Spinal Cord Transplants Research Articles

Gene Corrected Spinal Muscular Atrophy-Induced Pluripotent Stem Cells and Motoneuron as a Model and Cell Source for Transplantation (IN8-2.002)

Objective: To describe the correction of human spinal muscular atrophy (SMA)-induced pluripotent stem cells (iPSCs) and motoneurons using targeted gene correction with single-stranded oligonucleotides. Background Spinal muscular atrophy (SMA) is among the most common genetic neurological diseases causing infant mortality. Reprogramming adult human cells to induced pluripotent stem cells (iPSCs) allows obtaining patient-specific cells possible. However, the use of SMA iPSCs will require their genetic correction in a manner that is compatible with clinical applications. Design/Methods: We generated iPSCs from fibroblasts from a patient with SMA and his father using a non-viral method. Cells were nucleofected with oriP/EBNA1 vectors encoding six reprogramming factors. We used SMN2 sequence-specific ODNs to direct the exchange of a T to C at position +6 of exon 7, thus converting SMN2 into SMN1 in the SMA-iPSCs. The iPSCs were fully characterized and differentiated using a protocol to promote motoneuron commitment. The phenotype of cells was tested by morphological, gene expression, and protein analysis. iPSC-purified motoneurons were transplanted into the spinal cords of SMA mice. Histochemical and neuropathological analyses were conducted. Survival and neuromuscular function were investigated. Results: SMA corrected cell lines contained no exogenous sequences and appeared indistinguishable from healthy iPSCs. Non-viral SMA-iPSC-derived motoneurons reproduced disease-specific features (reductions in cell number, cell size, and axon length) while corrected SMA-specific-iPSCs gave rise to phenotypically rescued motoneurons in vitro and in vivo after transplantation in SMA spinal cord. Different splicing profiles, detected in vitro by microarray analysis in SMA motoneurons compared to wild-type, were normalized after the genetic correction. Transplantation of wild-type and corrected SMA motoneurons extended lifespan (>50%) and ameliorated the phenotype of SMA mice significantly more than SMA motoneurons and untreated animals. Conclusions: These results offer proof-of-concept that generating patient-specific corrected iPSCs and motoneurons free of exogenous elements may be possible, with potential for research and clinical applications. Disclosure: Dr. Corti has nothing to disclose. Dr. Nizzardo has nothing to disclose. Dr. Simone has nothing to disclose. Dr. Falcone has nothing to disclose. Dr. Nardini has nothing to disclose. Dr. Ronchi has nothing to disclose. Dr. Donadoni has nothing to disclose. Dr. Salani has nothing to disclose. Dr. Riboldi has nothing to disclose. Dr. Menozzi has nothing to disclose. Dr. Bonaglia has nothing to disclose. Dr. Magri has nothing to disclose. Dr. Bresolin has nothing to disclose. Dr. Comi has received research support from Telethon Italy and SMA Europe.

Human Glial-Restricted Progenitor Transplantation into Cervical Spinal Cord of the SOD1G93A Mouse Model of ALS

Cellular abnormalities are not limited to motor neurons in amyotrophic lateral sclerosis (ALS). There are numerous observations of astrocyte dysfunction in both humans with ALS and in SOD1G93A rodents, a widely studied ALS model. The present study therapeutically targeted astrocyte replacement in this model via transplantation of human Glial-Restricted Progenitors (hGRPs), lineage-restricted progenitors derived from human fetal neural tissue. Our previous findings demonstrated that transplantation of rodent-derived GRPs into cervical spinal cord ventral gray matter (in order to target therapy to diaphragmatic function) resulted in therapeutic efficacy in the SOD1G93A rat. Those findings demonstrated the feasibility and efficacy of transplantation-based astrocyte replacement for ALS, and also show that targeted multi-segmental cell delivery to cervical spinal cord is a promising therapeutic strategy, particularly because of its relevance to addressing respiratory compromise associated with ALS. The present study investigated the safety and in vivo survival, distribution, differentiation, and potential efficacy of hGRPs in the SOD1G93A mouse. hGRP transplants robustly survived and migrated in both gray and white matter and differentiated into astrocytes in SOD1G93A mice spinal cord, despite ongoing disease progression. However, cervical spinal cord transplants did not result in motor neuron protection or any therapeutic benefits on functional outcome measures. This study provides an in vivo characterization of this glial progenitor cell and provides a foundation for understanding their capacity for survival, integration within host tissues, differentiation into glial subtypes, migration, and lack of toxicity or tumor formation.

Olfactory Mucosal Transplantation After Spinal Cord Injury Improves Voiding Efficiency by Suppressing Detrusor-Sphincter Dyssynergia in Rats

Several recent studies showed that olfactory mucosal transplantation after spinal cord injury promotes extensive regeneration of the injured spinal cord. We examined the efficacy of olfactory mucosal transplantation for bladder dysfunction after spinal cord injury in rats. In adult female rats the Th9-10 spinal cord was completely transected, followed by olfactory mucosal transplantation or gelatin sponge filling as the control. Each group was examined by cystometrogram and external urethral sphincter electromyogram. Calcitonin gene-related peptide and growth associated protein 43 double positive expression in the L6/S1 dorsal horn was evaluated by immunohistochemistry. Transplant sites were examined by immunohistochemistry with antibodies against neurofilament M and neuronal class III beta-tubulin. On cystometrogram voiding efficiency was significantly higher in the transplantation group than in controls. On external urethral sphincter electromyogram with simultaneous cystometrogram the transplantation group showed a larger ratio of interburst silent periods to burst activity duration and a greater number of high frequency oscillations. In the transplantation group calcitonin gene-related peptide and growth associated protein 43 double positive expression in the L6/S1 dorsal horn was less dense than in controls. The transplantation group showed strong neurofilament M and neuronal class III beta-tubulin expression at the transplant site. Olfactory mucosal transplantation after spinal cord injury weakened external urethral sphincter excessive bursting and increased the urethral opening to improve voiding efficiency. Olfactory mucosal transplantation may modify emergence of the spinal micturition reflex after spinal cord injury. Transplantation resulted in new axons growing at the transplant site, implying the possible existence of interneuron bridging across the injured spinal cord.

Olfactory ensheathing cells (OECs) transplantation in spinal cord prolongs the survival of mutant SOD1 (G93A) ALS rats through neuroprotection and remyelination

Amyotrophic lateral sclerosis (ALS) is a fatal progressive neurodegenerative disease. SOD1G93A rat(n=20), medium+SOD1G93A rat(n=20), OECs+SOD1G93A rat(n=20), and another 20 wild type rats as control. 5μL(1×105) OECs was injected into the dorsal funiculus of the thoracic spinal cord (T7) through a glass micropipette at depths of 0.7 mm and 0.4 mm (0.5 lL). Kaplan Meier survival analysis revealed a significant increase (7 days) in the survival time of the OEC+SOD1G93A rats. Body weight record and Inclined board test showed a significant difference from onset 7days to 11days (P<0.05). Motor neuron count per ventral horn of thoracic spinal cord showed a significant motor neuron loss in SOD1G93A rats(P<0.001). Immunohistochemistry and Western blot ChAT analysis supported the motor neuron count. Confocal microscope images of tissue sections indicated that the transplanted OEC could have survived more than four weeks and migrated as long as 8.4mm through the spinal cord. Evidence of remyelination was captured. In conclusion, the transplanted OECs could serve as a source of neuroprotection and a remyelination to positively modify the ALS‐specific microenvironment.

Survival of neurally induced mesenchymal cells may determine degree of motor recovery in injured spinal cord rats

We recently developed a new method for efficient generation of neural-like cells from mice bone marrow (BM)-derived mesenchymal stem cells (MSC) by exposing MSCs to epigenetic modifiers and a neural stem cell environment. These neurally induced MSCs (NI-MSCs) differentiate into neuronal- and glial-like cells in vitro, release neurotrophic factors NGF and BDNF, survive and integrate after transplantation in intact spinal cord. The aim of this study was to determine whether transplanted NI-MSCs survive, differentiate, and integrate in injured spinal cord (ISC) rats and promote functional recovery. Twenty rats, half grafted with MSCs and half with NI-MSCs, were used for survival and differentiation studies. Results were analyzed using triple-labeled immunohistochemistry. For motor function studies the 3 group of adult female Sprague Dawley rats received PBS (vehicle), MSCs, or NI-MSCs, respectively. Functional outcome was measured using the BBB scale. Results demonstrated gradual improvement of locomotor function in NI-MSC-transplanted rats in comparison to vehicle and non-modified MSC-transplanted animals, with statistically significant differences at 7, 14, and 21 days post transplantation. Immunocytochemical studies revealed poor survival of NI-MSCs within the ISC as early as 3 weeks after transplantation. Thus, there is a correlation between the degree of surviving NI-MSCs and extent of functional recovery.

Using the ‘six thinking hats’ model of learning in a surgical nursing class: sharing the experience and student opinions

Objective: The purpose of the study was to test the use of a creative teaching method in developing nursing students’ critical thinking skills The ‘six thinking hats’ model, which was developed by Edward DeBono in 1994, is a teaching method for developing creative thinking by presenting and systematising thoughts and suggestions within a specific structure. The six different hats (white, yellow, black, red, green, and blue) symbolise the thinking system. In this method the students are expected to change their perspectives symbolised by a colour at regular intervals. Setting: This study was conducted during the 2006‑2007 academic year at Gaziosmanpaşa University. Subjects: The research population was comprised of 41 students who were registered for the surgical nursing class in the Department of Nursing at Tokat School for Health Sciences. Purposive sampling was not undertaken for the study with the entire population of students targeted. Method: The ‘six thinking hats’ model was used in theoretical lessons about breast cancer, spinal cord trauma and transplantation nursing in a surgical nursing class. At the end of the theoretic section of the class, the students’ opinions were sought about the ‘sixthinking hats’ model of learning. Data were analysed with percentage and Chi square test reporting of the responses. Findings: The majority of the students stated that this method facilitated their empathising with the patient; sharing different ideas and opinions; considering the patient holistically; generating creative ideas; looking at an event from positive and negativeaspects; and developing their system of thinking. They recommended that this class be taught using this method. Conclusion: This study is important because it tested the use of a creative teaching method that is useful for the development of nursing students’ critical thinking skills. The study is also important because there are a limited number of references in the literature about the use of the ‘six thinking hats’ model of learning in university education, and only one in nursing education, and having this type of study in the nursing literature adds to the improvement and enrichment of nursing education.

Human neural precursor cells continue to proliferate and exhibit low cell death after transplantation to the injured rat spinal cord

During the last decade, the interest in stem and progenitor cells, and their applications in spinal cord injuries have steadily increased. However, little is known about proliferation and cell death mechanisms in these cells after transplantation to the spinal cord. The aim of the present project was to study cell turn-over, i.e. total cell number, with time course of proliferation and cell death, in human neural precursor cells (NPCs) after transplantation to the injured rat spinal cord. Immunodeficient rats were subjected to lateral clip compression injuries, transplanted with neurospheres of human forebrain-derived NPCs two weeks after lesion, and sacrificed after 6 h, 1, 3, 10, or 21 days. Cell death was assessed by quantifying human cells immunoreactive for active caspase-3 and calpain 1-dependent fodrin breakdown products (FBDP). The results showed that after an initial drop, the number of implanted cells increased over time after transplantation. Cell proliferation was substantial, with 34% of human cells being immunoreactive for proliferating cell nuclear antigen at 6 h, but which declined over the next few days. The fractions of caspase-3-, and FBDP-immunoreactive cells were remarkably low, together representing 18% of all human cells at 6 h, and rapidly decreasing the next few days. Our results show that already 10 days after spinal cord transplantation of human NPCs as intact neurospheres, the number of human cells exceeded the initially implanted, which was the result of marked cell proliferation in combination with a low rate of apoptotic and non-apoptotic cell death taking place early after transplantation.

Effects of long-term FK506 administration on functional and histopathological outcome after spinal cord injury in adult rat.

FK506 (tacrolimus), a potent immunosuppressive drug primarily used for reduction of allograft rejection in organ transplantation, also offers neuroprotection after central nervous system injury. FK506-mediated immunosuppression and neuroprotection may occur through different mechanisms that could affect neurological recovery and the severity of spinal lesions where cells transplantation therapy is combined with FK506 application. We assessed effects of long-term FK506 administration using the same dose regiment (1 mg/kg/day for 6 weeks) as is used in spinal cord transplantation studies following a balloon-compression induced spinal cord injury (SCI). Body weight and locomotor recovery quantified by the BBB (Basso-Beattie-Bresnehan) locomotor rating scale were evaluated for up to 42 days post-injury. The area of the preserved spinal cord tissue within a 13 mm segment of the spinal cord (lesion epicenter and 6 mm rostral-caudal) was examined histologically. The results showed no significant effects of FK506 on spinal cord tissue sparing or improvement of locomotor recovery. However, body weight fell significantly (P < 0.05) with FK506 treatment when compared with placebo from day 7 until sacrifice. In our experimental design, long-term FK506 treatment did not affect the parameters of outcome following balloon-compression SCI in the rat; however, multiple effects of FK506 should be taken into account when evaluating the outcomes in transplantation studies.

Involvement of Acidic Fibroblast Growth Factor in Spinal Cord Injury Repair Processes Revealed by a Proteomics Approach

Acidic fibroblast growth factor (aFGF; also known as FGF-1) is a potent neurotrophic factor that affects neuronal survival in the injured spinal cord. However, the pathological changes that occur with spinal cord injury (SCI) and the attribution to aFGF of a neuroprotective effect during SCI are still elusive. In this study, we demonstrated that rat SCI, when treated with aFGF, showed significant functional recovery as indicated by the Basso, Beattie, and Bresnahan locomotor rating scale and the combined behavior score (p < 0.01-0.001). Furthermore proteomics and bioinformatics approaches were adapted to investigate changes in the global protein profile of the damaged spinal cord tissue when experimental rats were treated either with or without aFGF at 24 h after injury. We found that 51 protein spots, resolvable by two-dimensional PAGE, had significant differential expression. Using hierarchical clustering analysis, these proteins were categorized into five major expression patterns. Noticeably proteins involved in the process of secondary injury, such as astrocyte activation (glial fibrillary acidic protein), inflammation (S100B), and scar formation (keratan sulfate proteoglycan lumican), which lead to the blocking of injured spinal cord regeneration, were down-regulated in the contusive spinal cord after treatment with aFGF. We propose that aFGF might initiate a series of biological processes to prevent or attenuate secondary injury and that this, in turn, leads to an improvement in functional recovery. Moreover the quantitative expression level of these proteins was verified by quantitative real time PCR. Furthermore we identified various potential neuroprotective protein factors that are induced by aFGF and may be involved in the spinal cord repair processes of SCI rats. Thus, our results could have a remarkable impact on clinical developments in the area of spinal cord injury therapy.

Investigation of the potential of mesenchymal stem cells (MSCs) for gene delivery to breast tumours

Amyotrophic lateral sclerosis (ALS) is a lethal disease affecting motoneurons. In familial ALS, patients bear mutations in the superoxide dismutase gene (SOD1). We transplanted human bone marrow mesenchymal stem cells (hMSCs) into the lumbar spinal cord of asymptomatic SOD1G93A mice, an experimental model of ALS. hMSCs were found in the spinal cord 10 weeks after, sometimes close to motoneurons and were rarely GFAP- or MAP2-positive. In females, where progression is slower than in males, astrogliosis and microglial activation were reduced and motoneuron counts with the optical fractionator were higher following transplantation. Motor tests (Rotarod, Paw Grip Endurance, neurological examination) were significantly improved in transplanted males. Therefore hMSCs are a good candidate for ALS cell therapy: they can survive and migrate after transplantation in the lumbar spinal cord, where they prevent astrogliosis and microglial activation and delay ALS-related decrease in the number of motoneurons, thus resulting in amelioration of the motor performance.

Human mesenchymal stem cell transplantation extends survival, improves motor performance and decreases neuroinflammation in mouse model of amyotrophic lateral sclerosis

Amyotrophic lateral sclerosis (ALS) is a lethal disease affecting motoneurons. In familial ALS, patients bear mutations in the superoxide dismutase gene (SOD1). We transplanted human bone marrow mesenchymal stem cells (hMSCs) into the lumbar spinal cord of asymptomatic SOD1 G93A mice, an experimental model of ALS. hMSCs were found in the spinal cord 10 weeks after, sometimes close to motoneurons and were rarely GFAP- or MAP2-positive. In females, where progression is slower than in males, astrogliosis and microglial activation were reduced and motoneuron counts with the optical fractionator were higher following transplantation. Motor tests (Rotarod, Paw Grip Endurance, neurological examination) were significantly improved in transplanted males. Therefore hMSCs are a good candidate for ALS cell therapy: they can survive and migrate after transplantation in the lumbar spinal cord, where they prevent astrogliosis and microglial activation and delay ALS-related decrease in the number of motoneurons, thus resulting in amelioration of the motor performance.

Adult olfactory bulbs from primates provide reliable ensheathing glia for cell therapy

Olfactory bulb ensheathing glia (OB-OEG) from adult rodents promote functional and morphological repair after grafting into injured spinal cords. To provide insight into the feasibility of using OB-OEG in human therapy, we studied OB-OEG in primates to determine their suitability for spinal cord transplantation. Here, we show that OEG can be obtained from olfactory bulbs of adult macaca mulatta and nemestrina monkeys and compare their characteristics to those obtained from rats. In contrast to rodent OB-OEG, primate OB-OEG are nonsenescent, exhibit a longer lifespan, are less sensitive to high oxygen culture environment, and maintain a phenotype suitable for grafting for up to 2.5 months in vitro. Three-week cultures (short term) derived from a single macaca olfactory bulb provide enough OEG for autologous transplantation at the acute stage of injury, and after long-term cultures (2.5 months) may yield an additional 20 billion. OEG can be frozen for later use. Therefore, primate adult olfactory bulbs constitute a reliable source of OEG for cell therapy, and successful culture of these cells make autologous transplantation feasible.

Reproduction Medicine —The Present State and Problems in Spinal Cord Transplantation—

Reproduction Medicine —The Present State and Problems in Spinal Cord Transplantation—

Plasmid-based genetic modification of human bone marrow-derived stromal cells: analysis of cell survival and transgene expression after transplantation in rat spinal cord

BackgroundBone marrow-derived stromal cells (MSC) are attractive targets for ex vivo cell and gene therapy. In this context, we investigated the feasibility of a plasmid-based strategy for genetic modification of human (h)MSC with enhanced green fluorescent protein (EGFP) and neurotrophin (NT)3. Three genetically modified hMSC lines (EGFP, NT3, NT3-EGFP) were established and used to study cell survival and transgene expression following transplantation in rat spinal cord.ResultsFirst, we demonstrate long-term survival of transplanted hMSC-EGFP cells in rat spinal cord under, but not without, appropriate immune suppression. Next, we examined the stability of EGFP or NT3 transgene expression following transplantation of hMSC-EGFP, hMSC-NT3 and hMSC-NT3-EGFP in rat spinal cord. While in vivo EGFP mRNA and protein expression by transplanted hMSC-EGFP cells was readily detectable at different time points post-transplantation, in vivo NT3 mRNA expression by hMSC-NT3 cells and in vivo EGFP protein expression by hMSC-NT3-EGFP cells was, respectively, undetectable or declined rapidly between day 1 and 7 post-transplantation. Further investigation revealed that the observed in vivo decline of EGFP protein expression by hMSC-NT3-EGFP cells: (i) was associated with a decrease in transgenic NT3-EGFP mRNA expression as suggested following laser capture micro-dissection analysis of hMSC-NT3-EGFP cell transplants at day 1 and day 7 post-transplantation, (ii) did not occur when hMSC-NT3-EGFP cells were transplanted subcutaneously, and (iii) was reversed upon re-establishment of hMSC-NT3-EGFP cell cultures at 2 weeks post-transplantation. Finally, because we observed a slowly progressing tumour growth following transplantation of all our hMSC cell transplants, we here demonstrate that omitting immune suppressive therapy is sufficient to prevent further tumour growth and to eradicate malignant xenogeneic cell transplants.ConclusionIn this study, we demonstrate that genetically modified hMSC lines can survive in healthy rat spinal cord over at least 3 weeks by using adequate immune suppression and can serve as vehicles for transgene expression. However, before genetically modified hMSC can potentially be used in a clinical setting to treat spinal cord injuries, more research on standardisation of hMSC culture and genetic modification needs to be done in order to prevent tumour formation and transgene silencing in vivo.

Adult Olfactory Bulb Neural Precursor Cell Grafts Provide Temporary Protection From Motor Neuron Degeneration, Improve Motor Function, and Extend Survival in Amyotrophic Lateral Sclerosis Mice

Amyotrophic lateral sclerosis is a fatal disease caused by degeneration of motor neurons (MNs). We transplanted multipotent neural precursor cell (NPC)-neurospheres from mouse olfactory bulb (OB) into the spinal cord of transgenic mice that develop MN degeneration because of human mutant superoxide dismutase-1 (mSOD1). Adult NPCs were isolated from the OB core of transgenic mice expressing green fluorescent protein, human wild-type SOD1, or human mSOD1. mSOD1 mice received lumbar spinal cord transplants of OB-NPC neurospheres at preclinical stages of disease (70 days old). Control mSOD1 mice received dead cells or recombinant green fluorescent protein. OB-NPCs attenuated the loss of motor function and wasting. They delayed disease onset to approximately 117 days, compared with control onset at approximately 90 days. The lifespan of NPC recipient mice was extended (approximately 170 days) compared with the lifespan of controls (approximately 140 days). Transplanted OB-NPCs differentiated into large spinal neurons positive for choline acetyltransferase, interneurons, and glial cells. Loss of endogenous MNs was attenuated in mSOD1 mice with transplants. New neurons formed myelinated axons and synapses. NPC-derived neurons issued axons that grew into peripheral nerve. OB-NPCs also differentiated into oligodendrocytes and astrocytes that contacted neuronal processes. We conclude that transplantation of adult OB-NPCs is therapeutic for mouse amyotrophic lateral sclerosis.

Transplantation of Adult Rat Spinal Cord Stem/Progenitor Cells for Spinal Cord Injury

Stem/progenitor cells derived from the ependymal region of the spinal cord have the ability to self-renew and are multipotential for neurons and glia. These cells may have the ability to regenerate the injured mammalian spinal cord as they do in some lower vertebrates. However, the optimal conditions for transplantation and the fate of transplanted cells are not fully known. In the current study, spinal cord stem/progenitor cells were cultured from adult male rats expressing enhanced green fluorescent protein (eGFP). Neurospheres were transplanted at the time of clip compression injury (35-g force) into the injury site, or 1 mm rostral and caudal to the injury site. Neurospheres were also transplanted into a subacute model (day 9 after injury) and a chronic model (day 28 after injury). Functional recovery was also studied in an acute injury model with weekly locomotor testing over a 16-week period. A significant increase in cell survival at 7 days was seen in rats receiving rostral and caudal injections as compared to injection directly into the site of injury. A significant increase in cell survival was also seen in rats receiving subacute transplants at 9 days after injury. Transplanted cells differentiated primarily into astrocytes (31.2%) and oligodendrocytes (50.3%), and a small number of neurons (1%). No improvement was seen in the Basso, Beattie and Bresnahan (BBB) locomotor rating scale after acute transplantation as compared with injury only, although surviving transplanted cells were identified that had migrated across the injury site from the rostral and caudal injection sites.

Comparison of cell populations derived from canine olfactory bulb and olfactory mucosal cultures

To evaluate the numbers and proportions of olfactory ensheathing cells (OECs) in cell cultures derived from the olfactory bulb (OB) and olfactory mucosa of dogs. 7 dogs. OB tissue and olfactory mucosa from the nasal cavity and frontal sinus were obtained from euthanatized dogs and prepared for cell culture. At 7, 14, and 21 days of culture in vitro, numbers and proportions of OECs, astrocytes, and fibroblasts were determined via immunocytochemistry. Antibody against the low-affinity nerve growth factor receptor p75 was used to identify OECs, antibody against glial fibrillary acidic protein was used to identify astrocytes, and antibody against fibronectin was used to identify fibroblasts. Cultured OECs derived from the olfactory mucosa of the nasal cavity and frontal sinus had similar characteristics. However, whereas OECs in the OB cell cultures constituted approximately 50% of the cells at 7 days and approximately 75% at 21 days the proportion of OECs in cultures derived from both mucosal types was much lower, with approximately 40% OECs at 7 days and approximately 25% at 21 days. Analysis of OEC numbers revealed that these changes were accompanied by corresponding decreases and increases in the population of cells with fibronectin receptors. Although olfactory mucosal cell cultures yielded a sufficient number of OECs for spinal cord transplantation procedures in dogs, modification of culture conditions would be required to ensure that the derived cell population contained a sufficient proportion of OECs.

Delayed Intervention with Transplants and Neurotrophic Factors Supports Recovery of Forelimb Function after Cervical Spinal Cord Injury in Adult Rats

The adult central nervous system is capable of considerable anatomical reorganization and functional recovery after injury. Functional outcomes, however, vary greatly, depending upon size and location of injury, type and timing of intervention, and type of recovery and plasticity evaluated. The present study was undertaken to assess the recovery of skilled and unskilled forelimb function in adult rats after a C5/C6 spinal cord over-hemisection and delayed intervention with fetal spinal cord transplants and neurotrophins. Recovery of forelimb function was evaluated during both target reaching (a skilled behavior) and vertical exploration (an unskilled behavior). Anatomical tracing and immunohistochemistry were used to assess the growth of descending raphespinal, corticospinal, and rubrospinal fibers at the injury site, tracts that normally confer forelimb function. Delayed intervention with transplants and either brain-derived neurotrophic factor (BDNF) or neurotrophin-3 (NT-3) restored skilled left forelimb reaching to pre-injury levels. Animals showed recovery of normal reaching movements rather than compensation with abnormal movements. Transplants and NT-3 also improved right forelimb use during an unskilled vertical exploration, but not skilled right reaching. Intervention with fetal transplant tissue supported the growth of descending serotonergic, corticospinal, and rubrospinal fibers into the transplant at the lesion site. The addition of neurotrophins, however, did not significantly increase axonal growth at the lesion site. These studies suggest that the recovery of skilled and unskilled forelimb use is possible after a large cervical spinal cord injury following delayed intervention with fetal spinal cord and neurotrophins. Plasticity of both spared and axotomized descending pathways likely contributes to the functional recovery observed.

Apparent Diffusion Coefficients in Spinal Cord Transplants and Surrounding White Matter Correlate With Degree of Axonal Dieback After Injury in Rats

Apparent Diffusion Coefficients in Spinal Cord Transplants and Surrounding White Matter Correlate With Degree of Axonal Dieback After Injury in Rats

Degradation of chondroitin sulfate proteoglycans potentiates transplant-mediated axonal remodeling and functional recovery after spinal cord injury in adult rats

Transplantation of growth-permissive cells or tissues was used to bridge a lesion cavity and induce axonal growth in experimental spinal cord injury (SCI). Axonal interactions between host and transplant may be affected by upregulation of inhibitory chondroitin sulfate proteoglycans (CSPGs) following various transplantation strategies. The extent of axonal growth and functional recovery after transplantation of embryonic spinal cord tissue decreases in adult compared to neonatal host. We hypothesized that CSPGs contribute to the decrease in the extent to which transplant supports axonal remodeling and functional recovery. Expression of CSPGs increased after overhemisection SCI in adult rats but not in neonates. Embryonic spinal cord transplant was surrounded by CSPGs deposited in host cord, and the interface between host and transplant seemed to contain a large amount of CSPGs. Intrathecally delivered chondroitinase ABC (C'ase) improved recovery of distal forelimb usage and skilled motor behavior after C4 overhemisection injury and transplantation in adults. This behavioral recovery was accompanied by an increased amount of raphespinal axons growing into the transplant, and raphespinal innervation to the cervical motor region was promoted by C'ase plus transplant. Moreover, C'ase increased the number of transplanted neurons that grew axons to the host cervical enlargement, suggesting that degradation of CSPGs supports remodeling not only of host axons but also axons from transplanted neurons. Our results suggest that CSPGs constitute an inhibitory barrier to prevent axonal interactions between host and transplant in adults, and degradation of the inhibitory barrier can potentiate transplant-mediated axonal remodeling and functional recovery after SCI.