Ferulic Acid Improves Functional Recovery after Acute Spinal Cord Injury in Rats by Inducing Hypoxia to Inhibit microRNA-590 and Elevate Vascular Endothelial Growth Factor Expressions.
ABSTRACT: Spinal cord injury (SCI) is the leading cause of paralysis, disability and even death in severe cases, and neural stem cells (NSCs) transplant has been employed for repairing SCI. Ferulic acid (FA) is able to promote neurogenesis in various stem cell therapies. We aimed to investigate the effect of FA on NSC transplant therapy, and the underlying mechanism, in improving functional recovery in SCI rat model. A rat model of SCI was established, which then received transplant of NSCs with or without FA pre-treatment. Functional recovery of the SCI rats was then evaluated, in terms of spinal cord water content, myeloperoxidase activity and behavioral assessments. Effect of FA in inducing hypoxia in NSCs was also assessed, followed by identifying the hypoxic regulated microRNA and the subsequent target gene. Transplant of FA pre-treated NSCs improved functional recovery of SCI rats to a more significant extent than NSCs without FA pre-treatment. The beneficial effects of FA in repairing SCI was mediated by inducing hypoxia in NSCs, which in turn inhibited microRNA-590 to elevate vascular endothelial growth factor expression. Our findings support the clinical potential of FA in improving efficacy of NSC transplant therapy for treatment of SCI.
Project description:The body's capacity to restore damaged neural networks in the injured CNS is severely limited. Although various treatment regimens can partially alleviate spinal cord injury (SCI), the mechanisms responsible for symptomatic improvement remain elusive. Here, using a mouse model of SCI, we have shown that transplantation of neural stem cells (NSCs) together with administration of valproic acid (VPA), a known antiepileptic and histone deacetylase inhibitor, dramatically enhanced the restoration of hind limb function. VPA treatment promoted the differentiation of transplanted NSCs into neurons rather than glial cells. Transsynaptic anterograde corticospinal tract tracing revealed that transplant-derived neurons reconstructed broken neuronal circuits, and electron microscopic analysis revealed that the transplant-derived neurons both received and sent synaptic connections to endogenous neurons. Ablation of the transplanted cells abolished the recovery of hind limb motor function, confirming that NSC transplantation directly contributed to restored motor function. These findings raise the possibility that epigenetic status in transplanted NSCs can be manipulated to provide effective treatment for SCI.
Project description:Neural stem cell (NSC) transplantation is a major focus of current research for treatment of spinal cord injury (SCI). However, it is very important to promote the survival and differentiation of NSCs into myelinating oligodendrocytes (OLs). In this study, myelin basic protein-activated T (MBP-T) cells were passively immunized to improve the SCI microenvironment. Olig2-overexpressing NSCs were infected with a lentivirus carrying the enhanced green fluorescent protein (GFP) reporter gene to generate Olig2-GFP-NSCs that were transplanted into the injured site to differentiate into OLs. Transferred MBP-T cells infiltrated the injured spinal cord, produced neurotrophic factors, and induced the differentiation of resident microglia and/or infiltrating blood monocytes into an "alternatively activated" anti-inflammatory macrophage phenotype by producing interleukin-13. As a result, the survival of transplanted NSCs increased fivefold in MBP-T cell-transferred rats compared with that of the vehicle-treated control. In addition, the differentiation of MBP-positive OLs increased 12-fold in Olig2-GFP-NSC-transplanted rats compared with that of GFP-NSC-transplanted controls. In the MBP-T cell and Olig2-GFP-NSC combined group, the number of OL-remyelinated axons significantly increased compared with those of all other groups. However, a significant decrease in spinal cord lesion volume and an increase in spared myelin and behavioral recovery were observed in Olig2-NSC- and NSC-transplanted MBP-T cell groups. Collectively, these results suggest that MBP-T cell adoptive immunotherapy combined with NSC transplantation has a synergistic effect on histological and behavioral improvement after traumatic SCI. Although Olig2 overexpression enhances OL differentiation and myelination, the effect on functional recovery may be surpassed by MBP-T cells.
Project description:: Neural stem cells (NSCs) from embryonic or fetal/adult tissue sources have shown considerable promise in regenerative strategies for traumatic spinal cord injury (SCI). However, there are limitations with their use related to the availability, immunogenicity, and uncertainty of the mechanisms involved. To address these issues, definitive NSCs derived from induced pluripotent stem (iPS) cells generated using a nonviral, piggyBac transposon approach, were investigated. Committed NSCs were generated from iPS cells using a free-floating neurosphere methodology previously described by our laboratory. To delineate the mechanism of action, specifically the role of exogenous myelination, NSCs derived from wildtype (wt) and nonmyelinating Shiverer (shi) iPS cell lines were used following thoracic SCI with subacute intraspinal transplantation. Behavioral, histological, and electrophysiological outcomes were analyzed to assess the effectiveness of this treatment. The wt- and shi-iPS-NSCs were validated and shown to be equivalent except in myelination capacity. Both iPS-NSC lines successfully integrated into the injured spinal cord and predominantly differentiated to oligodendrocytes, but only the wt-iPS-NSC treatment resulted in a functional benefit. The wt-iPS-dNSCs, which exhibited the capacity for remyelination, significantly improved neurobehavioral function (Basso Mouse Scale and CatWalk), histological outcomes, and electrophysiological measures of axonal function (sucrose gap analysis) compared with the nonmyelinating iPS-dNSCs and cell-free controls. In summary, we demonstrated that iPS cells can generate translationally relevant NSCs for applications in SCI. Although NSCs have a diverse range of functions in the injured spinal cord, remyelination is the predominant mechanism of recovery following thoracic SCI.Gain-of-function/loss-of-function techniques were used to examine the mechanistic importance of graft-derived remyelination following thoracic spinal cord injury (SCI). The novel findings of this study include the first use of neural stem cells (NSCs) from induced pluripotent stem cells (iPSCs) derived using the clonal neurosphere expansion conditions, for the treatment of SCI, the first characterization and in vivo application of iPSCs from Shiverer mouse fibroblasts, and the first evidence of the importance of remyelination by pluripotent-sourced NSCs for SCI repair and regeneration.
Project description:The number of elderly patients with spinal cord injury (SCI) is increasing worldwide, representing a serious burden for both the affected patients and the community. Previous studies have demonstrated that neural stem cell (NSC) transplantation is an effective treatment for SCI in young animals. Here we show that NSC transplantation is as effective in aged mice as it is in young mice, even though aged mice exhibit more severe neurological deficits after SCI. NSCs grafted into aged mice exhibited better survival than those grafted into young mice. Furthermore, we show that the neurotrophic factor HGF plays a key role in the enhanced functional recovery after NSC transplantation observed in aged mice with SCI. The unexpected results of the present study suggest that NSC transplantation is a potential therapeutic modality for SCI, even in elderly patients.
Project description:Neural stem cell (NSC) grafts have demonstrated significant effects in animal models of spinal cord injury (SCI), yet their clinical translation remains challenging. Significant evidence suggests that the supporting matrix of NSC grafts has a crucial role in regulating NSC effects. Here we demonstrate that grafts based on porous collagen-based scaffolds (PCSs), similar to biomaterials utilized clinically in induced regeneration, can deliver and protect embryonic NSCs at SCI sites, leading to significant improvement in locomotion recovery in an experimental mouse SCI model, so that 12 weeks post-injury locomotion performance of implanted animals does not statistically differ from that of uninjured control animals. NSC-seeded PCS grafts can modulate key processes required to induce regeneration in SCI lesions including enhancing NSC neuronal differentiation and functional integration in vivo, enabling robust axonal elongation, and reducing astrogliosis. Our findings suggest that the efficacy and translational potential of emerging NSC-based SCI therapies could be enhanced by delivering NSC via scaffolds derived from well-characterized clinically proven PCS.
Project description:<h4>Background</h4>Spinal cord injury (SCI) is a common disease that results in motor and sensory disorders and even lifelong paralysis. The transplantation of stem cells, such as embryonic stem cells (ESCs), induced pluripotent stem cells (iPSCs), mesenchymal stem cells (MSCs), or subsequently generated stem/progenitor cells, is predicted to be a promising treatment for SCI. In this study, we aimed to investigate effect of human iPSC-derived neural stem cells (hiPSC-NSCs) and umbilical cord-derived MSCs (huMSCs) in a mouse model of acute SCI.<h4>Methods</h4>Acute SCI mice model were established and were randomly treated as phosphate-buffered saline (PBS) (control group), repaired with 1?×?10<sup>5</sup> hiPSC-NSCs (NSC group), and 1?×?10<sup>5</sup> huMSCs (MSC group), respectively, in a total of 54 mice (n?=?18 each). Hind limb motor function was evaluated in open-field tests using the Basso Mouse Scale (BMS) at days post-operation (dpo) 1, 3, 5, and 7 after spinal cord injury, and weekly thereafter. Spinal cord and serum samples were harvested at dpo 7, 14, and 21. Haematoxylin-eosin (H&E) staining and Masson staining were used to evaluate the morphological changes and fibrosis area. The differentiation of the transplanted cells in vivo was evaluated with immunohistochemical staining.<h4>Results</h4>The hiPSC-NSC-treated group presented a significantly smaller glial fibrillary acidic protein (GFAP) positive area than MSC-treated mice at all time points. Additionally, MSC-transplanted mice had a similar GFAP+ area to mice receiving PBS. At dpo 14, the immunostained hiPSC-NSCs were positive for SRY-related high-mobility-group (HMG)-box protein-2 (SOX2). Furthermore, the transplanted hiPSC-NSCs differentiated into GFAP-positive astrocytes and beta-III tubulin-positive neurons, whereas the transplanted huMSCs differentiated into GFAP-positive astrocytes. In addition, hiPSC-NSC transplantation reduced fibrosis formation and the inflammation level. Compared with the control or huMSC transplanted group, the group with transplantation of hiPSC-NSCs exhibited significantly improved behaviours, particularly limb coordination.<h4>Conclusions</h4>HiPSC-NSCs promote functional recovery in mice with acute SCI by replacing missing neurons and attenuating fibrosis, glial scar formation, and inflammation.
Project description:BACKGROUND Spinal cord injury (SCI) causes a rapid loss of motor neurons, leading to weakness and paralysis. Transplantation of neural stem cells is known to restore the neuronal activity but is inefficient due to limited regenerative capability and low rate of survival. There has been an emphasis on the use of growth factors along with neural stem cells (NSCs) to enhance the neuronal recovery. Transplantation of recombinant NSCs with vascular endothelial growth factor (VEGF) might promote neuronal repair. This effect might be attributed to the reduced transient receptor potential vanilloid 1 (TRPV1) expression following transplantation. MATERIAL AND METHODS NSCs were cultured from the embryos of Sprague-Dawley rats (E12.5). Four group of rats (n=10, each) were subjected to SCI and allowed to recover for 1 week. Recombinant VEGF-NSCs, normal NSCs and PBS were intrathecally administered to the rats. VEGF and TRPV-1 expression at mRNA and protein level was evaluated. ELISA was performed to determine the release of neurotrophic factors after the transplantation. Motor neurons and axons were counted and the motor behavioral outcome was assessed using the rota-rod test. RESULTS VEGF-NSC transgene transplantation resulted in an enhanced neuronal repair and motor behavioral outcome compared to the normal NSCs transplanted group. VEGF-NSCs increased the release of neurotrophic factors and reduced the expression of TRPV1. CONCLUSIONS Recombinant VEGF-NSCs transplantation following SCI is more efficacious compared to normal NSC transplantation. This might also be related to a reduced pain in the process of recovery due to reduced TRPV1 expression.
Project description:Neural stem cells (NSCs) differentiate into both neurons and glia, and strategies using human NSCs have the potential to restore function following spinal cord injury (SCI). However, the time period of maturation for human NSCs in adult injured CNS is not well defined, posing fundamental questions about the design and implementation of NSC-based therapies. This work assessed human H9 NSCs that were implanted into sites of SCI in immunodeficient rats over a period of 1.5 years. Notably, grafts showed evidence of continued maturation over the entire assessment period. Markers of neuronal maturity were first expressed 3 months after grafting. However, neurogenesis, neuronal pruning, and neuronal enlargement continued over the next year, while total graft size remained stable over time. Axons emerged early from grafts in very high numbers, and half of these projections persisted by 1.5 years. Mature astrocyte markers first appeared after 6 months, while more mature oligodendrocyte markers were not present until 1 year after grafting. Astrocytes slowly migrated from grafts. Notably, functional recovery began more than 1 year after grafting. Thus, human NSCs retain an intrinsic human rate of maturation, despite implantation into the injured rodent spinal cord, yet they support delayed functional recovery, a finding of great importance in planning human clinical trials.
Project description:Survival and migration of transplanted neural stem cells (NSCs) are prerequisites for therapeutic benefits in spinal cord injury. We have shown that survival of NSC grafts declines after transplantation into the injured spinal cord, and that combining treadmill training (TMT) enhances NSC survival via insulin-like growth factor-1 (IGF-1). Here, we aimed to obtain genetic evidence that IGF-1 signaling in the transplanted NSCs determines the beneficial effects of TMT. We transplanted NSCs heterozygous (+/-) for Igf1r, the gene encoding IGF-1 receptor, into the mouse spinal cord after injury, with or without combining TMT. We analyzed the influence of genotype and TMT on locomotor recovery and survival and migration of NSC grafts. In vitro experiments were performed to examine the potential roles of IGF-1 signaling in the migratory ability of NSCs. Mice receiving +/- NSC grafts showed impaired locomotor recovery compared with those receiving wild-type (+/+) NSCs. Locomotor improvement by TMT was more pronounced with +/+ grafts. Deficiency of one allele of Igf1r significantly reduced survival and migration of the transplanted NSCs. Although TMT did not significantly influence NSC survival, it substantially enhanced the extent of migration for only +/+ NSCs. Cultured neurospheres exhibited dynamic motility with cytoplasmic protrusions, which was regulated by IGF-1 signaling. IGF-1 signaling in transplanted NSCs may be essential in regulating their survival and migration. Furthermore, TMT may promote NSC graft-mediated locomotor recovery via activation of IGF-1 signaling in transplanted NSCs. Dynamic NSC motility via IGF-1 signaling may be the cellular basis for the TMT-induced enhancement of migration.
Project description:Objective:To judge the efficacies of neural stem cell (NSC) transplantation on functional recovery following contusion spinal cord injuries (SCIs). Data sources:Studies in which NSCs were transplanted into a clinically relevant, standardized rat model of contusion SCI were identified by searching the PubMed, Embase and Cochrane databases, and the extracted data were analyzed by Stata 14.0. Data selection:Inclusion criteria were that NSCs were used in in vivo animal studies to treat contusion SCIs and that behavioral assessment of locomotor functional recovery was performed using the Basso, Beattie, and Bresnahan lo-comotor rating scale. Exclusion criteria included a follow-up of less than 4 weeks and the lack of control groups. Outcome measures:The restoration of motor function was assessed by the Basso, Beattie, and Bresnahan locomotor rating scale. Results:We identified 1756 non-duplicated papers by searching the aforementioned electronic databases, and 30 full-text articles met the inclusion criteria. A total of 37 studies reported in the 30 articles were included in the meta-analysis. The meta-analysis results showed that transplanted NSCs could improve the motor function recovery of rats following contusion SCIs, to a moderate extent (pooled standardized mean difference (SMD) = 0.73; 95% confidence interval (CI): 0.47-1.00; P < 0.001). NSCs obtained from different donor species (rat: SMD = 0.74; 95% CI: 0.36-1.13; human: SMD = 0.78; 95% CI: 0.31-1.25), at different donor ages (fetal: SMD = 0.67; 95% CI: 0.43-0.92; adult: SMD = 0.86; 95% CI: 0.50-1.22) and from different origins (brain-derived: SMD = 0.59; 95% CI: 0.27-0.91; spinal cord-derived: SMD = 0.51; 95% CI: 0.22-0.79) had similar efficacies on improved functional recovery; however, adult induced pluripotent stem cell-derived NSCs showed no significant efficacies. Furthermore, the use of higher doses of transplanted NSCs or the administration of immunosuppressive agents did not promote better locomotor function recovery (SMD = 0.45; 95% CI: 0.21-0.70). However, shorter periods between the contusion induction and the NSC transplantation showed slightly higher efficacies (acute: SMD = 1.22; 95% CI: 0.81-1.63; subacute: SMD = 0.75; 95% CI: 0.42-1.09). For chronic injuries, NSC implantation did not significantly improve functional recovery (SMD = 0.25; 95% CI: -0.16 to 0.65). Conclusion:NSC transplantation alone appears to be a positive yet limited method for the treatment of contusion SCIs.