SCI)., (Published by Elsevier Ltd.)

Delivery of cellular and/or trophic factors to the site of injury may promote neural repair or axonal regeneration and return of function after spinal cord injury. Engineered scaffolds provide a platform to deliver therapeutic cells and neurotrophic molecules. To explore therapeutic potential of engineered neural tissue, we generated an artificial neural construct in vitro, and transplanted this construct into a completely transected spinal cord of adult rats. Two months later, behavioral analysis showed that the locomotion recovery was significantly improved compared with control animals. Immunoreactivity against microtubule associated protein 2 (Map2) and postsynaptic density 95 (PSD95) demonstrated that grafted cells had a higher survival rate and were able to differentiate toward neuronal phenotype with ability to form synapse-like structure at the injury site; this was also observed under the electron microscope. Immunostaining of neurofilament-200 (NF-200) showed that the number of nerve fibers regrowing into the injury site in full treatment group was much higher than that seen in other groups. Furthermore, Nissl staining revealed that host neuron survival rate was significantly increased in rats with full treatments. However, there were no biotin dextran amine (BDA) anterograde tracing fibers crossing through the injury site, suggesting the limited ability of corticospinal tract axonal regeneration. Taken together, although our artificial neural construct permits grafted cells to differentiate into neuronal phenotype, synaptogenesis, axonal regeneration and partial locomotor function recovery, the limited capacity for corticospinal tract axonal regeneration may affect its potential therapy in spinal cord injury., (Copyright © 2011 Elsevier B.V. All rights reserved.)