Current Status of Spinal Cord Regenerative Therapies: A Review

Abstract

Spinal cord injury (SCI) is any injury resulting from an insult to the spinal cord that disrupts its major functions, either completely or incompletely, and it can be caused by both traumatic and nontraumatic events. The number of SCI patients has been continuously increasing due to increasing number of motor vehicles and average age of patients is constantly decreasing. After SCI, nerve cells located at the injured site are severely damaged and eventually die, and these dead cells are cleared away by the immune system and in turn a cavity remains. Unfortunately, till date no effective treatment strategy exists ensuring functional recovery after SCI. There is an imperative need for the development of therapies to reduce the enormous physical and financial burdens of people afflicted with SCI. The surgical treatment has been known to aid in rehabilitation, but cannot substantially improve neurologic and functional outcome after SCI. The stem cell–based therapy has been proposed as a promising treatment strategy for SCI. Many of the current strategies for treatment of SCI involve replacing the cells lost to injury with cells derived from alternative sources, such as Schwann cells, oligodendrocyte precursor cells, and neural stem cells. This review discusses the present status of various cell-based therapies, which are being used for treating SCI.

Keywords

stem cell - spinal cord - regeneration - trauma - injury

Introduction

Spinal cord injury (SCI) is defined as any injury resulting from an insult to the spinal cord that disrupts its major functions, either completely or incompletely.[1] Spinal cord injuries can be caused by both traumatic and nontraumatic events. The leading causes of traumatic SCI include motor vehicle accidents, falls, sport-related injuries, industrial accidents, gunshot wounds, and assault. Nontraumatic causes of SCI include stroke, infections, degenerative joints/bone disease, neoplasms, and other neurologic conditions, such as multiple sclerosis.[2] [3] [4]

The number of patients suffering from SCI is continuously increasing, and each year an estimated 180,000 individuals over the world sustain new injury.[5] In the United States, the incidence of SCI has been estimated to be approximately 40 cases per million people per year or around 12,000 cases per year with around 250,000 individuals living with SCIs.[6] [7] The average age at the time of injury has gradually increased from a reported 29 years in the mid-1970s to a current average of around 40. Men are more at risk for SCI than women.[8] [9]

The spinal cord contains millions of nerve cells, which are organized into multiple tracts to convey electrical signals between the brain and the rest of the body. Once the spinal cord is injured, these nerve cells located at the injured site are severely damaged and eventually die. The dead cells are cleared away by the immune system and in turn a cavity remains.[10] The glial scar tissue that forms around the cavitation prevents axonal regeneration and, as a consequence of this, no more electrical signals can be transmitted, resulting in loss of motor, sensory, and autonomic functions.[11] Many of these injuries result in partial or complete paralysis, and only less than 1% of people are able to recover complete neurologic function.

Unfortunately, till date there has been no effective treatment strategy ensuring functional recovery after SCI. There is an imperative need for the development of therapies to reduce the enormous physical and financial burdens of people afflicted with SCI.

There have been various drug treatment strategies for SCI, but none of them, other than methylprednisolone, have achieved a substantial clinical usage. The surgical treatment has been known to aid in rehabilitation, but cannot substantially improve neurologic and functional outcome after SCI. There have been several experimental studies in animal models, involving strategies such as local spine cooling and oscillating field stimulation aimed at reducing the paralyzing effects of injury and promoting regrowth of functional nerve fibers.[12] Several these experimental treatments have now reached the stage of controlled human trials.[13] [14] The stem cell–based therapy has been proposed as a promising treatment strategy for SCI. Many of the current strategies for treatment of SCI involve replacing the cells lost to injury with cells derived from alternative sources, such as Schwann cells (SCs), oligodendrocyte precursor cells (OPCs), and neural stem cells (NSCs).[15] However, none of these developments have reached even limited use in the clinical care of human SCI.[16] Currently, interventions to promote recovery from SCI using the cellular regeneration therapies involve stem cell transplantation with an aim to supply new cells to replace lost ones and to optimize the spinal cord for natural recovery. [Fig. 1] shows schematic flow of how stem cell could potentially promote recovery after SCI. Furthermore, evidence continues to show that these implanted cells are able to induce neuronal plasticity and modify their microenvironment by promoting axonal elongation, collateral sprouting, remyelination, and synapse formation.[17]

Fig. 1 Schematic illustration showing the stem cell differentiation during recovery after SCI.

This review aims to discuss the present status of various cell-based therapies, which are being used for treating SCI.

Various cells used in the regeneration in spinal cord injuries are discussed as follows.

Schwann Cells

Schwann cells (SCs) produce the myelin sheath in neurons. The transplanted SCs play a key role in regeneration of axons after injury, by secreting a variety of growth factors, including neurotrophin-3 (NT-3), nerve growth factor (NGF), brain-derived nerve factor (BDNF), ciliary neurotrophic factor (CNTF), and fibroblast growth factor (FGF) that contribute to neuronal survival, generation of cell adhesion molecules, and extracellular matrix proteins that support axonal growth. The SC also migrates to the site of injury after SCI, despite being part of the peripheral nervous system, and they can be easily expanded in vitro, allowing for autologous cell transplantation to avoid immune response.[18]

Xu et al used nerve guidance conduits (NGCs) containing SC in a complete transaction injury rat model and concluded that the group with NGCs containing SC showed increased remyelination as well as propriospinal and sensory axonal regeneration as compared with controls.[19]

A few other studies have looked at genetically modifying Schwann cells to secrete increased amounts of growth factors, specifically NGF and BDNF.[20] [21] [22] Although the genetically modified cells promoted more axonal regeneration than unmodified Schwann cells, no functional recovery was observed with either cell line. Additionally, several studies have used Schwann cells in combination with other strategies for treatment of SCI. Chondroitinase ABC enhances Schwann cell–induced axonal regeneration, but it does not lead to functional recovery, as studied in T9 dorsal hemisection injury model.[23] Combining cyclic adenosine monophosphate (cAMP) with Schwann cells as therapy for treatment of a T8 mild contusion injury resulted in functional recovery as assessed by the Basso-Beattie-Bresnahan (BBB) score.[24] This study used an inhibitor against the phosphodiesterase that hydrolyzes cAMP to extend its signaling, which allows the neurons to extend into otherwise inhibitory substrates.

Thus, SC works best when used in combination with other therapeutic approaches that minimize the inhibitory nature of the lesion site.

Embryonic Stem Cells

Embryonic stem cells (ESCs) are pluripotent cells found in the blastocyst. They have a capability of differentiating into all three primary germ cell layers and generating all cell types, including oligodendrocytes, astrocytes, or neurons, making ESCs a perfect candidate for cell therapy.[25] However, there have been ethical considerations, surrounding the use of human ESCs, as obtaining them require the destruction of several human embryos or fertilized oocytes.

Mouse ESCs have been investigated as a starting point for developing potential therapies for SCI. These cells are attractive for preclinical studies because they are easy to culture and readily differentiate into cells found in CNS. One of the first studies that looked at using mouse ESCs for treatment of SCI involved a multistep process that promoted the formation of oligodendrocytes and astrocytes for transplantation.[26] Once transplanted into the spinal cord, these cells demonstrated the ability to restore myelination in myelin-deficient shiverer rats, indicating the potential of mouse ESCs to treat SCI.

Other studies have demonstrated that mouse ESCs can survive in the spinal cord for more than 50 days, suggesting their potential as a long-term treatment for SCI.[27] A more recent study showed that mouse ESCs pretreated with a lecithinized BDNF implanted 9 days after T9/10 contusion injury showed better differentiation and promoted increased functional recovery afterward, as assessed by BBB compared with cells treated with normal BDNF, demonstrating the influence of growth factor treatments on ESC differentiation.[28] Recent work has looked at different types of genetically modified mouse ESCs and their impact on SCI. Mouse ESCs were modified to overexpress the bcl-2 protein, which blocks apoptosis, and implanted 9 days after contusion injury.[29] However, these cells did not produce an increase in functional recovery and resulted in tumor-like growths. This study illustrates the importance of selecting the appropriate proteins when genetically modifying cells for transplantation.

Another study examined the effect of transfecting mouse ESCs with the L1 adhesion molecule, which promotes neuronal outgrowth, to determine the effect on cell behavior.[30] These cells were injected both rostral and caudal to the injury site 7 days after a T7–9 compression injury. After 1 month, the cells transfected with L1 showed increased survival and migration compared with untransfected ESCs. Overall, mouse ESCs serve a model cell culture system for determining effective strategies for treatment of SCI due to the ease of culture and ability to be genetically modified. These strategies provide a starting place for translating such methods to human ESCs.

Development of methods and strategies for using human ESCs for treatment of SCI allows for translation to clinical studies. One of the challenges of developing human ESC therapies is determining the necessary cues to promote differentiation in the desired cell types, such as oligodendrocytes and motor neurons. Although the work done with mouse ESCs provides a good starting point, the end goal is to determine human ESC differentiation protocols. Similar to the mouse ESCs, testing the ability of human ESCs to promote recovery in rat injury models requires immunosuppression. Many of the human ESC lines that currently exist have been cultured in the presence of mouse feeder cell layers, making them undesirable for use in clinical trials. Other major challenges include developing person-specific ESC lines to allow transplantation without immunosuppression in a clinical setting.

Keirstead laboratory developed protocols that produce large numbers of oligodendrocytes from human ESCs.[31] [32] [33] [34] Their time-intensive differentiation protocol involves multiple selection steps with serum-free media to produce OPCs that can differentiate into mature oligodendrocytes in vitro and in vivo.[31] They implanted these cells into myelin deficient shiverer mice 1 week after T10 contusion injury. The transplanted cells integrated into the spinal cord, differentiated into oligodendrocytes and restored myelination.[32]

The clinical applications of human embryonic stem cell (hESC) critically depend on their ability to differentiate toward defined and purified neural cell types in vitro. Recently, considerable progress has been achieved in improving the methods for differentiation of hESC into neural or neuronal precursors prior to cell transplantation in animal models of SCI.[35] [36] [37] [38] Despite promising results obtained in preclinical studies, the challenges in using ESCs include determining the correct cues to direct differentiation in the specific desired cell types in vitro and in vivo.[39]

Mesenchymal Stem Cells/Bone Marrow Stem cells

The bone marrow (BM) is a source of stem cells and it is the only organ where two types of stem cells can be found: coexisting and cooperating. The hematopoietic stem cells (HSCs) produce blood cells, whereas mesenchymal stem cells/mesenchymal stromal cells (MSCs) can be found in the stromal layer adjacent to hematopoietic cells and support hematopoietic cells, regulating the microenvironment and facilitating the maturation of blood cells. The MSCs can also be isolated from other regions such as adipose tissue, neonatal tissue (such as umbilical cord and placenta), and fetal tissues such as the lungs, liver, and blood.[40] From a clinical perspective, MSCs are attractive for transplantation because they are easily obtained from BM and can be transplanted back into the original donor, eliminating the risk of rejection.

In 2006, Moviglia et al reported a combined treatment with MSCs and autoimmune T (AT) cells in two SCI patients. The main goal behind this approach is to control inflammatory activity to create the ideal microenvironment for cell transplantation, as it has been shown that AT cells are essential for tissue repair. Both patients reported motor improvement and no adverse effects.[41]

Pal et al transplanted autologous MSCs via lumbar puncture in 30 patients of SCI. There were no significant differences in the MRI scans taken at baseline and at the 1-year follow-up. Similarly, somatosensory evoked potentials (SSEPs), motor evoked potentials (MEPs), and nerve conduction velocity (NCV) measurements revealed no significant changes. Despite these negative electrophysiologic findings, patients reported significant recovery: initially, the return of bladder sensation, followed by bladder regulation and bowel function, then improvement in sensory perception, and finally in motor function.[42]

Another study compared the transplantation of MSCs to the conventional treatment in SCI, and confirmed the safety and feasibility of using this technique via lumbar puncture; however, even though recovery was reported, it was not found to be statistically significant.[43]

Mazzini et al studied 19 patients of SCI with a goal to study and evaluate the safety and feasibility of MSC transplantation. All patients were monitored for adverse effects of therapy and magnetic resonance imaging (MRI), and tractography suggested no mass at the injection site or within the neuraxis, no syringomyelia or pseudomeningocele formation, and suggested the therapy to be safe. However, the patients did not report any improvement in the quality of life after either form of therapy.[44]

Hofstetter et al transplanted BMSCs into T7 contusion injury model of SCI both immediately following injury and 7 days postinjury.[45] The cells transplanted 7 days postinjury showed better rates of survival and formed bundles that bridged the lesion. Implantation of BMSCs also led to an increase in BBB scores compared with controls.

Various other studies have looked at the feasibility of different methods of intravenous injection for BMSC delivery,[46] [47] [48] but a study by Vaquero et al showed that direct injection of BMSCs into the site of a T6–8 contusion injury promoted superior functional recovery when compared with intravenous injections of BMSCs.[49]

Additional work has been done to clarify the mechanisms by which BMSCs promote functional recovery. One study demonstrated that BMSCs help guide regenerating axons across the injury site when implanted 2 days after a T8 contusion injury and can help promote recovery by restoring the stepping control circuitry.[50] A more recent study showed that BMSCs express the gamma aminobutyric acid (GABA) receptor.[51] BMSCs also stimulate phosphoinositide-3-kinase and mitogen-activated protein kinase signaling in neurons, which promotes their survival.[52] All of these mechanisms contribute to the success of BMSC transplantation as a treatment for SCI.

Olfactory Ensheathing Cells

The neurogenesis in olfactory system continues to take place throughout a person's life. The stem cells proliferate to generate new sensory neurons in the basal layer of olfactory epithelium. The stem cells proliferate in the subventricular zone (SVZ) of the forebrain, generating neural progenitors, which migrate to the olfactory bulb to create new interneurons, within the central nervous system (CNS). In case of injury, these neurons are immediately replaced through an increase in neurogenesis.

Because of their ability to guide the connections between the peripheral nervous system (PNS) and the CNS, and to their ability to differentiate into nonolfactory cell types, these multipotent cells are excellent candidates for cell transplantation.[53] OECs can be harvested and then transplanted back into the original human donor, eliminating rejection. Currently, phase I clinical trials using autologous OEC transplantation are underway where these cells have been demonstrated to be safe for up to 1 year after treatment.[54]

There have been several studies in literature supporting the use of OECs reporting improved outcomes in both experimental models[55] [56] [57] [58] [59] and clinical settings.[60] [61]

The use of genetically modified OECs with the aim of obtaining any additional benefit of recovery in SCI is being considered. The OECs are modified to secrete various growth factors such as NT-3, BDNF, and GDNF.[62] [63] The growth factor–secreting OECs have been found to promote the tissue sparing and improve functional recovery after SCI.[63]

Olfactory ensheathing cells can therefore be considered as a promising source to promote regeneration in cases of SCI. Another advantage is that they can be used as autografts. Nevertheless, several preclinical and clinical trials with high number of patients using olfactory ensheathing cells need to be performed to consider the possibility of using OECs as an effective treatment for SCI.

Neural Stem Cells

First described by Altman in 1960, these multipotent cells have the potential to transform into any cell type in the CNS. Neural stem cells (NSC) are remnants from neuroectoderm of early embryos, and are present in embryonic, fetal, and adult nervous systems. During development, these cells divide and differentiate to form the main components of the CNS, that is, the brain and the spinal cord. During adulthood, stem cells reduce and are confined to certain specific regions, such as the spinal cord, and, to a greater extent, SVZ and subgranular zone (SGZ) of the hippocampal dentate gyrus.[64]

Several studies have investigated the potential of NSCs derived from humans to promote recovery in animal models of SCI. One of the first studies looked at the ability of human NSCs derived from fetal brain cultured as neurospheres to promote recovery after T9 contusion injury in both severe combined immunodeficiency (SCID) and myelin-deficient shiverer mice.[65] These cells differentiated into functional neurons and oligodendrocytes while promoting an increase in functional recovery as assessed by BBB scores.

Other studies have reported good outcomes by using NSCs in combination with other strategies to treat SCI. Teng et al seeded mouse NSCs within polymer scaffolds[66] and implanted them into the lesion site resulting from a T9/10 hemisection. The authors reported good functional recovery as indicated by an increase in BBB scores as compared with animals receiving only cells, even though the transplanted cells did not stain positive for mature cell markers, indicating that they remained undifferentiated.

Some studies suggested that NSCs, genetically modified to produce neurotrophin-3 (NT-3), enhanced the axonal sprouting in animals.[67] Others have used NSC in conjunction with anticiliary neurotrophic factor (CNTF) antibodies, myelin-specific T cells, fibroblasts, combination of growth factors, heparin, laminin, and FGF, and found that the combination of cells was able to produce functional recovery as evidenced by an increase in the BBB.[68] [69] [70] [71] [72] [73]

Fibroblasts

Fibroblasts secrete extracellular matrix molecules, such as collagen and glycoproteins, and are easy to obtain and are expandable in culture, thereby making them attractive for use as a cellular therapy. Fibroblasts are also easy to genetically modify, allowing for additional functionality. However, the method of transfection should be carefully considered. Some transfection agents only produce transient expression of the target protein whereas other methods, such as lentiviral transfection, produce stable protein expression, but may cause unwanted mutations upon integrating into the chromosomal DNA of the fibroblasts.

Many studies have been performed to determine the effect of transplanting fibroblasts that overexpress neurotrophins, including NT-3, BDNF, and NGF.[74] [75] The growth factor–secreting fibroblasts have been found to promote axonal growth into the injury site as compared with unmodified fibroblasts.[76] [77]

Many other groups have also investigated the use of fibroblasts as a potential treatment for SCI. McTigue et al investigated the effects of several growth factors, including NT-3, BDNF, CNTF, NGF, and basic FGF, secreted by genetically modified fibroblasts on regeneration after SCI.[78] The cells modified to secrete NT-3 and BDNF promoted proliferation of endogenous oligodendrocytes and remyelination of damaged axons, advocating the efficacy of using fibroblasts in SCI.

Human Umbilical Cord Blood Stem Cells

Human umbilical cord blood stem cells (HUCBCs) have been shown to have the following properties: They have the potential to differentiate in vitro into cells that are morphologically similar to oligodendrocytes and express oligodendrocyte markers; they secrete factors that prevent further injury; have tropism for the injured area in the spinal cord; can be effective even through remote infusion either by intravascular or intrathecal administration; and improve neurologic function in animal studies.[79] Therefore, the clinical application of HUCBCs to treat SCI is very appealing. In cases of SCI, UCBC could, thus, control apoptosis, demyelination, and scar formation.[80]

Transplantation of CD34 (+) HUCBCs in traumatic SCI rats significantly increased BBB score and the infarct size and blood vessel density at the injured site. However, the transplanted cells survived at least 3 weeks at the injured site, but did not differentiate into neural cells.[81] In a study conducted by Yao et al, 25 patients with traumatic SCI (injury time > 6 months) were treated with HUCBCs via intravenous and intrathecal injection. After 12 months of treatment, they found that autonomic nerve functions were restored and the latent period of somatosensory evoked potentials was reduced.[79] Clinical studies with hUCB are still rare because of the concerns about safety and efficiency.

Conclusion

SCI is a life-changing event with a predilection for young patients who face a long life of physical and emotional struggles. The treatment of acute SCI is an ever-evolving landscape and one that is likely to see tremendous changes over coming years. SCI continues to be at the forefront of scientific research into stem cell and biotechnology innovation, with stem cell implantation, fast emerging as a promising therapeutic option, with the proven advantages of ease of availability, especially the hESCs, their easy administration, and fewer adverse effects. Various studies are underway, identifying the potential areas of damage in SCI, the histopathologic changes and the potential target areas for therapy in SCI models. The combination of basic science and clinical advances brings hope to the current and future sufferers of SCI. For better SCI recovery, more SCI research, systematic preclinical testing of promising therapies, diverse and abundant source of transplantable stem cells, genetically modified stem cells optimized for specific conditions, or the combination therapies are the areas that need to be further explored.

• References

• 1 Dumont RJ, Okonkwo DO, Verma S , et al. Acute spinal cord injury, part I: pathophysiologic mechanisms. Clin Neuropharmacol 2001; 24 (5) 254-264
• 2 Profyris C, Cheema SS, Zang D, Azari MF, Boyle K, Petratos S. Degenerative and regenerative mechanisms governing spinal cord injury. Neurobiol Dis 2004; 15 (3) 415-436
• 3 Tator CH, Fehlings MG. Review of the secondary injury theory of acute spinal cord trauma with emphasis on vascular mechanisms. J Neurosurg 1991; 75 (1) 15-26
• 4 McKinley W, Santos K, Meade M, Brooke K. Incidence and outcomes of spinal cord injury clinical syndromes. J Spinal Cord Med 2007; 30 (3) 215-224
• 5 Lee BB, Cripps RA, Fitzharris M, Wing PC. The global map for traumatic spinal cord injury epidemiology: update 2011, global incidence rate. Spinal Cord 2014; 52 (2) 110-116
• 6 “Spinal Cord Injury Facts.” Foundation for Spinal Cord Injury Prevention, Care & Cure; June 2009
• 7 Qin W, Bauman WA, Cardozo C. Bone and muscle loss after spinal cord injury: organ interactions. Ann N Y Acad Sci 2010; 1211: 66-84
• 8 Spinal Cord Injury in Men. http://menshealth.about.com/od/conditions/a/Spinal_Injury.htm . Retrieved June 15, 2015
• 9 Dedeepiya VD, Rao YY, Jayakrishnan GA , et al. Index of CD34+ cells and mononuclear cells in the bone marrow of spinal cord injury patients of different age groups: a comparative analysis. Bone Marrow Res 2012; 2012: 787414
• 10 Willyard C. Stem cells: a time to heal. Nature 2013; 503 (7475) S4-S6
• 11 Lu P, Kadoya K, Tuszynski MH. Axonal growth and connectivity from neural stem cell grafts in models of spinal cord injury. Curr Opin Neurobiol 2014; 27: 103-109
• 12 Knoller N, Auerbach G, Fulga V , et al. Clinical experience using incubated autologous macrophages as a treatment for complete spinal cord injury: phase I study results. J Neurosurg Spine 2005; 3 (3) 173-181
• 13 Hansebout RR, Tanner JA, Romero-Sierra C. Current status of spinal cord cooling in the treatment of acute spinal cord injury. Spine 1984; 9 (5) 508-511
• 14 Shapiro S, Borgens R, Pascuzzi R , et al. Oscillating field stimulation for complete spinal cord injury in humans: a phase 1 trial. J Neurosurg Spine 2005; 2 (1) 3-10
• 15 Silva NA, Sousa N, Reis RL, Salgado AJ. From basics to clinical: a comprehensive review on spinal cord injury. Prog Neurobiol 2014; 114: 25-57
• 16 Cadotte DW, Fehlings MG. Spinal cord injury: a systematic review of current treatment options. Clin Orthop Relat Res 2011; 469 (3) 732-741
• 17 Ruff CA, Wilcox JT, Fehlings MG. Cell-based transplantation strategies to promote plasticity following spinal cord injury. Exp Neurol 2012; 235 (1) 78-90
• 18 Beattie MS, Bresnahan JC, Komon J , et al. Endogenous repair after spinal cord contusion injuries in the rat. Exp Neurol 1997; 148 (2) 453-463
• 19 Xu XM, Guénard V, Kleitman N, Bunge MB. Axonal regeneration into Schwann cell-seeded guidance channels grafted into transected adult rat spinal cord. J Comp Neurol 1995; 351 (1) 145-160
• 20 Menei P, Montero-Menei C, Whittemore SR, Bunge RP, Bunge MB. Schwann cells genetically modified to secrete human BDNF promote enhanced axonal regrowth across transected adult rat spinal cord. Eur J Neurosci 1998; 10 (2) 607-621
• 21 Tuszynski MH, Weidner N, McCormack M, Miller I, Powell H, Conner J. Grafts of genetically modified Schwann cells to the spinal cord: survival, axon growth, and myelination. Cell Transplant 1998; 7 (2) 187-196
• 22 Hurtado A, Moon LD, Maquet V, Blits B, Jérôme R, Oudega M. Poly (D,L-lactic acid) macroporous guidance scaffolds seeded with Schwann cells genetically modified to secrete a bi-functional neurotrophin implanted in the completely transected adult rat thoracic spinal cord. Biomaterials 2006; 27 (3) 430-442
• 23 Chau CH, Shum DK, Li H , et al. Chondroitinase ABC enhances axonal regrowth through Schwann cell-seeded guidance channels after spinal cord injury. FASEB J 2004; 18 (1) 194-196
• 24 Pearse DD, Pereira FC, Marcillo AE , et al. cAMP and Schwann cells promote axonal growth and functional recovery after spinal cord injury. Nat Med 2004; 10 (6) 610-616
• 25 Elisseeff JH. Embryonic stem cells: potential for more impact. Trends Biotechnol 2004; 22 (4) 155-156
• 26 Brüstle O, Jones KN, Learish RD , et al. Embryonic stem cell-derived glial precursors: a source of myelinating transplants. Science 1999; 285 (5428) 754-756
• 27 Jendelová P, Herynek V, Urdzíková L , et al. Magnetic resonance tracking of transplanted bone marrow and embryonic stem cells labeled by iron oxide nanoparticles in rat brain and spinal cord. J Neurosci Res 2004; 76 (2) 232-243
• 28 Kitagawa A, Nakayama T, Takenaga M , et al. Lecithinized brain-derived neurotrophic factor promotes the differentiation of embryonic stem cells in vitro and in vivo. Biochem Biophys Res Commun 2005; 328 (4) 1051-1057
• 29 Howard MJ, Liu S, Schottler F, Joy Snider B, Jacquin MF. Transplantation of apoptosis-resistant embryonic stem cells into the injured rat spinal cord. Somatosens Mot Res 2005; 22 (1–2) 37-44
• 30 Chen J, Bernreuther C, Dihné M, Schachner M. Cell adhesion molecule l1-transfected embryonic stem cells with enhanced survival support regrowth of corticospinal tract axons in mice after spinal cord injury. J Neurotrauma 2005; 22 (8) 896-906
• 31 Nistor GI, Totoiu MO, Haque N, Carpenter MK, Keirstead HS. Human embryonic stem cells differentiate into oligodendrocytes in high purity and myelinate after spinal cord transplantation. Glia 2005; 49 (3) 385-396
• 32 Keirstead HS, Nistor G, Bernal G , et al. Human embryonic stem cell-derived oligodendrocyte progenitor cell transplants remyelinate and restore locomotion after spinal cord injury. J Neurosci 2005; 25 (19) 4694-4705
• 33 Faulkner J, Keirstead HS. Human embryonic stem cell-derived oligodendrocyte progenitors for the treatment of spinal cord injury. Transpl Immunol 2005; 15 (2) 131-142
• 34 Cloutier F, Siegenthaler MM, Nistor G, Keirstead HS. Transplantation of human embryonic stem cell-derived oligodendrocyte progenitors into rat spinal cord injuries does not cause harm. Regen Med 2006; 1 (4) 469-479
• 35 Lee H, Shamy GA, Elkabetz Y , et al. Directed differentiation and transplantation of human embryonic stem cell-derived motoneurons. Stem Cells 2007; 25 (8) 1931-1939
• 36 Erceg S, Ronaghi M, Stojković M. Human embryonic stem cell differentiation toward regional specific neural precursors. Stem Cells 2009; 27 (1) 78-87
• 37 Erceg S, Ronaghi M, Oria M , et al. Transplanted oligodendrocytes and motoneuron progenitors generated from human embryonic stem cells promote locomotor recovery after spinal cord transection. Stem Cells 2010; 28 (9) 1541-1549
• 38 Baizabal JM, Covarrubias L. The embryonic midbrain directs neuronal specification of embryonic stem cells at early stages of differentiation. Dev Biol 2009; 325 (1) 49-59
• 39 Keirstead HS. Stem cell transplantation into the central nervous system and the control of differentiation. J Neurosci Res 2001; 63 (3) 233-236
• 40 Wislet-Gendebien S, Laudet E, Neirinckx V, Rogister B. Adult bone marrow: which stem cells for cellular therapy protocols in neurodegenerative disorders?. J Biomed Biotechnol 2012; 2012: 601560
• 41 Moviglia GA, Fernandez Viña R, Brizuela JA , et al. Combined protocol of cell therapy for chronic spinal cord injury. Report on the electrical and functional recovery of two patients. Cytotherapy 2006; 8 (3) 202-209
• 42 Pal R, Venkataramana NK, Bansal A , et al. Ex vivo-expanded autologous bone marrow-derived mesenchymal stromal cells in human spinal cord injury/paraplegia: a pilot clinical study. Cytotherapy 2009; 11 (7) 897-911
• 43 Karamouzian S, Nematollahi-Mahani SN, Nakhaee N, Eskandary H. Clinical safety and primary efficacy of bone marrow mesenchymal cell transplantation in subacute spinal cord injured patients. Clin Neurol Neurosurg 2012; 114 (7) 935-939
• 44 Mazzini L, Mareschi K, Ferrero I , et al. Mesenchymal stromal cell transplantation in amyotrophic lateral sclerosis: a long-term safety study. Cytotherapy 2012; 14 (1) 56-60
• 45 Hofstetter CP, Schwarz EJ, Hess D , et al. Marrow stromal cells form guiding strands in the injured spinal cord and promote recovery. Proc Natl Acad Sci U S A 2002; 99 (4) 2199-2204
• 46 Bakshi A, Barshinger AL, Swanger SA , et al. Lumbar puncture delivery of bone marrow stromal cells in spinal cord contusion: a novel method for minimally invasive cell transplantation. J Neurotrauma 2006; 23 (1) 55-65
• 47 Shi E, Kazui T, Jiang X , et al. Therapeutic benefit of intrathecal injection of marrow stromal cells on ischemia-injured spinal cord. Ann Thorac Surg 2007; 83 (4) 1484-1490
• 48 Khalatbary AR, Tiraihi T. Localization of bone marrow stromal cells in injured spinal cord treated by intravenous route depends on the hemorrhagic lesions in traumatized spinal tissues. Neurol Res 2007; 29 (1) 21-26
• 49 Vaquero J, Zurita M, Oya S, Santos M. Cell therapy using bone marrow stromal cells in chronic paraplegic rats: systemic or local administration?. Neurosci Lett 2006; 398 (1–2) 129-134
• 50 Ankeny DP, McTigue DM, Jakeman LB. Bone marrow transplants provide tissue protection and directional guidance for axons after contusive spinal cord injury in rats. Exp Neurol 2004; 190 (1) 17-31
• 51 Yano S, Kuroda S, Shichinohe H , et al. Bone marrow stromal cell transplantation preserves gammaaminobutyric acid receptor function in the injured spinal cord. J Neurotrauma 2006; 23 (11) 1682-1692
• 52 Isele NB, Lee HS, Landshamer S , et al. Bone marrow stromal cells mediate protection through stimulation of PI3-K/Akt and MAPK signaling in neurons. Neurochem Int 2007; 50 (1) 243-250
• 53 Mackay-Sim A, St John JA. Olfactory ensheathing cells from the nose: clinical application in human spinal cord injuries. Exp Neurol 2011; 229 (1) 174-180
• 54 Féron F, Perry C, Cochrane J , et al. Autologous olfactory ensheathing cell transplantation in human spinal cord injury. Brain 2005; 128 (Pt 12): 2951-2960
• 55 Li Y, Field PM, Raisman G. Repair of adult rat corticospinal tract by transplants of olfactory ensheathing cells. Science 1997; 277 (5334) 2000-2002
• 56 Keyvan-Fouladi N, Raisman G, Li Y. Functional repair of the corticospinal tract by delayed transplantation of olfactory ensheathing cells in adult rats. J Neurosci 2003; 23 (28) 9428-9434
• 57 Ramón-Cueto A, Cordero MI, Santos-Benito FF, Avila J. Functional recovery of paraplegic rats and motor axon regeneration in their spinal cords by olfactory ensheathing glia. Neuron 2000; 25 (2) 425-435
• 58 Lu J, Féron F, Ho SM, Mackay-Sim A, Waite PM. Transplantation of nasal olfactory tissue promotes partial recovery in paraplegic adult rats. Brain Res 2001; 889 (1–2) 344-357
• 59 García-Alías G, López-Vales R, Forés J, Navarro X, Verdú E. Acute transplantation of olfactory ensheathing cells or Schwann cells promotes recovery after spinal cord injury in the rat. J Neurosci Res 2004; 75 (5) 632-641
• 60 Huang H, Chen L, Xi H , et al. Fetal olfactory ensheathing cells transplantation in amyotrophic lateral sclerosis patients: a controlled pilot study. Clin Transplant 2008; 22 (6) 710-718
• 61 Chen L, Chen D, Xi H , et al. Olfactory ensheathing cell neurorestorotherapy for amyotrophic lateral sclerosis patients: benefits from multiple transplantations. Cell Transplant 2012; 21 (Suppl. 01) S65-S77
• 62 Ruitenberg MJ, Plant GW, Hamers FP , et al. Ex vivo adenoviral vector-mediated neurotrophin gene transfer to olfactory ensheathing glia: effects on rubrospinal cord. J Neurosci 2003; 23: 7045-7058
• 63 Cao L, Liu L, Chen ZY , et al. Olfactory ensheathing cells genetically modified to secrete GDNF to promote spinal cord repair. Brain 2004; 127 (Pt 3): 535-549
• 64 Andressen C. Neural stem cells: from neurobiology to clinical applications. Curr Pharm Biotechnol 2013; 14 (1) 20-28
• 65 Cummings BJ, Uchida N, Tamaki SJ , et al. Human neural stem cells differentiate and promote locomotor recovery in spinal cord-injured mice. Proc Natl Acad Sci U S A 2005; 102 (39) 14069-14074
• 66 Teng YD, Lavik EB, Qu X , et al. Functional recovery following traumatic spinal cord injury mediated by a unique polymer scaffold seeded with neural stem cells. Proc Natl Acad Sci U S A 2002; 99 (5) 3024-3029
• 67 Lu P, Jones LL, Snyder EY, Tuszynski MH. Neural stem cells constitutively secrete neurotrophic factors and promote extensive host axonal growth after spinal cord injury. Exp Neurol 2003; 181 (2) 115-129
• 68 Ishii K, Nakamura M, Dai H , et al. Neutralization of ciliary neurotrophic factor reduces astrocyte production from transplanted neural stem cells and promotes regeneration of corticospinal tract fibers in spinal cord injury. J Neurosci Res 2006; 84 (8) 1669-1681
• 69 Ziv Y, Avidan H, Pluchino S, Martino G, Schwartz M. Synergy between immune cells and adult neural stem/progenitor cells promotes functional recovery from spinal cord injury. Proc Natl Acad Sci U S A 2006; 103 (35) 13174-13179
• 70 Pfeifer K, Vroemen M, Blesch A, Weidner N. Adult neural progenitor cells provide a permissive guiding substrate for corticospinal axon growth following spinal cord injury. Eur J Neurosci 2004; 20 (7) 1695-1704
• 71 Wu P, Tarasenko YI, Gu Y, Huang LY, Coggeshall RE, Yu Y. Region-specific generation of cholinergic neurons from fetal human neural stem cells grafted in adult rat. Nat Neurosci 2002; 5 (12) 1271-1278
• 72 Tarasenko YI, Gao J, Nie L , et al. Human fetal neural stem cells grafted into contusion-injured rat spinal cords improve behavior. J Neurosci Res 2007; 85 (1) 47-57
• 73 Engesser-Cesar C, Anderson AJ, Basso DM, Edgerton VR, Cotman CW. Voluntary wheel running improves recovery from a moderate spinal cord injury. J Neurotrauma 2005; 22 (1) 157-171
• 74 Grill R, Murai K, Blesch A, Gage FH, Tuszynski MH. Cellular delivery of neurotrophin-3 promotes corticospinal axonal growth and partial functional recovery after spinal cord injury. J Neurosci 1997; 17 (14) 5560-5572
• 75 Tuszynski MH, Grill R, Jones LL, McKay HM, Blesch A. Spontaneous and augmented growth of axons in the primate spinal cord: effects of local injury and nerve growth factor-secreting cell grafts. J Comp Neurol 2002; 449 (1) 88-101
• 76 Tobias CA, Shumsky JS, Shibata M , et al. Delayed grafting of BDNF and NT-3 producing fibroblasts into the injured spinal cord stimulates sprouting, partially rescues axotomized red nucleus neurons from loss and atrophy, and provides limited regeneration. Exp Neurol 2003; 184 (1) 97-113
• 77 Shumsky JS, Tobias CA, Tumolo M, Long WD, Giszter SF, Murray M. Delayed transplantation of fibroblasts genetically modified to secrete BDNF and NT-3 into a spinal cord injury site is associated with limited recovery of function. Exp Neurol 2003; 184 (1) 114-130
• 78 McTigue DM, Horner PJ, Stokes BT, Gage FH. Neurotrophin-3 and brain-derived neurotrophic factor induce oligodendrocyte proliferation and myelination of regenerating axons in the contused adult rat spinal cord. J Neurosci 1998; 18 (14) 5354-5365
• 79 Yao L, He C, Zhao Y , et al. Human umbilical cord blood stem cell transplantation for the treatment of chronic spinal cord injury: Electrophysiological changes and long-term efficacy. Neural Regen Res 2013; 8 (5) 397-403
• 80 Sobani ZA, Quadri SA, Enam SA. Stem cells for spinal cord regeneration: Current status. Surg Neurol Int 2010; 1: 93
• 81 Ning G, Tang L, Wu Q , et al. Human umbilical cord blood stem cells for spinal cord injury: early transplantation results in better local angiogenesis. Regen Med 2013; 8 (3) 271-281