Abstract
Cell transplantation therapies have become a major focus in pre-clinical research as a promising strategy for the treatment of spinal cord injury (SCI). In this article, we systematically review the available pre-clinical literature on the most commonly used cell types in order to assess the body of evidence that may support their translation to human SCI patients. These cell types include Schwann cells, olfactory ensheathing glial cells, embryonic and adult neural stem/progenitor cells, fate-restricted neural/glial precursor cells, and bone-marrow stromal cells. Studies were included for review only if they described the transplantation of the cell substrate into an in-vivo model of traumatic SCI, induced either bluntly or sharply. Using these inclusion criteria, 162 studies were identified and reviewed in detail, emphasizing their behavioral effects (although not limiting the scope of the discussion to behavioral effects alone). Significant differences between cells of the same “type” exist based on the species and age of donor, as well as culture conditions and mode of delivery. Many of these studies used cell transplantations in combination with other strategies. The systematic review makes it very apparent that cells derived from rodent sources have been the most extensively studied, while only 19 studies reported the transplantation of human cells, nine of which utilized bone-marrow stromal cells. Similarly, the vast majority of studies have been conducted in rodent models of injury, and few studies have investigated cell transplantation in larger mammals or primates. With respect to the timing of intervention, nearly all of the studies reviewed were conducted with transplantations occurring subacutely and acutely, while chronic treatments were rare and often failed to yield functional benefits.
Introduction
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The purpose of this review is to describe, for the most-studied and best-understood cell types used in SCI research, the current body of pre-clinical literature that might support the translation of such treatments into human clinical trials. Given the explosion of interest in therapeutic approaches to SCI and the large number of cell candidates that are reportedly beneficial in animal models of SCI, it is difficult – if not impossible – to cover the field comprehensively. We felt a need to limit this review to the “best studied cell types,” since unlike a specific pharmacologic agent, a given “cell type” can vary considerably from laboratory to laboratory due to differences in the source materials (age, gender, and species from which the cells are taken, or progenitors from which the cells are generated), cell purity and contamination with other cell types, culture conditions (such as number of passages), and variability of media used – to list only a few confounding factors. Hence, a “cell type” has become an umbrella term for several subtypes of cells, and a larger body of data from several laboratories is needed to build consensus regarding the identity, benefits, risks, and translational potential of a given cell as a therapeutic candidate for SCI.
Most cell transplantations are delivered directly into the site of injury or adjacent to it by injecting a few microliters of cell suspension (with several hundred thousand cells) via fine needles or glass capillaries. Attempts have been made to deliver cell substrates to the injured cord via intrathecal injection or even systemically via intravenous infusions. With a few exceptions, rodent models of SCI are used, and the transplantation is typically performed 1–2 weeks after the injury (herein referred to as “subacute” treatment, since transplantations performed immediately after injury – “acutely” – generally yield poor results due to the robust inflammatory response initiated at the time of injury). Only a few chronic studies have been reported, which is a disquieting issue for patients with chronic SCI who are often the most ardent consumers of information regarding cell transplant technologies. While the demonstration of neurologic benefit in rodent models is viewed as evidence that the cell therapy in question may have a “therapeutic” potential in human SCI, it is important to point out that the majority of such studies employ rodent or mouse cells that would be impossible to implant into humans. The number of studies in which human cells are actually tested in such rodent models is remarkably low.
Our strategy, therefore, was to select cell therapies that have reasonable “translational potential” by virtue of the fact that they had been under extensive pre-clinical investigation. While it was not the intention to attempt to cover the entire SCI transplantation field, our goal was to apply the tenets of systematic review to the specific cell therapies that met these conditions. By performing a systematic review of these cellular therapies, we hoped to provide the field with an overview of the body of pre-clinical evidence that supports (or fails to support) the translation of the therapy into human trials.
Methodology
In the summer of 2008, a PubMed search was conducted on “spinal cord injury” and the cell type of interest (e.g., “Schwann cell” or “neural stem cell”). We performed the PubMed search with the “cell type,” recognizing that this would be an umbrella search term for possible subtypes (e.g., “neural stem cells,” which might come from adult or embryonic sources).
From the list of studies generated through this fairly indiscriminant search, we applied the following criteria to systematically review the pre-clinical literature on these therapies. The inclusion criteria for these studies were: • Studies that evaluated the cell therapy in an in-vivo model of • Studies that included a control group for the cell transplantation experiment. • The presence of at least
Exclusion criteria for this systematic review were: • Studies using non-traumatic local or global ischemia models, photothrombotic models, demyelination models. • Studies of injury to the cauda equina or conus. • Studies of root avulsions or injuries to the dorsal root entry zone. • Studies that evaluated the cell therapy exclusively in vitro. • Studies with n = 3 or fewer animals in a rodent experiment. • Studies with reportedly greater than >30% loss of animals. • Studies with fewer than 7 days survival time. • Studies in which there were single reports from one laboratory only on a given “umbrella cell type.” • Studies describing the experimental application of a cell substrate into human SCI patients. As this was a review of pre-clinical literature, such human studies, while important, could not be included.
The data from those studies that fitted the criteria were then extracted into a table format to depict the animal model, injury model, the treatment's dose and timing, the experimental groups tested in the study and the “n per group”, and the reported behavioral and non-behavioral outcomes (e.g., histological, biochemical, or physiologic outcomes). A summary statement about the body of literature was then generated.
Results
Using this selection process, we identified the following cell “types” and grouped the studies according to the following “umbrella” cell type: Schwann cells, olfactory ensheathing glial cells, neural stem/progenitor cells (adult and embryonic), mesenchymal stem cells (most from bone marrow). The heterogeneity of each cell type is reflected in the tables, as the studies within each cell type were further organized according to the
The PubMed searches on these therapies were conducted in the spring/summer of 2008 by SCI researchers across Canada (plus one from the United States). By applying the previously described criteria (essentially, in-vivo animal studies utilizing a traumatic model of SCI), the following publications were selected, and the tables for each of these respective cell therapies are listed below.
Schwann cells and their combinations (Table 2)
Schwann cells (SCs) are the myelin-forming cells of the peripheral nervous system, and have been shown not only to myelinate (remyelinate) axons after transplantation into the injured spinal cord but also to form a permissive substrate for regenerating axons, as reported in many of the studies reviewed here.
d: day, days; hr: hour, hours; i.v.: intravenous; PI: post-injury; PT: post-transplant; SCI: spinal cord injury; SD – Sprague Dawley; Tx: transection; wk: week, weeks
5HT: serotonin; AdV: adenoviral; aFGF: acidic fibroblast growth factor; BBB: Basso, Beattie and Bresnahan locomotor test; BDNF – brain-derived neurotrophic factor; bFGF (FGF2): basic fibroblast growth factor; CCK: cholecystokinin; CGRP - calcitonin gene-related peptide; ChABC: chondroitinase ABC; CsA: Cyclosporine; CSF: cerebrospinal fluids; CSPG - chondroitin sulfate proteoglycan; CST - cortico-spinal tract; CTB: cholera toxin beta; DBH: dopamine-β-hydroxylase; DMEM: Dulbecco's modified Eagle's medium; DRG - dorsal root ganglion; EM: electron microscopy; ENK: enkephalin; FB: fibroblast; GDNF: glial cell line-derived neurotrophic factor; GFAP: glial fibrillary acidic proteins; GFP – green fluorescent protein; FBs: fibroblasts; IGF-1: insulinlike growth factor; LV: lentivirus; LacZ – beta-galactosidase; MG: matrigel; MRI-magnetic resonsnce imaging; MP: methylprednisolone; NF: neurofilament; NGF: nerve growth factor; NPC: neural progenitor/precursor cells; NT: neurotrophin; OEC: olfactory ensheahting cell; OEG: olfactory ensheathing glia; PAN/PVC: poly(acrilonitrile-vinyl chloride); PDGF: platelet-derived growth factor; PSA: polysialic acid; SCs:- Schwann cells; SKPs: skin precursors; SP: substance P; STX: sialyl-transferase; TH: tyrosine hydroxylase; TPH: tryptophan hydroxylase; VIP: vasoactive intestinal peptide.
Of all the cell types examined in the context of this review, SCs have the longest history of transplantation, with the first experiment involving the transplantation of purified SCs occurring in 1981 (Duncan et al., 1981). Much of the early work understanding the basic biology of SC transplantation involved transplanting SCs into the brain and spinal cord in models of demyelination, and is not discussed here (for a review, see Duncan and Milward, 1995). These early transplant studies demonstrated the ability of SCs to myelinate demyelinated CNS axons, as well as the regenerative ability of PNS axons, which made SCs a cell type of interest for SCI injury repair. More recently, it has been recognized that cell transplants (SCs but also OEG and BMSCs) facilitate the invasion of host SCs into the injured spinal cord (Biernaskie et al., 2007; Hill et al., 2006). This invasion of endogenous cells results in a transplant that is a mixture of transplanted cells and host SCs, and suggests that host SCs may contribute to the recovery observed in such transplants.
Most of the studies reviewed were performed with adult rodent (mostly rat) nerve-derived SCs (n = 35). Thirty-two of these studies inflicted injuries to the spinal cord at mid to low thoracic levels, and employed blunt contusion/compression type injuries (n = 11) or full (n = 14) and partial (n = 8) transection injuries. Experiments employing full or partial transection injuries have been used in combination with matrix filled channels to examine the ability of SCs to promote CNS axonal regeneration. A number of these studies clearly demonstrated that SCs are very good at enhancing the regeneration of sensory axons from the dorsal root ganglia, as well as propriospinal axons adjacent to the injury site. These studies also highlight the limits of SC transplants, in that SCs alone (at least in complete injury models) are not sufficient to promote regeneration of brainstem spinal axons, nor do they permit axons that enter SCs grafts to exit and reenter the host spinal cord. As a result, there has been demonstrable interest in enhancing the therapeutic utility of SCs by using them in combination with other co-treatments such as neuroprotective agents, with other cell substrates, or after transduction with growth factor expression vectors. All of these studies were carried out in rats.
Clinical translation emphasizes the need for the pre-clinical demonstration of behavioral benefits, and we emphasize this aspect in greater details in this summary below. Of these 35 publications with adult nerve derived SCs, 10 performed behavioral analyses using the Basso, Beattie, Bresnahan (BBB) open field locomotion scale: six after blunt lesions and four after full transection. Of the five studies involving thoracic contusion injury in which a comparison with injury alone can be made, two studies reported significant behavioral benefits in open field locomotion after SC transplantation alone. These included a report by Takami and colleagues (2002) in a subacute setting and a study by Barakat and colleagues (2005) in an 8-week chronic contusion injury (the only chronic study in the SC literature). The three other studies only saw benefits when SCs were used in combinations with either rolipram plus cAMP (Pearse et al., 2004a), olfactory ensheathing cells (OECs) (Pearse et al., 2007), or the combination of methylprednisolone, IL-10, and OECs (Pearse et al., 2004b). Despite the lack of effect of SCs alone in these studies, SCs score almost one point above injury controls in two of these studies with BBB scores ranging from 10 to 11.5 versus 9.5 to 10.7 for SCI controls. It is noteworthy that the two studies in which SCs were efficacious involve pre- and post-transplantation behavioral assessment, whereas the three studies in which SCs alone had no effect did not score behavior prior to one week post-transplantation. To date, the BBB score comparing SCs with matrix (Fibrin or Matrigel) only has not been reported, making it difficult to ascertain the functional benefits of SCs in either the complete or incomplete transection model. Only one of the four studies that performed behavioral analysis after full transections yielded clear behavioral benefits. In this study, the SCs were used in combination with a Matrigel-filled PAN/PVC channel plus OECs injections into the stumps plus ChABC and mouse IgG (Fouad et al., 2005), and it did not include a SC-only control. Given that the BBB score reported for the combination treatment (6.6) was similar to that of human SCs suspended in the same matrix (8.2) (Guest et al., 1997b) and rat SCs suspended in fibrin (6.3) (Hurtado et al., 2006), it remains unclear if the combination treatment is truly better than SCs alone. These behavioral findings suggest that additional, well-designed, and properly controlled studies are needed to assess the functional benefits of SCs. While the focus of interest is in the development of SCs as a transplantation therapy, as multiple centers move toward or have begun human clinical trails (Saberi et al., 2008), there is strong pre-clinical interest in the use of additional experimental co-treatments. However, translating such a combination of two experimental treatments into clinical trials has significant implications from a regulatory standpoint, particularly if each treatment in the “co-treatment” is experimental.
Despite the clinical observation that about 60% of human SCIs occur at the cervical level, experiments involving cervical injuries are rare. Only two studies that evaluated adult nerve-derived rodent SCs were performed at the cervical level of the spinal cord. One study that used a clinically relevant contusion injury without co-treatments reported improvements in forelimb grip strength and forelimb hang tests (Schaal et al., 2007).
SCs from nerves of newborn rodents were used in four studies, three of these employing a thoracic contusion injury and the other a lumbar partial transection. While all three contusion studies employed the BBB, only two showed behavioral benefits. It should be pointed out that in one of these two positive studies, efficacy was not observed with wildtype SCs, but rather in SCs transduced to express the cell adhesion molecule PSA. In the other positive study, an astonishing BBB improvement from 9 to 13.5 was reported using the mere injection of 50,000 neonatal SCs into the subarachnoid space after clip compression injury (Firouzi et al., 2006). It should be noted that the rats in this study were very young (100–140 grams), which corresponds with 45–60 days of age. Recently, it has been suggested that SC precursors may be more beneficial than SCs from newborn rodents. However, even with enhanced CST regeneration (something that SCs from adults typically do not promote), these SCs from newborns did not result in functional improvements in a cervical crush model (Agudo et al., 2008). Further studies are warranted to compare the effects of SCs derived from young versus adult animals. The ethical and logistical considerations of acquiring human SCs from embryonic or early postnatal sources makes them less attractive as a therapeutic approach than human SCs derived from the nerves of adults, particularly if an autologous transplantation can be performed.
Considering the fact that a clinical trial of SCs requires the transplantation of human cells, it is remarkable that human SCs have been reported in only two pre-clinical rodent studies. Both of these studies by Guest and colleagues were in thoracic full transection SCI models (Guest et al., 1997a, 1997b). In one study, in which behavior was assessed, the authors report a small but significant behavioral benefit in the BBB and the inclined plane test. The SCs were used in conjunction with a guidance channel and Matrigel, for which there is presently no FDA-approved formulation available. Pre-clinical experiments examining the survival and efficacy of human SC in contusion models of SCI are clearly needed. Despite the paucity of human SC experience in traumatic SCI, it is noted that a number of studies have looked at the effects of cultured human SCs in rodent models of demyelination (Kohama et al., 2001) and peripheral nerve injury (Hood et al., 2009). Additionally, autologous human SCs have been implanted into humans with multiple sclerosis (Brierly et al., 2001; Halfpenny et al., 2002).
An autologous transplantation approach is appealing as it eliminates concerns regarding immune rejection and avoids the controversy over embryonic or neonatal sources. However, an autologous approach necessitates sacrificing a peripheral nerve (the significance of which may be negligible in a completely paralyzed individual), and despite improvements in amplification techniques, a number of weeks are still needed before enough cells can be generated for transplantation. To obviate the need for harvesting a peripheral nerve from a patient, alternative sources of SCs from postnatal skin or adult bone marrow have recently been pursued and tested after thoracic transection (Kamada et al., 2005) or contusion (Biernaskie et al., 2007) injuries. Both studies reported modest (but significant) improvements on the BBB scale and subscale, suggesting that other sources, which may potentially be less invasive that peripheral nerve biopsies, may be an alternative source for autologous SCs. Intriguingly, the SCs from skin-derived progenitors formed bridges across the injury site, migrated into the host parenchyma, and formed myelin with minimal astrocyte hypertrophy (Biernaskie et al., 2007).
Despite the lack of pre-clinical data using human SCs, clinical trials involving the use of human SCs are moving ahead. Saberi and colleagues (2008) in Iran recently published the first results from 4 of 33 patients with chronic thoracic SCI (2.0–6.5 years post injury) that underwent autologous SC transplantation. No detrimental (or beneficial) effect was reported in the first four patients. While it has been possible to identify SC cables using MRI in rats (Iannotti et al., 2002) and fetal tissue transplants in the human spinal cord (Wirth et al., 2001), MRI failed to identify the SC transplants in the study by Saberi and colleagues. This trial, following ICCP guidelines, is a promising first step in the move toward human translation of SCs in SCI. A summary of the pros and cons and knowledge gaps for SC transplantation is depicted in Table 3.
Olfactory ensheathing cells (also known as olfactory ensheathing glia; Table 4)
OECs are found in the nerve fiber layer of the olfactory bulb, as well as in the nasal olfactory mucosa. These cells have garnered considerable interest because of their ability to facilitate the lifelong repeated regeneration of olfactory axons from the PNS environment of the nasal olfactory mucosa to the CNS environment of the olfactory bulb (Doucette, 1991). Significant differences have been revealed between OECs of various origins (Richter et al., 2005), and therefore these are considered separately in the tables. In addition, the properties of OECs can change considerably depending on the culture conditions (e.g., the number of passages in vitro) (Au et al., 2007). Hence, optimal cell source and treatment for transplantation into the injured spinal cord is subject to an ongoing debate.
d: day, days; hr: hour, hours; PI: post-injury; PT: post-transplant; SCI: spinal cord injury; s.c.: subcutaneous; SD: Sprague–Dawley; Tx: transection; wk: week, weeks
5HT: serotonin; BBB: Basso, Beattie and Bresnahan locomotor test; BDA: biotinylated dextran amine; BMSCs: bone marrow stromal cells; CGRP - calcitonin gene-related peptide; CsA: Cyclosporine; CSPG - chondroitin sulfate proteoglycan; CST - cortico-spinal tract; DMEM: Dulbecco's modified Eagle's medium; EGFP: enhanced green fluorescent protein; FBs: fibroblasts; GDNF: glial cell line-derived neurotrophic factor; GFAP: glial fibrillary acidic proteins; GFP – green fluorescent protein; LacZ – beta-galactosidase; LP: Lamina Propria; MEP: motor-evoked potentials; MG: matrigel; MP: methylprednisolone; NF = neurofilament; OEC: olfactory ensheathing cell; OEG: olfactory ensheathing glia; OB: olfactory bulb; ONL: olfactory nerve layer; SCs: Schwann cells.
OECs derived from the olfactory bulbs of adult rodents are the most commonly studied OECs, compared to OECs derived from the lamina propria of the olfactory mucosa. Thirteen such studies were reviewed, which employed both blunt and sharp injury models of the thoracic and cervical spinal cord (thoracic contusion in three, thoracic transection in four, and partial transection in six, five of which were performed in the cervical spinal cord). None of the three thoracic contusion studies reported that OECs alone conferred a behavioral benefit (as per BBB scores) when injected into the cord in either a subacute time frame (Pearse et al., 2007; Takami et al., 2002) or a chronic setting 8 weeks after injury (Barakat et al., 2005). The combination of OECs with SCs, however, appears to promote significant behavioral benefits (Pearse et al., 2007). Takami and colleagues (2002) and Barakat and colleagues (2005) have promoted SC transplantation for SCI, and in direct comparisons between OECs and SCs, these authors have reported that the latter yielded superior behavioral outcomes.
The evaluation of OECs in complete thoracic transection injury models has been based on the rationale that OECs may promote axonal regeneration across a spinal-cord lesion site and facilitate the reentry of axons into the host at the distal host/graft interface. The study by Ramon-Cueto and colleagues (2000) gained considerable international attention due to the apparent regeneration of corticospinal axons and improvements of motor behavior in a non-standardized climbing test after 3 and 7 months post injury. When combining this OEC treatment after full transection injury with treadmill step training, the ability of rats to perform plantar stepping was further improved (Kubasak et al., 2008). Similarly, in an independent study, Cao and colleagues (2004) studied both OECs, as well as OECs modified to overexpress glial cell line-derived neurotrophic factor, which were injected into the stumps of the fully transected spinal cord. They reported significant improvements on the BBB (by six to eight points), as well as on the inclined plane test. While both the Ramon-Cueto and Cao studies reported extensive axonal regeneration of various systems (CST, RST, raphespinal, coeruleospinal), there were no differences in serotonergic fibers below the injury site in the study by Kubasak and colleagues (2008). This is only one of many studies in which the claims of robust axonal regeneration after OEC transplantation were not confirmed, and possible differences in the types of OECs utilized may be one of many potential explanations. For example, a similar study to that of Ramon-Cueto and colleagues (2000), but using primate OECs implanted into nude rats, failed to reveal any CST regeneration but a modest regeneration of 5-HT fibers (Guest et al., 2008).
In partial lesion models (seven studies to date), the transplantation of adult bulb-derived OECs appeared to improve directed forepaw reaching after dorsal column transection, as well as electrolytic lesions of the dorsal columns (Li et al., 1997; Nash et al., 2002). Whether this is due to the claimed corticospinal axon regeneration facilitated by OECs or the enhancement of plasticity and sparing in the host spinal cord is subject of ongoing debate. There is heightened awareness within the field about the potential for sparing in partial lesion models and how this may influence both the interpretation of the histological and behavioral outcomes. No significant regeneration of ascending fibers after dorsal column transection was reported by Toft et al. (2007) although there was electorphysiological evidence for preserved function in rats transplanted with olfactory cells. Regeneration of rubrospinal or corticospinal axons across and beyond the lesion site was also not found in the studies using olfactory bulb derived OECs from adult rats (Deumens et al., 2006; Ruitenberg et al., 2003) or with OECs from the mucosa of newborn mice (Bretzner et al., 2008; Lu et al., 2006). Behavioral benefits were seen when the adult bulb-derived OECs were transfected to express BDNF/NT-3 (Ruitenberg et al., 2003). This might have been due to the enhanced plasticity and neuroprotection by these trophic factors in the absence of significant regeneration.
Four studies evaluated OECs derived from the olfactory bulb of prenatal or newborn rodents (one and four studies using a blunt or sharp injury model respectively) but none reported behavioral outcomes. There is consistent evidence for axonal growth into the OEC-filled lesion sites, yet only one study demonstrated axonal growth and myelination across and beyond the dorsal column lesions that was confirmed by electrophysiology (Imaizumi et al., 2000a,b). These same authors reported similar restorations of conduction in the dorsal columns of rats when transplanting OECs from the olfactory bulbs of pigs.
The approach of transplanting pieces of olfactory nasal mucosa into a T10 full transection injury model has been championed by Lu and colleagues (2001, 2002). When transplanted acutely after injury, the authors reported that OECs improve open field locomotor scores from values of around 0–2 in controls to 6–8 in treated animals. Intriguingly, significant improvements were still observed when these transplantations were performed 4 weeks after injury (Lu et al., 2002); however, these results were not seen in a formally conducted replication study that was performed in the laboratory of Steward and colleagues (2006). It is worth mentioning that a similar protocol is already used in chronically injured humans using an autotransplantation paradigm (Lima et al., 2006). While this systematic review of cell transplantation was in review for publication, an unblinded, non-randomized study by Lima and colleagues (2010) reported improvements in 11 of 20 individuals with chronic (>18 months) SCI, including six individuals who improved from an AIS score of A to C (motor/sensory complete to motor and sensory incomplete). It must not be overlooked that this intervention was combined with a very aggressive rehabilitation regimen.
Despite the description of culture conditions for human OECs (Barnett et al., 2000) only one animal spinal cord injury study to date has used human OECs (p75-positive “OEC-like” spinal cord injury cells) harvested from the outer layers of the olfactory bulbs from human fetuses 5 to 7 months gestation (Deng et al., 2008). After a severe contusion injury, the transplantation of these cells alone was reported to improve open field locomotor scores from 6 to 11, and from 6 to 15 when combined with human bone-marrow stromal cells. These data clearly require independent replication.
Taken together, the literature on OECs contains many claims of axonal regeneration that cannot be confirmed independently by others. The reasons for these discrepancies are not fully understood, although experimental bias, variability of the cell sources and culture conditions, and animal or injury model systems are all likely contributing factors. Importantly, about two thirds (13 of 18) of the studies reported improved behavioral outcomes, yet increased autotomy was seen in some studies (Guest et al., 2008; Richter et al., 2005), which raises the question of neuropathic pain and cautions against the indiscriminative application of OECs. Further studies with human OECs are clearly warranted, but these will require a better understanding of OEC biology with which to strengthen the rationale for human translation. A summary of the pros and cons and knowledge gaps for olfactory ensheathing cell transplantation is depicted in Table 5.
Neural stem/progenitor cells (Table 6)
Adult neural stem/progenitor cells (aNPCs) are typically harvested from the subventricular zone of the brain or the spinal cord of rodents, and amplified as neurospheres in EGF and/or bFGF for several rounds of passages. They contain precursors for neurons, astroglia, and oligodendrocytes – and likely some stem-like cells with capacity for self-renewal.
Generally, NSPCs are taken from the CNS of rodent (or human aborted) embryos and expanded as neuropheres (in the presence of EGF and/or bFGF or other trophic factors) before they are dissociated and injected. These neuropheres contain precursor cells for Neurons, Astrocytes, and Oligodendrocytes plus stem cells capable of self-renewal. Heterogeneities likely exist due to some variations in the donor species, ages, passages, anatomical origins and trophic factors/culture conditions used.
d: day, days; hr: hour, hours; IH – Infinite Horizon Impactor; i.v.: intravenous; n.s.: not significant; PI: post-injury; PT: post-transplant; s.c.: subcutaneous; SCI: spinal cord injury; SD: Sprague–Dawley; sig: significant; Tx: transection; wk: week, weeks; + ve: positive; -ve: negative
5HT: serotonin; AdV: adenoviral; APC: adenomatous polyposis coli gene protein; BBB: Basso, Beattie and Bresnahan locomotor test; BDA: biotinylated dextran amine; bFGF (FGF2): basic fibroblast growth factor; BLBP: brain lipid binding protein; BMSC: bone marro stromal cell; CGRP - calcitonin gene-related peptide; ChABC: chondroitinase ABC; ChAT: choline acetyltransferase; CMEP: cortical motor evoked potential; CNP: ciliary neurotrophic factor; CNTF: Ciliary neurotrophic factor; CNP: 2'3'-cyclic nucleotide 3'- phosphodiesterase/phosphohydrolase; CMEP: cortical motor evoked potential; CsA: Cyclosporine; CSEP: cortical somatosensory evoked potentials; CSF: cerebrospinal fluid; CSPG - chondroitin sulfate proteoglycan; CST - cortico-spinal tract; DMEM: Dulbecco's modified Eagle's medium; EGF: epidermal growth factor; EM: electron microscopy; EYFP: enhanced yellow fluorescent protein; FBs: fibroblasts; G-CSF: granulocyte colony stimulating factor;GDNF: glial cell line-derived neurotrophic factor; GFAP: glial fibrillary acidic proteins; GFP – green fluorescent protein; LacZ – beta-galactosidase; MAP-2: microtubule-associated protein-2; MG: matrigel; MRI: magnetic resonance imaging; NCAM: neural cell adhesion molecule; NSPCs: Neural Stem/Precursor Cells; NPCs: neuronal progenitor cells; OPC: oligodendrocyte precursor cell; OSP: oligodendrocyte-specific protein; PDGF: platelet derived growth factor; PLGA: poly(lactic-co-glycolic) acid; RA: retinoic acid; SCs: Schwann cells; Shh: sonic hedgehog; SVZ: sub-ventricular zone.
Adult rodent NPCs were applied to thoracic contusion or compression injuries in eight rodents studies (six rat, two mice) and to cervical dorsal column transections in four rat studies. A subacute regimen was chosen in most of these studies. While some authors reported mainly astrocytic differentiation of the transplanted aNPCs (e.g., Cao et al., 2001), many authors also observed the expression of up to 60% oligodendroglial markers (Karimi-Abdolrazaee et al,. 2006; Parr et al., 2007, 2008; Pfeifer et al., 2004; Vroemen et al., 2007); expression of neuronal markers was generally rare (0–1%). The extent to which these oligodendrocytes can mature in the injured spinal cord and generate compact myelin is inconsistently reported (Karimi-Abdolrezaee et al., 2006; Parr et al., 2008).
Six of the eight contusion studies evaluated behavioral recovery with open field BBB locomotor scores. Five of these six studies reported significant improvement with the transplantation of aNPCs, three in rats (Hofstetter et al., 2005; Karimi-Abdolrezaee et al., 2006; Parr et al., 2008) and two in mice (Bottai et al., 2008; Ziv et al., 2006). However, it needs to be pointed out that in some of these studies, co-treatments were also applied, such as myelin vaccination (Ziv et al., 2006) or a cocktail of trophic factors infused intrathecally for 1 week (Karimi-Abdolrezaee et al., 2006). Of note, Hofstetter and colleagues (2005) reported an alarming lowering of sensory thresholds to non-noxious stimuli (i.e., allodynia) in the naïve aNPCs transplanted animals, illustrating the very real potential that such cells may promote neuropathic pain. Interestingly, Bottai and colleagues (2008) observed even better behavioral outcomes with intravenous compared to intra-spinal delivery of mouse aNPCs into mice – while all other studies employed a direct transplantation approach with the cells injected rostrally and caudally.
Embryonic neural stem/progenitor cells (NSPCs) are taken from the CNS of rodent embryos and expanded as neurospheres before they are dissociated and injected into the injured spinal cord. These neurospheres contain precursor cells for neurons, astrocytes, and oligodendrocytes, plus stem cells capable of self-renewal. The group is somewhat heterogeneous, as they may be taken between embryonic day 13 and 16 from various parts of the CNS (forebrain to spinal cord) and expanded in EGF or bFGF or a combination thereof, plus other potential growth factors. In addition, the number of passages varies significantly between labs, which may favor different subpopulations within the neurospheres.
Embryonic NSPCs were applied in eight studies of compression/contusion injuries in rodents at the thoracic level, in one study with weight compression at the cervical level, and in two full transection and three different partial transection models at the thoracic level of rodents. Expression of astrocyte, oligodendrocyte, and neuronal markers was observed to a variable degree in several studies. Behaviorally, the cervical weight-compression model revealed improvements on a skilled reaching task (Ogawa et al., 2002). In the seven thoracic contusion studies, behavior was reported in only three, and all observed significant improvements on the BBB locomotor scale (Meng et al., 2008; Okada et al., 2005; Setoguchi et al., 2004). In two of these studies, the effects were further enhanced with adjuvent treatments of noggin (Setoguchi et al., 2004) or bFGF expressing rat amniotic epithelial cells (Meng et al., 2008). All three studies used a subacute time frame for transplantation (7–9 days), while one direct comparison with the cells transplanted acutely demonstrated the failure of this approach. This again underlines the general notion in the field that the acutely injured spinal cord is a hostile environment for many transplanted cells, and in this regard points to an important distinction from the neuroprotection field.
Four of the five studies using sharp models of SCI reported on behavioral outcomes. Pan and colleagues (2008) claimed BBB scores of 9.6 versus 3.8 (control) after filling a complete spinal cord transection site immediately after injury (i.e., acute intervention) with eNPSC and fibrin glue from embryonic rats. Administering five injections of G-CSF over 5 days further improved the scores to 11.7. Using the same model, Guo and colleagues (2007) reported BBB scores of 3.6 after transplantation of NSPCs from neonatal rats plus type1 collagen compared to 0.54 in controls. In the latter study, the benefit was greatly enhanced by co-transplantation with SCs from neonatal rats, especially when these were transduced to express NT-3 (BBB ∼ 10.7).
The transplantation of eNSPCs from human fetuses at 8 weeks of gestation into cervical contusion sites of marmoset monkeys (Iwanami et al., 2005) is interesting from a translational perspective for both the human source of cells and the primate model of cervical injury. Expression of astrocytic, neuronal, and a small percentage of oligodendrocyte markers was observed. Behaviorally, bar grip power and spontaneous motor activity was improved, which is promising, although validation of these test models is still pending. Given the ethical controversy around the use of human abortion material, as well as the technical variability and logistical problems involved, several authors have pursued human immortalized neural stem-cell lines (HB1.F3 clone; line K048) or long-term human neurosphere cultures (Cummings et al., 2005) and transplanted them to dogs, mice, and rats. However, only two of the four studies listed were met with behavioral success (Cummings et al., 2005), including the transplantation into dogs (motor scores of 15 vs. 10) (Lee et al., 2009). It is conceivable that eventually these approaches will yield viable sources of human cells for clinical translation. A general summary of the pros and cons and knowledge gaps for neural stem-cell transplantation is depicted in Table 7.
Neural and glial restricted precursors (Table 8)
The nine studies of glial restricted precursors (GRPs) and/or neural restricted precursors (NRPs) included here were performed with rodent cells transplanted into the injured rodent spinal cord – six of these employed blunt contusion models. While transplantation of NRPs alone into uninjured spinal cords resulted in neural differentiation, such neuronal differentiation is far less complete in the environment of the SCI site, underlining the fact that the environment of the injured spinal cord inhibits neuronal differentiation (Cao et al., 2002). Similarly, GRPs differentiate mainly into astroglial cells in the lesion centre, while only some express oliogdendrocyte markers, usually after they migrate into the spared host spinal cord (Enzmann et al., 2005; Han et al., 2004; Hill et al., 2004). Still, the degree to which GRPs form myelinating oligodendrocytes in the contused spinal cord is somewhat limited (Enzmann et al., 2005). Furthermore, it appears that behavioral recovery requires the transduction with the neurotrophin D15A (Cao et al., 2005), which has BDNF and NT-3 activities and also enhances oligodendrocyte differentiation. The extent to which the observed benefits are related to increased myelination, neuroprotection, or neural plasticity can only be speculated on.
These cells are isolated from embryos and rather than propagating them as neurospheres they are immunoselected (immunopanned) with antibodies to select glial precursors (A2B5 antibody for a tripotential glial precursor or O-2A for oligodendrocyte precursors) or neuronal precursors (PS-N-CAM). The main rationale behind the use of the GRPs is to replace lost oliogdendrocytes in order to remyelinate demyelinated axons in the spared host spinal cord. In addition, these cells may provide neuronal replacement and deliver trophic factors for neuroprotection and enhancement of plasticity.
d: day, days; hr: hour, hours; IH – Infinite Horizon Impactor; PI: post-injury; PT: post-transplant; s.c.: subcutaneous; SCI: spinal cord injury; SD: Sprague–Dawley; Tx: transection; wk: week, weeks; + ve: positive; -ve: negative.
5HT: serotonin; BBB: Basso, Beattie and Bresnahan locomotor test; BDNF – brain-derived neurotrophic factor; ChAT: choline acetyltransferase; CRF: corticotrophin releasing factor; CsA: Cyclosporine; CSPG - chondroitin sulfate proteoglycan; CST: corticospinal tract; eNCAM: embryonic neural cell adhesion molecule; EGFP: enhanced green fluorescent protein; DH:dorsal horn; DLF: dorsolateral funiculus; ESC: embryonic stem cell; FACS: fluorescence activated cell sorting; FG: fluorogold; FGF: fibroblast growth factor; GFAP: glial fibrillary acidic proteins; GFP – green fluorescent protein; GRP: glial restricted precursor cell; LF: lateral funiculus; LP: lumbar puncture; MEP: motor-evoked potentials; MP: methylprednisolone; NCAM: neural cell adhesion molecule; NPC: precursor cell; NRP: neural restricted precursor cell; PLAP: placental alkaline phosphatase; Shh: sonic hedgehog; SPN: spinal parasympathetic nucleus; SSEP: somatosensory evoked potentials.
Four studies transplanted a mixture of GRPs and NRPs from rodent embryos, and in two experiments this was performed in the context of severe thoracic contusion injuries. Both cases reported moderate but significant improvements on the BBB scale from ∼7 to ∼9; in addition, bladder control was improved and the hypersensitivity to thermal stimuli ameliorated (Mitsui et al., 2005; Neuhuber et al., 2008). The cells were neuroprotective, and many differentiated into astrocytes, some into oligodendrocytes.
The isolation and transplantation of oligodendrocyte precursors from newborn rodents (as opposed to embryos) using antibodies to A2B5 or O-2A resulted in improved BBB scores after mild as well as after moderate contusion injury (Bambakidis and Miller, 2004; Lee et al., 2005). Both studies reported reduced latencies of motor evoked potentials, consistent with either remyelination and/or neuroprotection.
From a translational perspective, harvesting human GRPs and NRPs from abortion materials is met with logistic and ethical concerns in many countries. Hence, alternative sources for oligodendrocyte precursors have been pursued. Most prominent is the differentiation of oligodendrocyte precursors (OPC) from a human ES-cell line, an approach that received FDA approval to proceed with a Phase 1 clinical trial in January 2009, but was subsequently put on hold (Geron Corp, Menlo Park, CA). In essence, these ESC-derived OPCs enhance myelination, are neuroprotective, and they appear to mediate moderate improvement of locomotor function when transplanted after subacute but not after chronic SCI (Keirstead et al., 2005). This study has not been independently replicated by other laboratories, although Geron performed extensive “in-house” safety and efficacy studies prior to obtaining FDA approval. Such studies further characterizing the efficacy of this technology have yet to be released to the academic community. Efficacy in blunt cervical models would be desirable if that will be a major human target for translation. Similarly, no larger animal models with OPC transplants exist so far. Concerns regarding the risk of teratoma formation have been voiced. A summary of the pros and cons and knowledge gaps of GFP/NRP transplantation is given in Table 9.
Bone-marrow-derived stromal cells – mesenchymal stem cells (Table 10)
The stromal cells from bone marrow are isolated and separated from the hematopoietic cell fraction of the bone marrow by their property to adhere to plastic. Some authors go further by using FACS to purify hematopoietic cells (which are CD34 positive). Bone-marrow-derived stromal cells (BMSCs) are hence typically a crude mixture of stromal cells that support the growth of hematopoietic stem cells and mesenchymal stem cells, and some authors do provide additional (albeit somewhat unspecific) markers to characterize these mesenchymal stem cells. This heterogeneity and uncertainty of origin likely explains the highly variable results among different laboratories regarding the ability of these cells to survive, integrate, and differentiate as neural cells in the injured spinal cord. In addition, there is evidence that rather non-specific treatments can induce the expression of a neuronal marker without truly specifying these cells as neuron or glial cells (Lu et al., 2004).
The stromal cells from the bone marrow are isolated and separated from the hematopoetic cell fraction of the bone marrow by their predilection to adhere to plastic. Some authors exclude the contamination by hematopoetic cells (which are CD 34 positive) by FACS. BMSCs are hence a crude mixture of stromal cells which support the growth of hematopoetic stem cells and mesenchymal stem cells. Some authors provide some additional (somewhat unspecific) markers to characterize these mesenchymal stem cells. Hence the actual stromal versus mesenchymal stem cell nature of the transplanted cells is unclear in many studies.
d: day, days; hr: hour, hours; PI: post-injury; IH: Infinite Horizon Impactor; i.v.: intravenous; PT: post-transplant; s.c.: subcutaneous; SCI: spinal cord injury; SD: Sprague–Dawley; Tx: transection; wk: week, weeks; + ve: positive; -ve: negative.
5HT: serotonin; APC: adenomatous polyposis coli gene protein; BBB: Basso, Beattie and Bresnahan locomotor test; BDA: biotinylated dextran amine; bFGF: basic fibroblast growth factor; BMSC: Human Bone Marrow Stromal Cell; CFDA-SE: carboxy fluorescein diacetate; CGRP - calcitonin gene-related peptide; ChAT: choline acetyltransferase; CNP: 2', 3'-cyclic nucleotide 3'-phosphodiesterase/ phosphohydrolase; CsA: Cyclosporine; CSEP: cortical somatosensory-evoked potential; CSPG - chondroitin sulfate proteoglycan; CST: corticospinal tract; EGF: epidermal growth factor; DMEM: Dulbecco's modified Eagle's medium; DRG: dorsal root ganglion; FLIP: FLICE-inhibitory protein; GAP-43 - growth associated protein-43; G-CSF: granulocyte colony stimulating factor; GFAP: glial fibrillary acidic proteins; GFP – green fluorescent protein; hPAP: human placental alkaline phosphatase; HSCs: hematopoetic stem cells; ICAM-1: intercellular adhesion molecule-1 (CD54); IHC: immunohistocemistry; LacZ – beta-galactosidase; LV: lentiviral vectors; MAP-2: microtubule-associated protein-2; MG: Matrigel; MP: methylprednisolone; MRI: magnetic resosnance imaging; NCAM: neural cell adhesion molecule; NF: neurofilament; NSE: neuron specific enolase; NGF: neural growth factor; OEC: olfactory ensheahting cell; NGF: nerve growth factor; PARP: poly [ADP- ribose] polymerase; RA: retinoic acid; SC: stem cells; Sca-1: stem cell antigen 1; SSEP: somatosensory evoked potentials; tcMMEP: transcranial magnetic motor evoked potentials; TH – tyrosine hydroxylase; UCBCs: umbilical cord blood cells; XIAP: X-linked inhibitor of apoptosis protein.
A narrative review of BMSC transplantation in TBI, stroke, and SCI has recently been published by Parr and colleagues (2007). Nine SCI studies were performed with human BMSC, of which six used a blunt contusion or compression model. Three of these report beneficial behavioral effects, while three groups observed no transplant-related improvements. Deng and colleagues (2008) claimed impressive BBB scores of 13 (weight-supported stepping with frequent coordination) versus a BBB of 6 in their controls. However, Kim and colleagues (2006) found less dramatic benefits (13 vs. 10) in a milder contusion model when combining these cells with FGF. Similarly, Cizkova and colleagues (2006) reported benefits after balloon compression. The behavioral benefits in the sharp models are questionable, since in the studies by Mansilla and colleagues (2005), as well as the study by Zhao and colleagues (2004), a large percentage (80% and 35% respectively) of control animals died (but not in the treated groups), which suggests suboptimal animal care standards (and leading to exclusion of the former study from this review). Neuhuber and colleagues (2005) tested human BMSC from four different donors and found highly variable outcomes in a rat hemisection model using various tests, which illustrates the heterogeneity of these cells. Hence, it appears that we need a better understanding of the types of cells in the BMSC fraction that might mediate these benefits.
Unfortunately, the rodent-to-rodent transplantations do not provide more insights regarding the cell characteristics. Twenty-two studies with rodent BMSCs employed blunt injury models (17 contusions) in mostly rat studies (only two murine studies). Given the perceived relevance of blunt models to support a rationale for clinical translation, these studies will be discussed. A large proportion of these 22 studies reported positive behavioral effects (12 studies), while six did not show behavioral data and four failed to see benefits. It must not be overlooked that the transplantation of any cell may confer benefits over saline or media injections, and this is rarely controlled for by using additional cell types as controls (e.g., fibroblasts). In most studies, the cells were injected directly into or next to the SCI site, yet, in some hands, intrathecal (Ohta et al., 2004) and even intravenous (Urdzikova et al., 2006) delivery seems successful. Other researchers did not have success with intravenous delivery (Fan et al., 2008). Most studies used a subacute or acute timing for the transplantation, except Zurita and Vaquero (2004, 2006) and Vaquero and colleagues (2006) who delayed the treatment to 3 months after spinal-cord contusion by weight drop at T6–8. These authors allowed the rats to survive for up to 12 months, and reported improvement to a BBB of around 17, whereas the control animals were completely paralyzed throughout (BBB = 0). Such poor performance in controls is highly surprising, as even rats with completely transected spinal cords typically score a few points on the BBB. Nevertheless, these long survival times should be considered, as most rodent studies are terminated around 6–8 weeks when the performance is deemed to reach a “plateau.”
In the light of the widely observed behavioral benefits, it is somewhat surprising that the histological data are very divergent. Histological observations of these cells range from good survival and differentiation of BMSC into neural cells, to poor survival and no differentiation into neural cells. Claims of differentiation are less credible when in-vitro dyes have been used to label the transplanted cells (e.g., the chromatin stain Hoechst; see Guest et al., 2008, in OEC section). Still, the heterogeneity of histological results once more underlines that beneficial behavioral effects can be brought about by multiple factors. These range from neuroprotection (via secretion of trophic factors and modification of inflammation) to the recruitment of endogenous cells, including stem cells and remyelinating cells, and – although hotly debated – the differentiation and integration of neural cells originating from the transplant. Indeed, several studies reported more preserved white matter or less cell death, indicative of neuroprotection (Ankeny et al., 2004; Bakshi et al., 2006; Dasari et al., 2007; Ohta et al., 2004; Urdzikova et al., 2006), while another could not confirm these effects (Yoshihara et al., 2006).
The claims of axonal regeneration in contusion studies can only be partially interpreted within the site of the lesion but not the host spinal cord itself where spared axons and regenerated axons are not easily distinguishable. Such questions are better addressed in sharp models of SCI, and 10 studies transplanted BMSCs into fully (n = 3) or partially (n = 7) transected spinal cords. Interestingly, two of the three studies with full transection reported behavioral improvements on the BBB score from 3 to 7 (Kamada et al., 2005; Koda et al., 2007). While this may be due to some axonal regeneration, other mechanisms like trophic effects on spinal circuits below the level of the injury site cannot be ruled out. BMSCs do promote axonal growth and these effects may be attributed to invading SCs; axon growth can be greatly enhanced with co-expression of trophic factors by the transplanted BMSCs (Lu et al., 2007).
From a translational perspective, BMSCs are the most widely studied cells using rodents, large mammals, and primates. This and the easy access to BMSCs for auto-transplantation explains their use in several human treatment studies using a mixture of BMSCs and hematopoietic (mononuclear) cells (Callera et al., 2006; Chernykh et al., 2007; Saito et al., 2008; Yoon et al., 2007). Unfortunately, these reports included small patient cohorts only, used uncharacterized mixtures of bone-marrow cells, and were mostly not controlled. Hence, a systematic clinical validation is needed. A summary of the pros and cous and knowledge gaps of BMSC transplantation for SCI is given in Table 11.
Discussion
The potential to bridge the injured spinal cord and repopulate the area of injury with cells that might restore axonal continuity, bridge the area for axonal regeneration, and promote axonal growth back to its distal targets has fascinated SCI researchers for decades. At first, it would seem to be a relatively simple task: fill the inhospitable area of parenchymal devastation with growth-promoting cells, and let them “do their thing.” The exciting discovery that CNS axons regenerated quite robustly into peripheral nerve grafts and SC environments was followed by the difficult realization that they were also rather reticent about reentering the host CNS. Excitement around the potential to overcome this barrier with OECs was accompanied by explosive interest in “stem cell” candidates that might have this and other therapeutic capabilities. Around the world, the enthusiasm of clinicians and desperation of patients thrust cell transplantation approaches into the translational spotlight, as individuals with cord injuries traveled around the globe to receive these technologies (at substantial financial cost). The announcement of FDA approval to conduct a stem-cell-based trial in North America in January 2009 ignited tremendous excitement, only to be put on hold some 6 months later for safety concerns.
Along the way, a great deal has been learned, and this should not be overlooked. It has become appreciated that the cells themselves may be the source of growth factors that can positively influence the environment. The initial goal of inducing rampant long-distance axonal regeneration has been dampened to a large degree by the realization that remyelination of demyelinated axons may be the most realistic therapeutic objective, although some authors believe that endogenous remyelination is effective albeit somewhat slower. Advances in cell biology have led to more sophisticated strategies of modifying cells with certain genetic traits, and developing purified sources of cells. The field has learned a great deal about evaluating what cells do once implanted into the cord: How many survive? Do they integrate and migrate? What do they associate with? How do they influence the host environment?
Alas, amongst this undeniable progress are significant questions, and a systematic review of the literature on this therapeutic approach exposes many gaps in our current knowledge. As an overview, one can quickly appreciate that while many labs around the world are studying cell transplantation therapies, substantial heterogeneity and uncertainty exists around the very nature of the cells that they are studying. This is perhaps best exemplified in the BMSC literature, but is equally perplexing in the discussion of precursor cells and stem-cell-based approaches. While our approach in this systematic review groups these under “umbrella” subtypes, the differences in source and culture conditions between a cell from one lab and the “same” cell from another may make them quite distinct in important ways. It is, unfortunately, far more complex than obtaining minocycline from Sigma, preparing it according to the manufacturer's protocol, and infusing a standard dose into a rodent after an acute SCI.
In this regard, while this review found many studies under each “umbrella” cell type, the actual pre-clinical substantiation for many cells within those subtypes seems quite modest. The “knowledge gaps” are fairly applicable across all of the cell types: the lack of independent replication (although this is inherently more difficult for cells than for pharmacologic treatments), the relative majority of laceration-type injuries over the more clinically relevant contusion injuries, the paucity of work in large animal models, and importantly, the near absence of chronic injury work. Given that patients with chronic injuries are the typically the most vocal proponents of cell transplantation therapies, and are the ones travelling around the world to be subjected to these unproven treatments, it is remarkable that so little has been reported in models of chronic injury. For those who have tackled this problem in pre-clinical studies, the results have generally not been very promising. This remains a daunting yet important task for the future, as chronically injured individuals with SCI will continue to be the most vocal “consumers” of this technology. Many of these chronically injured people seeking a cell transplantation treatment are likely to be unaware that the pre-clinical studies done at chronic time points on that particular cell have not been performed, or have shown negative results. Ironically, even for clinicians and scientists, they themselves might be surprised to note the gap between what they think “should” be done in pre-clinical studies before moving a cell transplant treatment into clinical trials and what is actually occurring in the field (or has happened in the past) (Kwon et al., 2009).
Despite these misgivings, the interest in cell transplantation for SCI will remain high, and it seems quite likely that FDA-sanctioned and closely regulated trials will be initiated in the foreseeable future. It is hoped that this systematic review has illustrated the pervasive interest in cell transplantation treatments for SCI, the experimental heterogeneity that inherently comes from such widespread interest, and has revealed both the promise and the knowledge gaps in these approaches, as they stood in the summer of 2008. The field will obviously continue to evolve, with hopes that further refinement and understanding will increase the chances that cell transplantation will someday emerge as a fruitful treatment for patients.
Footnotes
Acknowledgment
This review was supported by the Canadian SCI Solutions Network (Now Rick Hansen Institute). BKK holds a CIHR New Investigator Award, and WT is the Rick Hansen Man in Motion Chair in Spinal Cord Injury Research.
Author Disclosure Statement
No competing financial interests exist.