Regenerative Therapies for Spinal Cord Injury

Abstract

Spinal cord injury (SCI) is a serious problem that primarily affects younger and middle-aged adults at its onset. To date, no effective regenerative treatment has been developed. Over the last decade, researchers have made significant advances in stem cell technology, biomaterials, nanotechnology, and immune engineering, which may be applied as regenerative therapies for the spinal cord. Although the results of clinical trials using specific cell-based therapies have proven safe, their efficacy has not yet been demonstrated. The pathophysiology of SCI is multifaceted, complex and yet to be fully understood. Thus, combinatorial therapies that simultaneously leverage multiple approaches will likely be required to achieve satisfactory outcomes. Although combinations of biomaterials with pharmacologic agents or cells have been explored, few studies have combined these modalities in a systematic way. For most strategies, clinical translation will be facilitated by the use of minimally invasive therapies, which are the focus of this review. In addition, this review discusses previously explored therapies designed to promote neuroregeneration and neuroprotection after SCI, while highlighting present challenges and future directions.

Impact Statement

To date there are no effective treatments that can regenerate the spinal cord after injury. Although there have been significant preclinical advances in bioengineering and regenerative medicine over the last decade, these have not translated into effective clinical therapies for spinal cord injury. This review focuses on minimally invasive therapies, providing extensive background as well as updates on recent technological developments and current clinical trials. This review is a comprehensive resource for researchers working towards regenerative therapies for spinal cord injury that will help guide future innovation.

Introduction

Spinal cord injury (SCI) results from direct trauma to the tissue and is associated with loss of motor, sensory, and autonomic functions caudal to the site of injury.¹ According to the World Health Organization (WHO),² between 250,000 and 500,000 people around the world sustain an SCI each year. In the United States alone, there are around 11,000–20,000 new cases each year.³ However, the annual incidence of SCI varies in different countries; for example, it is 9.2 patients per million in Denmark and 40.1 per million in the United States.⁴ Around the world, the incidence ranges from 3.6 to 195.4 patients per million.⁵

In the United States, there are around 250,000 patients who are at present living with SCI.³ Over a lifetime, direct costs of SCI can reach 1.1–4.6 million USD per patient.⁶ Most often SCI occurs in young and middle-aged adults⁷ and those with SCI are two to five times more likely to die prematurely.² SCI affects not only a patient's physical and psychological health but also one's family and the broader community and economy.

Due to the lack of plasticity and limited regenerative capacity of the central nervous system (CNS), recovery of neural function after SCI is rare. After injury, the spinal cord undergoes several changes on the cellular and molecular level that interfere with axonal regeneration. These complex events are poorly understood, hindering the development of treatments that can lead to complete recovery.⁸ Present SCI treatment is mostly conservative relying on stabilization of the patient, prevention of complications, and physical rehabilitation.

Extensive research into regenerative strategies for SCI has been performed over the last couple of decades. Improvements in our understanding of the pathophysiology of SCI coupled with advances in cell-based therapies, biomaterials, and biomolecules have enabled us to develop new therapeutic approaches for neuroregeneration. Promising results from in vivo experiments evaluating therapies based on biomaterials, cells, or biomolecules are being continuously reported.^9–13 Because of the complexity of SCI pathophysiology, the use of combinatorial therapies may be more effective and lead to better regeneration.^14,15 Although various procedures can be used to administer therapy after SCI (Fig. 1), the use of minimally invasive strategies, such as injection, is needed to reduce risk of complications, including introducing additional injury, and thus spare neuronal circuitry.¹⁶

FIG. 1.

A schematic illustration showing different methods that can be used for the treatment of SCI. SCI, spinal cord injury. Adapted from Führmann et al.¹⁸² Color images are available online.

This review focuses on minimally invasive SCI therapeutics in clinical trials,^1,3,6,17 as well as those in preclinical^9–13 and in vitro phases of development.^18–21 In addition, major challenges are highlighted. Future successful approaches for the treatment of SCI will likely include the integration of several recent advances in various fields as combinatorial therapies in minimally invasive formats. This review summarizes accumulating knowledge, examines evidence and development, and highlights potential paths forward.

Pathophysiology

SCI is characterized by sequential primary, secondary, and chronic phases. The primary injury to the spinal cord is the result of initial trauma. The primary mechanical insult may occur from compression, shearing, laceration, stretch, distraction, hemorrhage, or vasospasm. Bone or tissue fragments from the primary injury can exacerbate swelling of the spinal cord and add to tissue damage.

Secondary injury follows in a progressive way, as a result of ischemia, inflammation, and development of a cytotoxic microenvironment, leading to death of functional cells and damage to the tissue microenvironment. The chronic stage of SCI is characterized by formation of astroglial and fibrous scar tissue around cystic cavitations. As a result, regeneration is thought to be inhibited, at least partially, by the extracellular matrix (ECM) and soluble factors secreted by inflammatory cells within the scar tissues (Fig. 2).⁶

FIG. 2.

An illustration showing the development of pathophysiological changes following SCI. The acute phase takes 0–48 h and it involves hemorrhage, edema, and proapoptotic factors (A). This leads to further loss of function, more than that resulting from the initial insult occurs due to injury to neurons and oligodendrocytes. Astrocyte infiltration and release of additional proinflammatory factors are seen while demyelinated and injured axons begin to die back. In the late subacute (B) and intermediate (C) stages, microcystic cavities follow cell death. These cavities then coalesce forming barriers to regeneration in the chronic stage (>6 months). The final chronic stage scar, which is composed of a network of astrocytic processes and a dense fibrous deposit, acts as a physical and biochemical barrier to neurite outgrowth and cell migration. (D) A schematic illustration showing demyelination and axonal loss that follow SCI and various regenerative therapeutics that can be used including the use of biomaterials, cells, molecules, such as an anti-NOGO-A antibody treatment and Rho-ROCK inhibition, or agents to mobilize endogenous cells such as metformin. ROCK, Rho-associated protein kinase. Adapted from Ahuja et al.⁶ Color images are available online.

Any therapeutic intervention should consider and address the dynamic pathophysiological events occurring in SCI. For example, combating measures against increased extracellular amino acids should be applied early on, in minutes, while measures against inflammatory mediators in hours and measures against myelin-associated inhibitors within weeks after injury.²² Because inflammation continues and may persist for years, the administration of therapeutic measures should be considered to continue accordingly.²²

Given practical limitations of treating acute SCI in a clinical setting, many therapeutic interventions in development aim to target either the secondary or chronic stages of injury. For example, accumulation of myelin debris during secondary injury is associated with inhibition of regeneration. Myelin-associated proteins lead to inhibition of neurite outgrowth, thus named NOGO inhibitors, which act through NOGO receptors leading to activation of GTPase Rho A. Its effector, Rho-associated protein kinase (ROCK), leads ultimately to apoptosis, axonal collapse, and neurite retraction.⁶ While a detailed discussion of the mechanisms of myelin-associated inhibition of axonal outgrowth is outside the scope of this review, we refer the reader to a review by Zhang et al. for more information.²³

There is also an accumulation of extracellular amino acids at the site of injury, whose abundance relates to severity of SCI and neurotoxicity.²² These molecules are attractive targets for pharmacological intervention aimed at enhancing neuroprotection and neuroregeneration post-SCI and are discussed more in the following sections. During secondary injury, damage occurs on the cellular level through several mechanisms, including mitochondrial dysfunction leading to high calcium levels, increased production of oxygen free radicals, and oxidative injury.²⁴

In chronic injury, scar formation is a problem and its disruption is a common strategy for promoting regeneration. For example, a recent study found that the protein tyrosine phosphatase σ (PTPσ) contributes to a transition of regenerating growth cones into dystrophic bulbs through interactions with chondroitin sulfate proteoglycans (CSPGs), which are abundant in the glial scar and many forms of which have been widely reported to inhibit axonal outgrowth.²⁵ Experimental inhibition of PTPσ was found to promote regeneration in a rodent model of SCI by enabling axons to overcome CSPG-mediated growth inhibition and, consequently, serotonergic fiber navigation across the SCI lesion leading to some functional recovery.²⁵

While the CSPG-rich glial scar was previously thought to be completely inhibitory to axonal growth, more recent studies have found that some aspects of scar ECM, which is secreted by astrocytes, are permissive and may even promote regeneration.²⁶ Furthermore, prevention, attenuation, or ablation of chronic astrocytic scars increased lesion size and reduced axonal ingrowth.²⁷ Recent studies found also that the ECM secreted by protoplasmic, but not fibrous, astrocytes can promote axonal elongation when injected into the rats' spinal cord after SCI.²⁶ Taken together, these findings suggest that while some aspects of the astrocytic scar may be detrimental, other components may be required for regeneration.

Therapeutic Approaches

Standard treatments

The management of SCI represents a formidable challenge, and there is no treatment at present available that can effectively restore lost tissue function. Several factors are responsible for the failure of healing after SCI, including chronic local inflammation and the release of antiregenerative factors. Present treatment methods include the use of neuroprotective and neuroregenerative strategies. The first line of management for SCI patients includes resuscitation, stabilization, and critical care in specialized centers with early determination and localization of specific injuries. Stabilization may include removal of bone fragments or foreign objects, physical stabilization of the vertebral column, and spinal decompression.

Nonpharmacological methods to reduce inflammation, including hypothermia²⁸ and cerebrospinal fluid drainage,²⁹ have also been in phase I/II clinical trials with promising early results. Early surgical decompression (within 24 h), administration of anti-inflammatory drugs, and augmentation of blood pressure lead to reduced acute complications and hospital stay.⁶ Timely and early intervention is important and can be achieved by using neuroprotective measures (ideally in the acute stage).

The management of SCI also includes prevention and treatment of complications, and physical rehabilitation. Without these, there is very limited recovery of neurological function.¹ Besides failure of regeneration, SCI is often complicated by other problems such as infection and pressure sores, which can be serious and ultimately lead to death, such as in the case of the actor Christopher Reeve following his SCI.³⁰ Development of regenerative therapies to treat neurological deficit represents an active area of research and is a rapidly growing field.⁶ Improved clinical outcomes will emerge by harnessing the potential of recent advances in different fields and integrating them to create innovative therapeutic strategies.

Neuroregenerative and neuroprotective therapies

Despite active research and clinical studies, there is no effective and globally accepted standard treatment for SCI.⁸ Various minimally invasive, neuroprotective, and neuroregenerative strategies using cells, biomaterials, or pharmacological agents are in various stages of preclinical research and clinical trials around the world (Table 1 and Fig. 3). Clinical trials have evaluated agents that can neutralize axonal growth inhibitors, such as the Rho-ROCK inhibitor, Cethrin/VX-210, and anti-NOGO monoclonal antibody.^6,31 However, no pharmacological agent for neuroregeneration (restoration of neurological function) has been approved by the U.S. Food and Drug Administration (FDA) for treatment of SCI.³²

FIG. 3.

An illustration showing the global trends of clinical and preclinical studies in which cell transplantation is involved. Number of studies, type of cells (ESCs, BM-MSCs, A-MSCs, AF-MSCs, UCB-MSCs, or OECs), and routes of administration (IV, intrathecal, fourth ventricle, or intracisterna) are shown. When no data available, it was indicated as NA. AF-MSCs, amniotic fetal MSCs; A-MSCs, adipose tissue-derived mesenchymal stem cells; BM-MSCs, bone marrow-derived mesenchymal stem cells; ESCs, embryonic stem cells; IV, intravenous; OECs, olfactory ensheathing cells; UCB-MSCs, umbilical cord blood MSCs. Reproduced with permission from Vismara et al.⁷⁰ Color images are available online.

Table 1.

Biomaterials Studied for Treatment of Spinal Cord Injury

Materials used	Cells or factors included	Injury model and delivery method	Notable findings	References
Phase-separated poly (2-hydroxyethyl methacrylate) (pHEMA)	N/A	Spinal lesion, injection	Reduced accumulation of GFAP and Neurocan; however, scaffold did not integrate into tissue	¹³⁴
Alginate	EGF, bFGF	Clip-compression, intrathecal injection	Improved functional recovery; enhanced outgrowth of corticospinal tracts, and angiogenesis	¹³⁵
HPMA copolymers	Bivalirudin peptides	Clip-compression, intrathecal injection	Decreased cellular proliferation, inflammation, and astrogliosis	¹⁴⁴
Hyaluronic acid	IKVAV, BDNF	Clip-compression, intrathecal injection	BDNF-hydrogels showed greatest improvement in locomotor functional recovery	^152,155
Agarose/carbomer hydrogels	RGD, ECM deposition	Clip-compression, intrathecal injection	Increased MP M2 population	⁴³
Agarose/carbomer hydrogels	Human mesenchymal cells	Clip-compression, intrathecal injection	Increased MP M2 population	⁴³
BD PuraMatrix synthetic peptide	Human Schwann cells	Clip-compression, intrathecal injection	Reduced astrogliosis and promoted infiltration of endogenous S100-positive cells	⁵⁶
Collagen	Laminin	Clip-compression, intrathecal injection	Enhanced long-term cell survival, vascularization, axonal regrowth, and infiltration	¹¹
Collagen	Schwann cells	Clip-compression, intrathecal injection		¹¹

ECM, extracellular matrix.

Cell-based therapies may provide multiple benefits, including neuroprotection, angiogenesis, immune modulation, and tissue regeneration. Cell types evaluated have included stem cells (embryonic [ESCs], neural [NSPCs], or mesenchymal [MSCs]) and glial cells (Schwann cells, olfactory ensheathing cells [OECs], or oligodendrocyte precursor cells [OPCs]).^1,6 In addition, electrical field stimulation of the injured spinal cord has been used to enhance neuronal plasticity and direct rewiring of circuitry into functional relays.³³ As in wound healing of other tissues, angiogenesis within the injured spinal cord tissue has been correlated with positive outcomes, particularly in studies delivering biomaterial, cell, or angiogenic soluble factors.^13,34,35

Biomaterials have also been investigated for their inherent regenerative potential as well as for the delivery of therapeutic agents, including cells,^9,16,36–44 drugs,^36,45 growth factors,^16,37,38,46 or genes (Fig. 4).⁴⁷ Combinations of these are also being investigated.^{16,36–38,48–51} Nevertheless, only one biomaterial has reached clinical trials,⁵² while most remain in preclinical animal^43,53–56 or in vitro stages of investigation.^57,58 Even developed therapies with potential for clinical translation still require extensive testing to prove their safety and efficacy in patients. Further complicating results is the fact that there are no comprehensive studies to compare different strategies under identical conditions. Research groups can vary greatly in the choice of animal or injury model and are understandably restricted in scope. To date, none of these neuroregenerative therapies is used routinely in clinical practice.

FIG. 4.

Illustration showing that scaffolds (made of synthetic or natural biomaterials) can be used for the treatment of SCI, as micropatterned devices or as matrices combined with drug delivery, or gene delivery to enable guided tissue regeneration. Adapted from Iyer et al.²⁰¹ Color images are available online.

Cell-based therapies

Cell-based therapy may be suitable for the treatment of discrete lesions in SCI (Figs. 3 and 5) to provide the following: (1) trophic support,⁵⁹ (2) immunomodulation,^60,61 (3) angiogenesis,³⁵ (4) and generation of new CNS cells,^1,6 which may form functional relays (Fig. 6).^15,16,48 In addition, paracrine factors released by administered cells may aid in the activation of resident progenitors.⁶² While stem cell therapies have shown promising results in clinical trials,^63,64 translation faces substantial challenges, including timing and method of administration,¹ and risks that have to be carefully evaluated, including (1) poor survival of transplanted cells,^46,63 (2) migration away from the delivery site,^65,66 (3) formation of ectopic stem cell colonies or tumors,⁶⁷ (4) excessive proliferation,³⁷ (4) differentiation into unwanted, nonregenerative cell types,⁶⁸ and (5) aberrant axonal growth leading to allodynia.⁶⁹ Controlling stem cell differentiation or production of specific regenerative factors remains as additional key objectives.^70,71

FIG. 5.

Schematic illustration showing the main cellular targets of cell therapy in SCI and how this works (A), with particular focus on the mechanism by which stem cells act to achieve immunomodulatory action, differentiation into neurons and oligodendrocytes, or providing trophic factors that can support axonal regeneration (B). Reproduced with permission from Vismara et al.⁷⁰ Color images are available online.

FIG. 6.

Schematic illustration showing bridge (A) or relay (B) formation during the repair process after the transplantation of cells for the treatment of SCI leading to neuronal connectivity. Reproduced with permission from Assinck et al.¹⁷ Color images are available online.

While various cell therapies are being evaluated in clinical trials, none of these trials has progressed past phase I/IIa.³ The absence of controls in these trials has limited their value, and the efficacy and long-term outcomes after treatment of SCI are yet to be demonstrated.¹⁷ Financial costs of cell-based therapies are another major hurdle. Of the more successful clinical trials, one was terminated due to limited long-term effects,⁷² while another was terminated for financial reasons.⁷³ Questions relating to the type of cells used, number, and safety remain to be addressed.⁷⁴ Stem cell therapy remains experimental and it cannot ethically be offered to patients as a proven form of treatment.

Despite these limitations, clinical and preclinical studies evaluating effects of transplanting several different cell types, including NSPCs, MSCs, Schwann cells, OPCs, and OECs, have shown promising results to date.^1,6,17 NSPCs have been derived from human fetal spinal cord,^63,75 human fetal brain,^44,72,76 or pluripotent or ESCs⁶⁴ and can differentiate into the glial and neuronal cells, which normally make up the spinal cord. MSCs, including those derived from adipose tissue (A-MSCs),⁷⁷ umbilical cord blood,⁷⁸ and Wharton's jelly,⁷⁹ human OPCs derived from human ESC (hESC) lines,⁶⁴ and human NSPCs derived from fetal spinal cord tissues⁶³ have been investigated in clinical trials.

While not in clinical trials, there has also been recent interest in using human dental pulp cells (hDPCs) because they contain neural crest-derived ecto-MSCs. These cells were found to be more effective than bone marrow-derived MSCs in inducing functional recovery in rodents following SCI.⁸⁰ Another promising cell source yet to be used in clinical trials is the human inducible pluripotent stem cell (iPSC), which may help overcome issues with immune rejection of allogenic transplants.

These types of cells vary greatly in their tissue source and processing methods, often making it difficult to directly compare results obtained from different studies. For example, there is strong evidence that NSPCs or OPCs derived from hESCs using different protocols or obtained from varying sources may have different regenerative capacities.⁴⁶ For example, some preclinical studies reported robust results with the use of NSPCs, and it was found that axonal regeneration can be obtained even with the use of NSPCs obtained from a different species, for example, with the use of human NSCs in immunodeficient rats.¹⁶ In contrast, other reports indicated that CNS-derived stem cells were not efficient in the treatment of cervical spine contusion injury in mice and may even have negative effects.⁴⁶

Recently, questions regarding the translation of preclinical data describing cell therapies to the clinic have been raised.^46,81,82 It was found that there are differences in therapeutic effects between cell lots, such as that observed with the cell lots used for research and those of clinical grade of an NSPC product (human central nervous system stem cell [HuCNS-SC]) cell in preclinical studies by Anderson et al.⁴⁶ Although the use of a research-grade cell lot was successful in rat and mouse thoracic SCI models, transplantation of the corresponding clinical-grade cell lot into a cervical SCI mouse model provided no benefits. Moreover, some data suggested negative effects.⁴⁶

One possible reason for the difference is the heterogenicity of the fetal tissue from which these cells were derived. Donor genetic makeup, the location from which cells were taken, and the developmental stage at which the cells were isolated may all vary. In addition, protocols and reagents used to maintain and differentiate therapeutic cells may all lead to differences among study outcomes.⁸¹ These difficulties may have contributed to the termination of clinical trials using HuCNS-SC transplantation for the treatment of SCI, which were terminated based on business decisions unrelated to any safety concerns.^72,83 Differences between research-grade and clinical-grade cells will undoubtedly be a critical factor in the translational capacity of stem cell therapies.

Timing of the administration of cell-based therapy after SCI is very important. Early delivery after an acute injury may not be appropriate because the hyperinflammatory environment may lead to death of transplanted cells. On the contrary, delayed administration may result in no neuroregenerative effects because of the loss of cell plasticity at the site of injury and extensive scar and cyst formation.⁷⁴ Studies have suggested that cell transplantation during the subacute stage is more effective than that during acute or chronic stages.¹ Hence, in most previous studies, cells were administered during the subacute stage.⁸⁴ It was also found from a clinical study that cell application within the therapeutic window of 3–4 weeks post-SCI will have an important role in clinical settings to balance the feasibility of transplantation with optimal therapeutic efficacy.⁸⁵

Soluble bioactive molecule-based therapies

Soluble bioactive molecules, such as growth factors and drugs, have been explored as SCI therapies for their neuroprotective, neuroregenerative, angiogenic, and immunomodulatory effects. As discussed in detail later in this review, biomolecules are often delivered in combination with biomaterials.^16,86,87 The use of carriers⁸⁸ such as hydrogels, and nanoparticles (NPs) can improve therapeutic drug delivery and efficacy.⁸⁹ Promising neuroprotective therapies for use in the acute stage of SCI that are being investigated for translation to the clinic include the use of various pharmacological agents such as riluzole,⁹⁰ magnesium,⁹¹ minocycline,⁹² hepatocyte growth factor (HGF),⁹³ and granulocyte-colony stimulating factor (G-CSF).^6,94

In preclinical studies, improved axonal sprouting has been demonstrated in experimental SCI in rats to which antibodies against NOGO were delivered by cells placed into the parietal cortex.⁹⁵ Inhibition of phosphatase and tensin homologue (PTEN), a downstream target of NOGO-RhoA-ROCK in the sensorimotor cortex, has also been shown to promote corticospinal tract regeneration in rodents.^96,97 Another recent study reported that the administration of connective tissue growth factor (CTGF) in zebrafish, after SCI leads to healing of SCI in these fish within weeks.⁹⁸ Recently, pregabalin, an established drug treatment for several neurological disorders, was also found to block the axonal regeneration inhibitor in mice,⁹⁹ and it is worth exploring its use further in SCI studies.

Biomolecules modulating the immune response are also candidates for SCI treatment. For example, blocking of the interleukin-7 (IL-7) receptor favors the generation of M2 phenotype macrophages and thus improves functional recovery after experimental SCI in mice.¹⁰⁰ Delivery of IL-4 and IL-10 chemokines to promote macrophages to adopt an M2-like phenotype has also been shown to improve recovery after SCI in a mouse model.^101,102 The use of fractalkine, a chemokine that preferentially recruits reparative monocytes, was found to dramatically increase the regeneration of experimental nerve defects in rats¹⁰³ and thus may have benefits if used as a treatment for SCI.

Factors released by stem cells may also be useful in the treatment of SCI.⁷⁰ Intrathecal injection of conditioned medium of MSCs into experimental SCI in rats was found to increase axonal regeneration and improve locomotor recovery 1–4 weeks after the treatment.⁵⁹ Stem cell-conditioned media contain several potentially regenerative factors, including insulin-like growth factor 1 (IGF-1), vascular endothelial growth factor (VEGF), transforming growth factor β1 (TGF-β1), and HGF. It has been reported that the use of such media are more effective than using individual factors or their combinations in enhancing the survival of neurons and growth of neurites.¹⁰⁴

Factors secreted by stem cells can also have an important role in regulating immune cell phenotypes, that is, immunomodulatory action. Such secreted factors may influence neutrophils, macrophages, dendritic cells, NK cells, T cells, and B cells.⁷¹ Furthermore, stem cell-derived exosomes can be used to modulate the recruitment and function of macrophages.¹⁰⁵ It is important to have a deep understanding of stem cell-derived exosomes, and their effect, to enable the development of novel therapeutic methods.⁷¹ The use of these media or exosomes provides many future possibilities for their application in minimally invasive therapy of SCI.

Therapeutic biomolecules can also be used to stimulate resident NSPCs, various types of which are present throughout the spinal cord.¹⁰⁶ It has been suggested that ependymal progenitor cells, present in the central canal of the spinal cord, may enable regeneration in some animals even after severe injury.¹⁰⁷ Furthermore, ependymal progenitor cells in the glial scar can be beneficial. They can limit secondary injury by maintaining tissue integrity and releasing bioactive factors that promote neuronal survival.^108,109 Thus, manipulation of the local ependymal stem cell niche¹⁰⁷ or mobilization of other resident progenitors¹⁰⁶ to stimulate self-repair through these cells may be a useful approach.¹¹⁰ For example, delivery of factors known to promote oligodendrocyte-lineage differentiation during development in a mouse model of acute SCI, including sonic hedgehog, platelet-derived growth factor-AA (PDGF-AA), and noggin, has been reported to increase the numbers of myelinated, regenerating axons and functional outcomes.^111,112

It is worth highlighting some recent outcomes of clinical trials investigating the use of antiregeneration inhibitors (Rho-ROCK inhibitors¹¹³ and G-CSF¹¹⁴) for the treatment of SCI. Rho-ROCK inhibitors are thought to have neuroprotective and neuroregenerative effects.¹¹⁵ In a phase I/IIa clinical trial that involved 48 patients who had acute and complete cervical (C4-T1) or thoracic (T2-T12) SCI, the application of Rho inhibitor to the epidural space in a fibrin glue resulted in no increased adverse effects. Improvement in long-term motor function was best seen in cervical SCI patients who had the drug administered during 7.8–146 h after injury.¹¹⁵

Despite these promising results, a phase IIb trial for treatment of acute SCI with Cethrin/VX-120 was halted in late 2018 due to lack of efficacy.¹¹⁶ G-CSF is also being investigated for the treatment of SCI.¹¹⁴ Two phase I/IIa trials^94,117 and a small-scale double-blind, randomized clinical trial¹¹⁸ have reported improved outcomes. However, future clinical trials that are larger in scale and randomized will be required to demonstrate the efficacy of the treatment with G-CSF.

While the majority of previous clinical studies have delivered potentially therapeutic agents either intravenously or orally, intrathecal delivery may provide better access to the injury and better approximates the design of many preclinical experimental studies with positive results. Thus, despite its more invasive nature, it has been argued that this route of administration will have better clinical success.¹¹⁹ A recent phase I clinical trial reported the safety and tolerability of intrathecal infusion of the human anti-NOGO antibody ATI355 in patients after acute SCI.³¹ In addition, a phase I/II clinical trial in which the HGF was intrathecally injected at the lumbar level in patients with cervical SCI was completed in 2018 in Japan, and results are expected in the near future.¹¹⁹

Biomaterial-based therapies

So far, no biomaterial-based regenerative therapy of SCI is in routine clinical practice. However, a biomaterial technology, the Neuro-Spinal Scaffold, has recently reached clinical trials.^52,120,121 Biomaterials used for the treatment of SCI can be in the form of conduits,^101,112,122 sheets,¹²³ scaffolds,^121,124 fibers,¹²⁵ particles, or hydrogels (Table 1).^126–130 Biomaterials are used to mechanically stabilize the injury site and provide an environment for interactions with host cells, physically fill SCI-associated cavities, reconstituting ECM, and bridging the injury to guide axonal growth across the gap.^{70,101,112,121,131–135} To guide axonal regeneration, several biomaterial architectures have been investigated, including channels,^86,136–138 fibers,¹³⁹ scaffolds, and magnetic microgels.⁵⁸

Biomaterials for the treatment of SCI can also be used for the delivery of therapeutic agents and cells (Fig. 4).^6,132 In addition, they can support cell survival of delivered cells in situ,^44,50 as discussed later in this review, and recruit migrating endogenous cells, such as Schwann cells¹²¹ and stem cells,^111,112,140 that can support regeneration. To avoid additional complexities associated with the use of cells, the use of acellular biomaterials for supporting spinal cord regeneration and remodeling after injury has been important to investigate.^{121,136,141,142}

It is preferable that biomaterials be biodegradable and injectable for delivery through minimally invasive means.⁷⁰ In the acute phase of SCI, the use of a hydrogel is preferred as it may be used to seal the dura and release anti-inflammatory drugs. At later stages the use of fibers and conduits may be better for achieving guided neural regeneration.¹³² However, it would be ideal to design a system that could be implanted acutely and remain for months to both guide regeneration and prevent chronic inflammation.

To achieve appropriate tissue repair, the implanted biomaterial should degrade as the host tissue regenerates. While biomaterials such as poly(lactide-co-glycolide) (PLGA) degrade hydrolytically,^142,143 naturally derived materials such as fibrin are degraded by cell-produced enzymes^15,48 and thus may have better potential for coupling degradation to native matrix formation, which is a challenging objective. Hybrids of synthetic and natural biomaterials can combine desirable properties of both types of materials. For example, synthetic biomaterials with controlled properties and tailored degradation profiles can be functionalized with peptides, which are susceptible to degradation by cell-produced enzymes,^138,144 or provide sites for cell adhesion.^121,125

Several different types of scaffolds have been investigated for the treatment of SCI. A scaffold made of a block copolymer of PLGA and poly-l-lysine (PLL) with a highly interconnected porous structure (approximately 250–500-μm-diameter pores) was shown to lead to improved functional recovery after SCI in rats.¹⁴⁵ Later, the safety and efficacy of these scaffolds were evaluated in partial and complete hemisection models of thoracic SCI in Old-World primates (Chlorocebus sabaeus) and significant recovery of locomotion was observed.¹⁴¹ A significant increase in remodeled tissue with neural sprouting was seen 12 weeks after injury.

Results from preclinical studies, where these PLGA-PLL devices known as “Neuro-Spinal Scaffold” were implanted into a thoracic-level contusion in rats and minipigs after internal decompression was performed, reported reduced inflammation and lesion cavitation volumes and increased tissue sparing and deposition of new, endogenous laminin-rich tissue.¹²¹ However, no functional benefits were observed. The Neuro-Spinal Scaffold has recently been evaluated in small-scale clinical trials for treatment of thoracic-level SCI and has shown promising results.^52,120

While porous polymeric scaffolds, such as those based on PLGA, have shown promise, hydrogel materials can be fabricated to better match hydration and mechanical properties of native spinal cord tissue, both of which are thought to reduce inflammation and scarring. In addition, hydrogels can be readily formulated to be injected and formed in situ in the spinal cord. In addition, microgels can be used for the delivery of cells,^146,147 drugs, and/or biomolecules,^70,89,148 and they can be interesting formulations to use in regenerative therapy of the spinal cord. For detailed reviews of hydrogels as minimally invasive therapies for spinal cord repair, the authors refer the readers to Perale et al.,¹²⁷ Macaya and Spector,¹²⁶ Pakulska et al.¹⁴⁸ and Khaing et al.¹⁴⁹

The use of polyethylene glycol (PEG), an FDA-approved biomaterial used for many applications, is considered promising for the treatment of SCI because it can support stem cell growth, migration, proliferation, and differentiation. PEG hydrogels can also help to reduce local glial scar invasion, and promote and guide axonal regeneration.^138,150 While PEG is essentially biologically inert, hydrogels containing bioactive polymers provide the opportunity to more directly affect cells in the spinal cord, and thus, the repair process. In particular, polymers derived from the native ECM, including hyaluronic acid (HA),^{26,44,50,151–155} fibrin,^37,42,48 and laminin,^125,156,157 have shown success in preclinical studies.

In contrast to including individual ECM components, cell-secreted ECM compositions have also been shown to aide in regeneration.^43,125,158 Alternatively, spinal cord tissue, decellularized to preserve not only the ECM composition but also the structure, has been found to enhance axonal regeneration in the spinal cord of rats.¹⁵⁹

A number of studies have found that provision of a physical structure to guide regenerating axons along the longitudinal axis to their targets in the form of conduits or channels enhances functional recovery after SCI. For example, collagen-based neural conduits were functionalized with the neurotrophin-3 (NT-3) gene and used for the treatment of completely transected spinal cord in rats.⁴⁷ Increased NT-3 levels seen in surrounding tissues and axonal regeneration, observed 1 month postoperatively, were significantly higher than in controls.

PLGA scaffolds with multiple guidance channels implanted in mice were found to be sufficient for promoting corticospinal tract axonal regeneration into the channels, through the injury site, and out of the conduits, where they have the opportunity to connect with synaptic targets caudal to the lesion.¹⁶⁰ In this study, the extent of axonal regeneration was found to correlate with functional recovery. Hydrogels made from multiple base materials and containing guidance channels have been reported to promote regeneration in animal models of SCI.^{9,138,161,162} A recent report by Koffler et al. described the use of a microscale continuous projection printing method to create hydrogel scaffolds with guidance architectures that conform to contusion lesions in unique animals.¹⁶¹

While conduits and channels provide macroscale guidance structures, nanofibers that mimic the nanostructure of the native ECM have been developed for regenerative biomaterials and studied in vitro and in vivo.^163,164 Nanofibers have been shown to guide the orientation of regenerating neurites parallel to their orientation.¹⁶⁵ In addition, the use of electrospun nanofibers has been associated with ESC differentiation toward neural lineages.¹⁶⁶ Recently, it was found that peptide nanofibers containing long laminin motif can influence neurogenesis both in vitro and in vivo.¹⁵⁷ Synthetic poly(ɛ-caprolactone) (PCL) nanofibers were also investigated and found to successfully create electrically active human three-dimensional (3D) neuronal networks with synapses using brain neural progenitors.¹⁶⁷

Engineered NPs have been explored for their use for the regeneration of the spinal cord. For example, PLGA NPs were used for local delivery of flavopiridol (a cyclin-dependent kinase inhibitor that inhibits astrocyte growth and inflammatory agent synthesis), and they resulted in recovery following SCI in rats.¹⁶⁸ Using a biological ligand, gold NPs were bonded to neurons for optical excitation and they exhibited potential benefits over optogenetics.¹⁶⁹ Pulsed infrared light and plasmonic gold nanorods were also used to enhance infrared neural stimulation by increasing neural responsivity.¹⁷⁰

Intravenously injected NPs made of ferulic acid-modified glycol chitosan were shown to be effective in achieving functional restoration after SCI in rats.¹⁷¹ Moreover, magnetic NPs coupled to HA-based hydrogels can be used to modulate the activation of mechanosensitive ion channels in dorsal root ganglia neurons as it was demonstrated in vitro.²⁰ Nanofibers can be combined with therapeutic agents and NPs¹⁶⁴ to enhance their function. For example, gold NPs were incorporated into electrospun PCL/gelatin nanofiber-based scaffolds. Neuronal cells were subsequently seeded onto these scaffolds and resulted in axonal elongation forming 3D networks in vitro.¹⁸ In another example, zero valent zinc NPs included in a PCL matrix using electrospinning were found to promote neuroglial cell proliferation in vitro.¹⁹

In addition, electroconductive (e.g., graphene) or magnetic (e.g., iron oxide NPs) elements can be incorporated into scaffolds. The use of electroconductive materials and applied electromagnetic fields may help to influence cell migration, adhesion, proliferation, and differentiation.¹⁷² For example, magnetic NPs can be used to guide neurite growth¹⁷³ and enable the control of cell organization and differentiation (Fig. 7).¹⁷⁴ It was found that when polypyrrole-coated poly (l-lactide-co-ɛ-caprolactone) nanofiber nerve conduits were combined with electric stimulation and used in the treatment of experimental sciatic nerve defects in rats, results were similar to those obtained with the use of autografts.¹⁷⁵ Cell elongation was found to occur in a direction parallel to that of the nanofibers.¹⁷⁶

FIG. 7.

The use of magnetic rods in fibrin hydrogel to help guide regeneration that can be aligned upon exposure to magnetic field. (A). An illustration showing the method of preparation of injectable hybrid hydrogel. A unidirectional structure made of aligned rod-shaped, magnetoceptive microgels within the fibrin hydrogel is generated in situ by exposure to magnetic field. Following this, the surrounding liquid prehydrogel is crosslinked, so that microgel orientation is fixed to guide aligned cell ingrowth. Adapted from Rose et al.⁵⁸ (B) Premixed fibroblasts can be seen to extend along the microgel axis (green), as visualized by stretched F-actin filaments (stained in red). Reproduced with permission Rose et al.⁵⁸ Color images are available online.

Combined with electrical stimulation, conductive carbon nanofiber-based scaffolds were found to enhance the regenerative function of NSPCs.¹⁷⁷ Moreover, core-shell-structured, piezoelectric polyvinylidene fluoride (PVDF) nanofibers that were chemically wrapped by graphene oxide lamellae showed a desirable out-of-plane piezoelectric constant higher than that of the conventional pure PVDF nanofibers.¹⁷⁸ To produce graphene-containing silk, a recent method reported feeding of graphene to silk-producing worms to create silk with new properties,¹⁷⁹ adding to the available fabrication methods.

The concept of biomaterials that can be injected and form a defined guidance architecture in situ is attractive; nevertheless, development of such biomaterials has posed a formidable challenge. Promising strategies to overcome this challenge include the use of scaffolds that are based on self-assembling fibers^{56,95,124,180} or that are embedded with magnetically responsive microgels.⁵⁸ In an interesting study, poly(ethylene oxide-stat-propylene oxide) gels containing supermagnetic iron oxide nanorods were used to help the orientation of nerve extension parallel to the rods that were aligned by using an external magnetic field.^58,181 This offers a great opportunity to develop minimally invasive and regenerative therapies for SCI.

Combinatorial therapeutic approaches

Because of the complex nature of pathophysiological changes that occur following SCI, combinatorial approaches that address different aspects of the problem are expected to be more effective. In such an approach, biomaterials may be combined with cells and soluble molecules and administered locally to an injury. It is evident that biomaterials can provide sustained drug delivery, enhance survival, growth, host tissue integration, and even differentiation of delivered cells (Fig. 4).¹⁸² Attention should be paid to proper biomaterial selection, tailored drug release, source and processing of therapeutic cells, and timing of intervention. Despite recent advances in these therapeutics, there is still a likely long way to go to achieve a therapy that can lead to significant recovery after SCI.¹⁸³

Biomaterials and soluble biomolecules

In one approach for the treatment of SCI, biomaterials and bioactive molecules can be combined (Table 1). There are many examples of procedures using soluble molecule delivery, for example, growth factors,^{86,87,184,185} drugs,¹¹⁵ or other therapeutic agents,^54,186–189 for the treatment of SCI. Combining biomaterials with diffusible factors, and even gene therapies, can maximize therapeutic effects by targeting more than one aspect involved in SCI.

Biomaterials can also maintain appropriate local therapeutic concentrations at or near the injury site over longer periods of time than bolus injection and hence better support regeneration.⁷⁰ For example, hydrogels fabricated from a hydrolytically degradable macromer [poly(l-lactide)(PLA)-b-PEG-b-PLA] were used for sustained release of NT-3 into the injured spinal cords of rats over the course of 2 weeks, where this combinatorial therapy induced more axonal regrowth than either individual therapy.¹⁸⁸ Such controlled release systems help to overcome the need for multiple injections or implantation events. Anderson et al. recently demonstrated that injectable biomaterials could be used as reservoirs for localized delivery of a combination of three growth factors (FGF2, EGF, and GDNF) whose synergistic actions were required to enhance recovery after SCI in a rodent model.¹⁸⁵

Biomaterials and cells

The use of biomaterials to deliver cells after SCI has been reported by several studies to improve treatment efficacy (Table 1).¹⁹⁰ Biomaterials may physically protect loaded cells from the hostile environment of SCI, retain administered cells near the injury site, and provide an adhesive matrix for cells and other cues to improve their survival.^43,191 By minimizing the loss of delivered cells, the number of cells required for achieving a therapeutic effect can also be reduced.⁴³ Promising results with the use of different biomaterial/cell combinations have been observed in experimental SCI models. These include neurotrophic factor expression,¹³ improved postimplantation cell survival,^{11,16,44,50,138} neuronal differentiation,¹⁰ and directed axonal regeneration.⁹

Although various forms of biomaterials can be used, hydrogels represent an attractive form as they can be easily used as minimally invasive therapies. A study by Kadoya et al. demonstrated some potential advantages of delivering therapeutic cells via methods that are minimally invasive, specifically those that maintain an intact dura.¹⁶ The study observed formation of ectopic stem cell colonies in rats that were administered a cell-laden fibrin matrix, which also contained a cocktail of growth factors, after opening the dura. However, no ectopic colonies were observed in closed dura delivery, where no growth factor cocktail was used.¹⁶ In addition, injection delivery of cells within hydrogel can help to reduce cell death induced by mechanical stresses.¹⁹²

Several combinations of biomaterials with cells have previously reported varying outcomes of axonal regeneration and functional recovery in experimental models of SCI (Table 1). These have included HA hydrogels with NSPCs,⁴⁴ self-assembling peptides with NSPCs,¹² PEG hydrogels with human iPSC-derived NSPCs,¹⁰ polyurethane-based gel with MSCs,¹⁹³ and Matrigel with Schwann cells.¹⁹⁴ A fibronectin-mimetic, peptide-grafted gellan gum hydrogel was combined with A-MSCs and OECs and evaluated for the treatment of experimental SCI in rats. It was found that hydrogel encapsulation of both A-MSCs and OECs led to increased axonal regeneration and significant improvement in motor function.¹⁹⁵ Delivery of OECs via PQLA/PLGA scaffolds improved cell survival and proregenerative efficacy in the rat SCI model.¹³ Delivery of mouse E14 spinal progenitors, loaded onto multichannel PLGA bridges providing a guidance architecture, showed marked improvement of axonal and functional regeneration in a mouse model of thoracic SCI.¹⁹⁶ However, another study found that collagen microfibers combined with neural progenitor cells from an alternative source failed to form a bridge across the injury site, or recovery of the motor function in a mouse SCI model.³⁹

It has been proposed that the combination of human MSCs and biomimetic hydrogels could modulate the immune cell microenvironment in SCI, leading to increased proregenerative M2 macrophage population.⁴³ The use of an MSC-seeded alginate hydrogel was found to promote linear axonal regeneration in an experimental rat C5 hemisection SCI model.⁹

In a separate study, MSCs combined with a collagen sponge were found to facilitate neurite elongation in a rat model of SCI.¹⁹⁷ In another study, injectable scaffolds made of self-assembling peptide nanofibers containing bone marrow homing, bioactive peptide motifs were described. When combined with human endometrium-derived stromal cells (hEnSCs), these materials were explored for the treatment of experimental chronic SCI in rats. The nanofibers were found to support neural differentiation of hEnSCs. In addition, higher axonal regeneration and myelination were also demonstrated.¹⁸¹ MSCs are thought to promote neuronal regeneration through neurotrophic protection and immunomodulation.⁶⁰

Cells and soluble biomolecules

Although there are some studies that have combined cells or growth factors with biomaterials for SCI treatment,^16,36–38 relatively few have used combinations of cells and growth factors alone. In some studies, cells themselves were applied as delivery vehicles for specific factors. For example, FGF2 was used to prime hDPCs that were subsequently injected into a complete transection model of SCI in rats. Some axonal regeneration and locomotor recovery were observed.¹⁹⁸ In vitro, astrocytes were transfected to overexpress nerve growth factor (NGF) and encapsulated into a collagen scaffold. They were then added to rat dorsal root ganglion culture and found to significantly enhance axonal growth.²¹ Cells were conjugated with adjuvant drug-loaded NPs and were found to enable pseudoautocrine stimulation of transplanted cells.¹⁹⁹ Finally, ESC-derived OPCs were combined with the ciliary neurotrophic factor (CNTF) and used to treat experimental SCI in rats leading to significant improvement in hind limb locomotor function.²⁰⁰

Biomaterials, soluble biomolecules, and cells

Combining biomaterials, stem cells, and biomolecules represents a critical²⁰¹ and promising¹⁴ approach that may improve recovery after SCI.³⁶ Exciting recent work has demonstrated that delivery of human NSPCs within a simple fibrin matrix containing a growth factor cocktail after SCI supports extensive engraftment with host neuronal circuits leading to functional recovery in both rodent¹⁵ and nonhuman primate models.⁴⁸ Another recent study used a combinatorial therapy comprising a scaffold containing Schwann cells modified to overexpress NT-3 and adult stem cells modified to overexpress the NT-3 receptor TrkC.²⁰² In each of these studies, results suggest that NSPCs can integrate with the host tissue and act as neuronal relays across SCI leading to functional gains.

A separate study incorporated NT-3 within silk fibroin coatings on poly(ɛ-caprolactone)-block-poly(l-lactide-co-ɛ-caprolactone) (PCLA) conduits, with enabled sustained release of the growth factor,²⁰³ which may provide advantages over delivery from a bolus or cell source. When combined with rat E14 NSPCs and transplanted into the rat thoracic spinal cavity after complete transection, incorporation of NT-3 increased NSPC survival, axonal regeneration, and functional improvement. Others have developed biomaterials as affinity-based delivery platforms (e.g., based on heparin-binding domains) for growth factors, including NT-3 and PDGF-AA, in combination with NSPCs and have found similar benefits in rodent models of SCI.^37,50

Beyond growth factors, a hydrogel delivery platform based on SH3 affinity has been used to deliver chondroitinase ABC in combination with human iPSCs, resulting in better survival of both host and transplanted cells.⁵¹ Liu et al. have described an approach where alginate biomaterials with longitudinally aligned channels to provide guidance were seeded with Schwann cells and implanted into cervical lesions in rats.⁴⁹ Adenovirus encoding BDNF was delivered caudally to the lesion to encourage axons to regenerate along the rostral/caudal axis and allow axons to exit biomaterial implants and re-enter the caudal spinal cord.

Other therapeutic approaches

Because of the complexity of the healing process of the spinal cord after SCI, taking advantage of non tissue engineering approaches may further support restoration of function. For example, small-scale clinical studies have demonstrated that the use of epidural electrical stimulation, in particular in conjunction with dynamic, task-specific training, can restore voluntary, coordinated motor activity in previously paralyzed individuals following SCI.^204–207 It is envisioned that combining proregenerative drugs, biomolecules, and/or cells with electrical stimulation and/or rehabilitative training will lead to better results. This type of approach would be expected to create a proregenerative environment supportive of axonal growth and plasticity, which would then be guided to form and maintain relays for different bodily functions via direct neuromuscular stimulation.³³ For example, Chen et al. found that treatment of rats with anti-NOGO-A antibodies after incomplete thoracic SCI followed by treadmill locomotive training improved motor recovery.²⁰⁸

Challenges and Future Perspectives

Many obstacles to the development of successful treatments for SCI remain, including reproducibility of the therapeutic paradigm (in particular with combinatorial or multimodal therapies), variations in the location and severity of SCI across patients, and clinical variability and recognizing the appropriate timing for treatment intervention, remain to be addressed. Cell-based therapies will face challenges concerning both preclinical processing and clinical administration, such as identifying the optimal number of cells for transplantation that balances efficacy and safety, improving survival of transplanted cells, and directing cell differentiation and functional integration with the host circuitry.

Important challenges also include issues related to interspecies variation and preclinical experimental models. For example, while rat SCI is more pathologically similar to the human case than mouse SCI,²⁰⁹ mice have been used more frequently in preclinical studies. Even the use of rat models is not ideal as these have fundamental differences from humans, and thus, results cannot always be translated to humans. For example, the efficacy of successful cell therapy observed in animal studies has not been reproduced in human clinical trials.¹⁷ In addition, while cervical SCI is the most common injury in human patients, it has been less often investigated in preclinical studies.⁷⁰ Instead, many preclinical studies have used lumbar and thoracic models that are technically easier to perform.²¹⁰ Therefore, more clinically representative models should be considered and developed in the future.

Given these considerations, preclinical results should be interpreted cautiously. For example, axonal regeneration does not always mean functional recovery. Efforts should be made to identify specific populations of axons responsible for functional recovery. The reversible neuronal silencing technique is a possible method for this purpose.²¹¹ However, regeneration of large numbers of axons, which can exit spinal cord lesions caudally, remains a significant challenge.¹⁷ There are also differences between animals and humans in biological cues, and hence, the extrapolation of results can be difficult.

Although there have been several reports of cell-based therapies for the treatment of SCI, including clinical trials, treatment efficacy is yet to be demonstrated in phase I-II trials.³ Recently, questions regarding the translation of preclinical data to the clinic have been raised.^46,81,82 It is difficult to define potency in cell products and current good manufacturing practice (cGMP) requires testing for the markers that are essential to identify desired cells and contaminants and as indicators of potency. In addition, the FDA does not require a definitive potency assay for early clinical studies. While this policy may serve to speed up translation of some therapies, it leaves room for variations in final products leading to a risk of failure in clinical trials.⁸¹ Thus, researchers must balance the need for extensive product characterization with that for helping patients sooner. It should be noted that no preclinical model is ideal, and thus, the ultimate testing for efficacy lies in the human clinical setting.²¹²

In the future, the use of iPSC-derived cells may help to avoid hurdles associated with possible immune rejection of allogenic cells. They may also mitigate the need to address issues with variability across lots, as each batch of therapeutic cells would be tailored for each patient. However, iPSCs will require thorough investigation into the safety of their clinical use, as possible genetic and epigenetic abnormalities may occur during their induction and tumorigenicity after their transplantation.⁸⁴ Tools, including the use of clustered regularly interspaced short palindromic repeats (CRISPR)-mediated gene editing, have helped to identify key players involved in the cell response to injury, and may aid in developing methods to enhance the ability of stem cell-based therapies to restore function after SCI.²¹³

Real-time, in vivo imaging, data communication and control can be integrated in future therapeutic strategies.^214,215 For example, the next wave of innovations would likely include smart, multifunctional materials that can sense, respond, actuate, and report with remote control.^215–217 The use of dynamically bioresponsive materials^216,217 for developing a new generation of therapeutic constructs may also be useful.²¹⁸ Such materials can be designed to provide graded, temporally and spatially controlled release of various agents,²¹⁹ and even stimuli-responsive behavior than can be triggered by external or internal means. Biomaterials may also have electroconductive elements and electroresponsive properties. In addition, including in situ organizable structures (e.g., with the use of magnetic rods⁵⁸) may promote organized guidance of regenerating axons/neurites. For example, the incorporation of superparamagnetic iron oxide NPs would help targetability under the influence of external magnetic field. This type of approach has the potential to yield new nonsurgical treatment options for SCI.²²⁰

Organ-on-a-chip technologies have been developed and used to build various models for studying normal physiology and pathological conditions.^221–223 Such models may be used in the future to study the various stages of pathophysiology of SCI, and to develop and test new therapeutics. Organ-on-a-chip models can be more representative to native tissues than conventional two-dimensional cell culture and can possibly recapitulate human cell behavior better than animal studies.^223–225 It would be a great advantage to have SCI-on-a-chip models that may help to overcome some of the present challenges, especially those related to the development of effective therapy for a difficult condition.

Commercialization of newly developed therapeutics will depend on the feasibility of scaling up for mass cGMP production, demonstration of clinical safety and efficacy, and obtaining regulatory approvals. When developing therapies integrating diverse technologies (e.g., devices, drugs, and cells), regulatory approval presents additional challenges, such as how these combination therapies affect the body differently than might be expected for each component alone. Thus, new combination products will require extensive preclinical characterization. Targeting more specific SCI pathologies when designing trials, and the use of new imaging technologies and biomarkers⁶ also promise to improve evaluation of outcomes.

There is a need to build upon already obtained clinical results to refine therapies and, in particular, cell-based therapies that are relatively new and for which the optimal potency, cell types, and dosing for therapeutic benefits are crucial to characterize. In addition, successful clinical translation will require development of safe methods for cell transplantation. Complications and risks related to the use of stem cells, such as the formation of ectopic colonies, need to be carefully assessed and resolved. More studies are needed to optimize minimally invasive delivery methods that may lead to reduced risk of cell migration.¹⁶

Moreover, a multidisciplinary approach involving clinicians, researchers, pharmacologists, materials scientists, and bioengineers will be indispensable to achieve successful treatment of patients with SCI, a very challenging, yet important, clinical goal. Development of a global consensus and standardization for cell-based therapies should be encouraged to curb the misuse of these interventions as curative means and maximize the ability of scientific studies to advance innovative treatments for SCI.⁷⁴

Conclusions

SCI is a devastating condition with a complex pathophysiology for which no present therapies can restore lost neurological functions. The last decade has produced increasingly intensive research that has led to the development of a few cell-based therapies that have proceeded to clinical trials. Unfortunately, despite successes in preclinical animal studies, efficacy has not been reproduced in humans. To date, the most successful results have been obtained using a combinatorial therapy that used biomaterials, cells, and soluble molecules. It is anticipated that addition of electrical stimulation of the spinal cord to these therapies will achieve a better outcome. At present, functional recovery after SCI is a long process that may take years and it is unlikely that a full recovery will ever be achieved. The development of minimally invasive approaches is desired to reduce complications and avoid additional injury to the spinal cord. Nevertheless, several barriers remain to be overcome through a multidisciplinary approach and consensus based on integrated evidence and recruitment of advances made in different fields of science and technology.

Footnotes

Acknowledgment

Thanks to Mohammed Xohdy for help with drawing figures.

Funding Information

The authors also acknowledge funding from the National Institutes of Health (EB021857, AR066193, AR057837, CA214411, HL137193, EB024403, EB023052, EB022403, and EB021857), Air Force Office of Sponsored Research under award No. FA9550-15-1-0273, and a National Science Foundation Career Award (S.K.S., 1653730).

Disclosure Statement

The authors acknowledge that they have no competing interests.

References

O'Shea

T.M.

, Burda

J.E.

, and Sofroniew

M.V.

Cell biology of spinal cord injury and repair. J Clin Invest, 127, 3259, 2017.

World Health Organization; The International Spinal Cord Society. International Perspectives on Spinal Cord Injury. Geneva, Switzerland: World Health Organization, 2013.

Cox

C.S.

Jr.

Cellular therapy for traumatic neurological injury. Pediatr Res, 83, 325, 2018.

Singh

, Tetreault

, Kalsi-Ryan

, Nouri

, and Fehlings

M.G.

Global prevalence and incidence of traumatic spinal cord injury. Clin Epidemiol, 6, 309, 2014.

Jazayeri

S.B.

, Beygi

, Shokraneh

, Hagen

E.M.

, and Rahimi-Movaghar

Incidence of traumatic spinal cord injury worldwide: a systematic review. Eur Spine J, 24, 905, 2015.

Ahuja

C.S.

, Nori

, Tetreault

, et al. Traumatic spinal cord injury-repair and regeneration. Neurosurgery, 80, S9, 2017.

Guilcher

S.J.

, Munce

S.E.

, Couris

C.M.

, et al. Health care utilization in non-traumatic and traumatic spinal cord injury: a population-based study. Spinal Cord, 48, 45, 2010.

Zeb

, Khan

I.N.

, Munir

, et al. Updates on therapeutics in clinical trials for spinal cord injuries: key translational applications of human embryonic stem cells-derived neural progenitors. CNS Neurol Disord Drug Targets, 15, 1266, 2016.

Gunther

M.I.

, Weidner

, Muller

, and Blesch

Cell-seeded alginate hydrogel scaffolds promote directed linear axonal regeneration in the injured rat spinal cord. Acta Biomater, 27, 140, 2015.

10.

Mosley

M.C.

, Lim

H.J.

, Chen

, et al. Neurite extension and neuronal differentiation of human induced pluripotent stem cell derived neural stem cells on polyethylene glycol hydrogels containing a continuous Young's Modulus gradient. J Biomed Mater Res A, 105, 824, 2017.

11.

Patel

, Joseph

, Patel

, et al. Suspension matrices for improved Schwann-cell survival after implantation into the injured rat spinal cord. J Neurotrauma, 27, 789, 2010.

12.

Zweckberger

, Ahuja

C.S.

, Liu

, Wang

, and Fehlings

M.G.

Self-assembling peptides optimize the post-traumatic milieu and synergistically enhance the effects of neural stem cell therapy after cervical spinal cord injury. Acta Biomater, 42, 77, 2016.

13.

Blumenthal

, Cohen-Matsliah

S.I.

, and Levenberg

Olfactory bulb-derived cells seeded on 3D scaffolds exhibit neurotrophic factor expression and pro-angiogenic properties. Tissue Eng Part A, 19, 2284, 2013.

14.

Shrestha

, Coykendall

, Li

, Moon

, Priyadarshani

, and Yao

Repair of injured spinal cord using biomaterial scaffolds and stem cells. Stem Cell Res Ther, 5, 91, 2014.

15.

, Wang

, Graham

, et al. Long-distance growth and connectivity of neural stem cells after severe spinal cord injury. Cell, 150, 1264, 2012.

16.

Kadoya

, Lu

, Nguyen

, et al. Spinal cord reconstitution with homologous neural grafts enables robust corticospinal regeneration. Nat Med, 22, 479, 2016.

17.

Assinck

, Duncan

G.J.

, Hilton

B.J.

, Plemel

J.R.

, and Tetzlaff

Cell transplantation therapy for spinal cord injury. Nat Neurosci, 20, 637, 2017.

18.

Baranes

, Shevach

, Shefi

, and Dvir

Gold nanoparticle-decorated scaffolds promote neuronal differentiation and maturation. Nano Lett, 16, 2916, 2016.

19.

Aydemir Sezer

, Ozturk

, Aru

, Yanikkaya Demirel

, Sezer

, and Bozkurt

M.R.

Zero valent zinc nanoparticles promote neuroglial cell proliferation: a biodegradable and conductive filler candidate for nerve regeneration. J Mater Sci Mater Med, 28, 19, 2017.

20.

Tay

, Sohrabi

, Poole

, Seidlits

, and Di Carlo

A 3D magnetic hyaluronic acid hydrogel for magnetomechanical neuromodulation of primary dorsal root ganglion neurons. Adv Mater, 30, e1800927, 2018.

21.

Berndt

, Li

, Seyedhassantehrani

, and Yao

Fabrication and characterization of microspheres encapsulating astrocytes for neural regeneration. ACS Biomater Sci Eng, 3, 1313, 2017.

22.

Saghazadeh

, and Rezaei

The role of timing in the treatment of spinal cord injury. Biomed Pharmacother, 92, 128, 2017.

23.

Zhang

, Chen

, and Wang

Serum amyloid A induces a vascular smooth muscle cell phenotype switch through the p38 MAPK signaling pathway. Biomed Res Int, 2017, 4941379, 2017.

24.

Springer

J.E.

, Visavadiya

N.P.

, Sullivan

P.G.

, and Hall

E.D.

Post-injury treatment with NIM811 promotes recovery of function in adult female rats after spinal cord contusion: a dose-response study. J Neurotrauma, 35, 492, 2018.

25.

Lang

B.T.

, Cregg

J.M.

, DePaul

M.A.

, et al. Modulation of the proteoglycan receptor PTPsigma promotes recovery after spinal cord injury. Nature, 518, 404, 2015.

26.

Thompson

R.E.

, Pardieck

, Smith

, et al. Effect of hyaluronic acid hydrogels containing astrocyte-derived extracellular matrix and/or V2a interneurons on histologic outcomes following spinal cord injury. Biomaterials, 162, 208, 2018.

27.

Anderson

M.A.

, Burda

J.E.

, Ren

, et al. Astrocyte scar formation aids central nervous system axon regeneration. Nature, 532, 195, 2016.

28.

Tracy

, Armola

, and Micham

The “cold cord”: a review of therapeutic hypothermia for traumatic spinal cord injuries. Am J Crit Care, 24, 540, 2015.

29.

Kwon

B.K.

, Curt

, Belanger

L.M.

et al. Intrathecal pressure monitoring and cerebrospinal fluid drainage in acute spinal cord injury: a prospective randomized trial. J Neurosurg Spine, 10, 181, 2009.

30.

Sen

C.K.

, Gordillo

G.M.

, Roy

, et al. Human skin wounds: a major and snowballing threat to public health and the economy. Wound Repair Regen, 17, 763, 2009.

31.

Kucher

, Johns

, Maier

, et al. First-in-man intrathecal application of neurite growth-promoting anti-Nogo-A antibodies in acute spinal cord injury. Neurorehabil Neural Repair, 32, 578, 2018.

32.

Ross

C.L.

, Syed

, Smith

T.L.

, and Harrison

B.S.

The regenerative effects of electromagnetic field on spinal cord injury. Electromagn Biol Med, 36, 74, 2017.

33.

Hamid

, and Hayek

Role of electrical stimulation for rehabilitation and regeneration after spinal cord injury: an overview. Eur Spine J, 17, 1256, 2008.

34.

Rocha

L.A.

, Sousa

R.A.

, Learmonth

D.A.

, and Salgado

A.J.

The role of biomaterials as angiogenic modulators of spinal cord injury: mimetics of the spinal cord, cell and angiogenic factor delivery agents. Front Pharmacol, 9, 164, 2018.

35.

, Guo

G.H.

, Wang

G.S.

, Guan

C.X.

, and Yue

Influence of neural stem cell transplantation on angiogenesis in rats with spinal cord injury. Genet Mol Res, 13, 6083, 2014.

36.

Wilems

T.S.

, Pardieck

, Iyer

, and Sakiyama-Elbert

S.E.

Combination therapy of stem cell derived neural progenitors and drug delivery of anti-inhibitory molecules for spinal cord injury. Acta Biomater, 28, 23, 2015.

37.

Johnson

P.J.

, Tatara

, McCreedy

D.A.

, Shiu

, and Sakiyama-Elbert

S.E.

Tissue-engineered fibrin scaffolds containing neural progenitors enhance functional recovery in a subacute model of SCI. Soft Matter, 6, 5127, 2010.

38.

Johnson

P.J.

, Tatara

, Shiu

, and Sakiyama-Elbert

S.E.

Controlled release of neurotrophin-3 and platelet-derived growth factor from fibrin scaffolds containing neural progenitor cells enhances survival and differentiation into neurons in a subacute model of SCI. Cell Transplant, 19, 89, 2010.

39.

Sugai

, Nishimura

, Kato-Negishi

, et al. Neural stem/progenitor cell-laden microfibers promote transplant survival in a mouse transected spinal cord injury model. J Neurosci Res, 93, 1826, 2015.

40.

Hatami

, Mehrjardi

N.Z.

, Kiani

, et al. Human embryonic stem cell-derived neural precursor transplants in collagen scaffolds promote recovery in injured rat spinal cord. Cytotherapy, 11, 618, 2009.

41.

McCreedy

D.A.

, Wilems

T.S.

, Xu

, et al. Survival, differentiation, and migration of high-purity mouse embryonic stem cell-derived progenitor motor neurons in fibrin scaffolds after sub-acute spinal cord injury. Biomater Sci, 2, 1672, 2014.

42.

Itosaka

, Kuroda

, Shichinohe

, et al. Fibrin matrix provides a suitable scaffold for bone marrow stromal cells transplanted into injured spinal cord: a novel material for CNS tissue engineering. Neuropathology, 29, 248, 2009.

43.

Caron

, Rossi

, Papa

, et al. A new three dimensional biomimetic hydrogel to deliver factors secreted by human mesenchymal stem cells in spinal cord injury. Biomaterials, 75, 135, 2016.

44.

Mothe

A.J.

, Tam

R.Y.

, Zahir

, Tator

C.H.

, and Shoichet

M.S.

Repair of the injured spinal cord by transplantation of neural stem cells in a hyaluronan-based hydrogel. Biomaterials, 34, 3775, 2013.

45.

Wilems

T.S.

, and Sakiyama-Elbert

S.E.

Sustained dual drug delivery of anti-inhibitory molecules for treatment of spinal cord injury. J Control Release, 213, 103, 2015.

46.

Anderson

A.J.

, Piltti

K.M.

, Hooshmand

M.J.

, Nishi

R.A.

, and Cummings

B.J.

Preclinical efficacy failure of human CNS-derived stem cells for use in the pathway study of cervical spinal cord injury. Stem Cell Reports, 8, 249, 2017.

47.

Yao

, Daly

, Newland

, et al. Improved axonal regeneration of transected spinal cord mediated by multichannel collagen conduits functionalized with neurotrophin-3 gene. Gene Ther, 20, 1149, 2013.

48.

Rosenzweig

E.S.

, Brock

J.H.

, Lu

, et al. Restorative effects of human neural stem cell grafts on the primate spinal cord. Nat Med, 24, 484, 2018.

49.

Liu

, Sandner

, Schackel

, et al. Regulated viral BDNF delivery in combination with Schwann cells promotes axonal regeneration through capillary alginate hydrogels after spinal cord injury. Acta Biomater, 60, 167, 2017.

50.

Fuhrmann

, Tam

R.Y.

, Ballarin

, et al. Injectable hydrogel promotes early survival of induced pluripotent stem cell-derived oligodendrocytes and attenuates longterm teratoma formation in a spinal cord injury model. Biomaterials, 83, 23, 2016.

51.

Fuhrmann

, Anandakumaran

P.N.

, Payne

S.L.

, et al. Combined delivery of chondroitinase ABC and human induced pluripotent stem cell-derived neuroepithelial cells promote tissue repair in an animal model of spinal cord injury. Biomed Mater, 13, 024103, 2018.

52.

Theodore

, Hlubek

, Danielson

, et al. First human implantation of a bioresorbable polymer scaffold for acute traumatic spinal cord injury: a clinical pilot study for safety and feasibility. Neurosurgery, 79, E305, 2016.

53.

Hakim

J.S.

, Esmaeili Rad

, Grahn

P.J.

, et al. Positively charged oligo[poly(ethylene glycol) fumarate] scaffold implantation results in a permissive lesion environment after spinal cord injury in rat. Tissue Eng Part A, 21, 2099, 2015.

54.

Chen

B.K.

, Madigan

N.N.

, Hakim

J.S.

, et al. GDNF Schwann cells in hydrogel scaffolds promote regional axon regeneration, remyelination and functional improvement after spinal cord transection in rats. J Tissue Eng Regen Med, 12, e398, 2018.

55.

Ruzicka

, Romanyuk

, Hejcl

, et al. Treating spinal cord injury in rats with a combination of human fetal neural stem cells and hydrogels modified with serotonin. Acta Neurobiol Exp (Wars), 73, 102, 2013.

56.

Moradi

, Bahktiari

, Joghataei

M.T.

, et al. BD PuraMatrix peptide hydrogel as a culture system for human fetal Schwann cells in spinal cord regeneration. J Neurosci Res, 90, 2335, 2012.

57.

Yang

Y.H.

, Khan

, Ma

, Lim

H.J.

, and Smith Callahan

L.A.

Optimization of adhesive conditions for neural differentiation of murine embryonic stem cells using hydrogels functionalized with continuous Ile-Lys-Val-Ala-Val concentration gradients. Acta Biomater, 21, 55, 2015.

58.

Rose

J.C.

, Camara-Torres

, Rahimi

, Kohler

, Moller

, and De Laporte

Nerve cells decide to orient inside an injectable hydrogel with minimal structural guidance. Nano Lett, 17, 3782, 2017.

59.

Kanekiyo

, Wakabayashi

, Nakano

, et al. Effects of intrathecal injection of the conditioned medium from bone marrow stromal cells on spinal cord injury in rats. J Neurotrauma, 35, 521, 2018.

60.

Han

, Wang

, Li

, et al. Bone marrow-derived mesenchymal stem cells in three-dimensional culture promote neuronal regeneration by neurotrophic protection and immunomodulation. J Biomed Mater Res A, 104, 1759, 2016.

61.

Torres-Espin

, Hernandez

, and Navarro

Gene expression changes in the injured spinal cord following transplantation of mesenchymal stem cells or olfactory ensheathing cells. PLoS One, 8, e76141, 2013.

62.

Kobolak

, Dinnyes

, Memic

, Khademhosseini

, and Mobasheri

Mesenchymal stem cells: identification, phenotypic characterization, biological properties and potential for regenerative medicine through biomaterial micro-engineering of their niche. Methods, 99, 62, 2016.

63.

Curtis

, Martin

J.R.

, Gabel

, et al. A first-in-human, phase I study of neural stem cell transplantation for chronic spinal cord injury. Cell Stem Cell, 22, 941, 2018.

64.

Asterias Biotherapeutics. Dose escalation study of AST-OPC1 in spinal cord injury. 2018. Available at: https://ClinicalTrials.gov/show/NCT02302157 (accessed July 3, 2019).

65.

Tuszynski

M.H.

, Wang

, Graham

, et al. Neural stem cell dissemination after grafting to CNS injury sites. Cell, 156, 388, 2014.

66.

Steward

, Sharp

K.G.

, and Matsudaira Yee

Long-distance migration and colonization of transplanted neural stem cells. Cell, 156, 385, 2014.

67.

Steward

, Sharp

K.G.

, Yee

K.M.

, Hatch

M.N.

, and Bonner

J.F.

Characterization of ectopic colonies that form in widespread areas of the nervous system with neural stem cell transplants into the site of a severe spinal cord injury. J Neurosci, 34, 14013, 2014.

68.

Nguyen

H.X.

, Hooshmand

M.J.

, Saiwai

, et al. Systemic neutrophil depletion modulates the migration and fate of transplanted human neural stem cells to rescue functional repair. J Neurosci, 37, 9269, 2017.

69.

Hofstetter

C.P.

, Holmstrom

N.A.

, Lilja

J.A.

, et al. Allodynia limits the usefulness of intraspinal neural stem cell grafts; directed differentiation improves outcome. Nat Neurosci, 8, 346, 2005.

70.

Vismara

, Papa

, Rossi

, Forloni

, and Veglianese

Current options for cell therapy in spinal cord injury. Trends Mol Med, 23, 831, 2017.

71.

Lin

, and Du

The role of secreted factors in stem cells-mediated immune regulation. Cell Immunol, 326, 24, 2018.

72.

StemCells, Inc. Study of human central nervous system (CNS) stem cell transplantation in cervical spinal cord injury. 2016. Available at: https://ClinicalTrials.gov/show/NCT02163876 (accessed July 3, 2019).

73.

Scott

C.T.

, and Magnus

Wrongful termination: lessons from the Geron clinical trial. Stem Cells Transl Med, 3, 1398, 2014.

74.

Chhabra

H.S.

, and Sarda

Clinical translation of stem cell based interventions for spinal cord injury—are we there yet? Adv Drug Deliv Rev, 120, 41, 2017.

75.

Neuralstem, Inc. Safety study of human spinal cord-derived neural stem cell transplantation for the treatment of chronic SCI. 2017. Available at: https://ClinicalTrials.gov/show/NCT01772810 (accessed July 3, 2019).

76.

Ghobrial

G.M.

, Anderson

K.D.

, Dididze

, et al. Human neural stem cell transplantation in chronic cervical spinal cord injury: functional outcomes at 12 months in a phase II clinical trial. Neurosurgery, 64, 87, 2017.

77.

Ferrer Internacional S.A.. Safety and preliminary efficacy of FAB117-HC in patients with acute traumatic spinal cord injury (SPINE). 2018. Available at: https://ClinicalTrials.gov/show/NCT02917291 (accessed July 3, 2019).

78.

Rong

Study of human central nervous system (CNS) stem cell transplantation in cervical spinal cord injury. Available at: https://ClinicalTrials.gov/show/NCT02163876 (accessed July 3, 2019).

79.

Banc de Sang i Teixits. Intrathecal administration of ex-panded Wharton's jelly mesenchymal stem cells in chronic traumatic spinal cord injury. 2018. Available at: https://ClinicalTrials.gov/show/NCT03003364 (accessed July 3, 2019).

80.

Sakai

, Yamamoto

, Matsubara

, et al. Human dental pulp-derived stem cells promote locomotor recovery after complete transection of the rat spinal cord by multiple neuro-regenerative mechanisms. J Clin Invest, 122, 80, 2012.

81.

Temple

, and Studer

Lessons learned from pioneering neural stem cell studies. Stem Cell Reports, 8, 191, 2017.

82.

Monuki

E.S.

, Anderson

A.J.

, Blurton-Jones

, and Cummings

B.J.

Response to StemCells Inc. Stem Cell Reports, 8, 195, 2017.

83.

StemCells, Inc.. Long-term follow-up of transplanted hu-man central nervous system stem cells (HuCNS-SC) in spinal cord trauma subjects. 2016. Available at: https://ClinicalTrials.gov/show/NCT01725880 (accessed July 3, 2019).

84.

Nagoshi

, and Okano

iPSC-derived neural precursor cells: potential for cell transplantation therapy in spinal cord injury. Cell Mol Life Sci, 75, 989, 2018.

85.

Sykova

, Homola

, Mazanec

, et al. Autologous bone marrow transplantation in patients with subacute and chronic spinal cord injury. Cell Transplant, 15, 675, 2006.

86.

Tsai

E.C.

, Dalton

P.D.

, Shoichet

M.S.

, and Tator

C.H.

Matrix inclusion within synthetic hydrogel guidance channels improves specific supraspinal and local axonal regeneration after complete spinal cord transection. Biomaterials, 27, 519, 2006.

87.

Nomura

, Katayama

, Shoichet

M.S.

, and Tator

C.H.

Complete spinal cord transection treated by implantation of a reinforced synthetic hydrogel channel results in syringomyelia and caudal migration of the rostral stump. Neurosurgery, 59, 183, 2006.

88.

Ashammakhi

Drug release: proper control to help clinical application. J Craniofac Surg, 29, 124, 2018.

89.

Rossi

, Perale

, Papa

, Forloni

, and Veglianese

Current options for drug delivery to the spinal cord. Expert Opin Drug Deliv, 10, 385, 2013.

90.

AOSpine North America Research Network. Riluzole in spinal cord injury study (RISCIS). 2019. Available at: https://ClinicalTrials.gov/show/NCT01597518 (accessed July 3, 2019).

91.

Acorda Therapeutics. AC105 in patients with acute trau-matic spinal cord injury (AC105). 2018. Available at: https://ClinicalTrials.gov/show/NCT01750684 (accessed July 3, 2019).

92.

Rick Hansen Institute. Minocycline in acute spinal cord injury (MASC). 2014. Available at: https://ClinicalTrials.gov/show/NCT01828203 (accessed July 3, 2019).

93.

Kringle Pharma, Inc.. Phase I/II study of KP-100IT in acute spinal cord injury. 2019. Available at: https://ClinicalTrials.gov/show/NCT02193334 (accessed July 3, 2019).

94.

Takahashi

, Yamazaki

, Okawa

, et al. Neuroprotective therapy using granulocyte colony-stimulating factor for acute spinal cord injury: a phase I/IIa clinical trial. Eur Spine J, 21, 2580, 2012.

95.

Bregman

B.S.

, Kunkel-Bagden

, Schnell

, Dai

H.N.

, Gao

, and Schwab

M.E.

Recovery from spinal cord injury mediated by antibodies to neurite growth inhibitors. Nature, 378, 498, 1995.

96.

Danilov

C.A.

, and Steward

Conditional genetic deletion of PTEN after a spinal cord injury enhances regenerative growth of CST axons and motor function recovery in mice. Exp Neurol, 266, 147, 2015.

97.

Lewandowski

, and Steward

AAVshRNA-mediated suppression of PTEN in adult rats in combination with salmon fibrin administration enables regenerative growth of corticospinal axons and enhances recovery of voluntary motor function after cervical spinal cord injury. J Neurosci, 34, 9951, 2014.

98.

Mokalled

M.H.

, Patra

, Dickson

A.L.

, Endo

, Stainier

D.Y.

, and Poss

K.D.

Injury-induced ctgfa directs glial bridging and spinal cord regeneration in zebrafish. Science, 354, 630, 2016.

99.

Tedeschi

, Dupraz

, Laskowski

C.J.

, et al. The calcium channel subunit Alpha2delta2 suppresses axon regeneration in the adult CNS. Neuron, 92, 419, 2016.

100.

Bao

, Wang

, Yang

, and Chen

Blockade of interleukin-7 receptor shapes macrophage alternative activation and promotes functional recovery after spinal cord injury. Neuroscience, 371, 518, 2018.

101.

Park

, Decker

J.T.

, Smith

D.R.

, Cummings

B.J.

, Anderson

A.J.

, and Shea

L.D.

Reducing inflammation through delivery of lentivirus encoding for anti-inflammatory cytokines attenuates neuropathic pain after spinal cord injury. J Control Release, 290, 88, 2018.

102.

Margul

D.J.

, Park

, Boehler

R.M.

, et al. Reducing neuroinflammation by delivery of IL-10 encoding lentivirus from multiple-channel bridges. Bioeng Transl Med, 1, 136, 2016.

103.

Mokarram

, Dymanus

, Srinivasan

, et al. Immunoengineering nerve repair. Proc Natl Acad Sci U S A, 114, E5077, 2017.

104.

Nakano

, Nakai

, Seo

T.B.

, et al. Characterization of conditioned medium of cultured bone marrow stromal cells. Neurosci Lett, 483, 57, 2010.

105.

Shen

, Liu

, Zhang

, et al. CCR2 positive exosome released by mesenchymal stem cells suppresses macrophage functions and alleviates ischemia/reperfusion-induced renal injury. Stem Cells Int, 2016, 1240301, 2016.

106.

Barnabe-Heider

, Goritz

, Sabelstrom

, et al. Origin of new glial cells in intact and injured adult spinal cord. Cell Stem Cell, 7, 470, 2010.

107.

Marichal

, Reali

, Rehermann

M.I.

, Trujillo-Cenoz

, and Russo

R.E.

Progenitors in the ependyma of the spinal cord: a potential resource for self-repair after injury. Adv Exp Med Biol, 1015, 241, 2017.

108.

Sabelstrom

, Stenudd

, Reu

, et al. Resident neural stem cells restrict tissue damage and neuronal loss after spinal cord injury in mice. Science, 342, 637, 2013.

109.

Sabelstrom

, Stenudd

, and Frisen

Neural stem cells in the adult spinal cord. Exp Neurol, 260, 44, 2014.

110.

Horner

P.J.

, and Gage

F.H.

Regenerating the damaged central nervous system. Nature, 407, 963, 2000.

111.

Smith

D.R.

, Margul

D.J.

, Dumont

C.M.

, et al. Combinatorial lentiviral gene delivery of pro-oligodendrogenic factors for improving myelination of regenerating axons after spinal cord injury. Biotechnol Bioeng, 116, 155, 2019.

112.

Thomas

A.M.

, Seidlits

S.K.

, Goodman

A.G.

, et al. Sonic hedgehog and neurotrophin-3 increase oligodendrocyte numbers and myelination after spinal cord injury. Integr Biol (Camb), 6, 694, 2014.

113.

Forgione

, and Fehlings

M.G.

Rho-ROCK inhibition in the treatment of spinal cord injury. World Neurosurg, 82, e535, 2014.

114.

Khorasanizadeh

, Eskian

, Vaccaro

A.R.

, and Rahimi-Movaghar

Granulocyte colony-stimulating factor (G-CSF) for the treatment of spinal cord injury. CNS Drugs, 31, 911, 2017.

115.

Fehlings

M.G.

, Theodore

, Harrop

, et al. A phase I/IIa clinical trial of a recombinant Rho protein antagonist in acute spinal cord injury. J Neurotrauma, 28, 787, 2011.

116.

Vertex Pharmaceuticals, Incorporated. Study to assess the efficacy and safety of VX-210 in subjects with acute traumatic cervical spinal cord injury. 2018. Available at: https://ClinicalTrials.gov/show/NCT02669849 (accessed July 3, 2019).

117.

Kamiya

, Koda

, Furuya

, et al. Neuroprotective therapy with granulocyte colony-stimulating factor in acute spinal cord injury: a comparison with high-dose methylprednisolone as a historical control. Eur Spine J, 24, 963, 2015.

118.

Derakhshanrad

, Saberi

, Yekaninejad

M.S.

, Joghataei

M.T.

, and Sheikhrezaei

Granulocyte-colony stimulating factor administration for neurological improvement in patients with postrehabilitation chronic incomplete traumatic spinal cord injuries: a double-blind randomized controlled clinical trial. J Neurosurg Spine, 29, 97, 2018.

119.

Kitamura

, Nagoshi

, Tsuji

, Matsumoto

, Okano

, and Nakamura

Application of hepatocyte growth factor for acute spinal cord injury: the road from basic studies to human treatment. Int J Mol Sci, 20, 1054, 2019.

120.

InVivo Therapeutics. The INSPIRE study: probable benefit of the neuro-spinal scaffold for treatment of AIS A thoracic acute spinal cord injury. 2018. Available at: https://ClinicalTrials.gov/show/NCT02138110 (accessed July 3, 2019).

121.

Guest

J.D.

, Moore

S.W.

, Aimetti

A.A.

, et al. Internal decompression of the acutely contused spinal cord: differential effects of irrigation only versus biodegradable scaffold implantation. Biomaterials, 185, 284, 2018.

122.

, Zhang

, Zeng

, et al. Manufacture of PLGA multiple-channel conduits with precise hierarchical pore architectures and in vitro/vivo evaluation for spinal cord injury. Tissue Eng Part C Methods, 15, 243, 2009.

123.

Kim

, Tator

C.H.

, and Shoichet

M.S.

Chitosan implants in the rat spinal cord: biocompatibility and biodegradation. J Biomed Mater Res A, 97, 395, 2011.

124.

Gelain

, Panseri

, Antonini

, et al. Transplantation of nanostructured composite scaffolds results in the regeneration of chronically injured spinal cords. ACS Nano, 5, 227, 2011.

125.

Tysseling-Mattiace

V.M.

, Sahni

, Niece

K.L.

, et al. Self-assembling nanofibers inhibit glial scar formation and promote axon elongation after spinal cord injury. J Neurosci, 28, 3814, 2008.

126.

Macaya

, and Spector

Injectable hydrogel materials for spinal cord regeneration: a review. Biomed Mater, 7, 012001, 2012.

127.

Perale

, Rossi

, Sundstrom

, et al. Hydrogels in spinal cord injury repair strategies. ACS Chem Neurosci, 2, 336, 2011.

128.

Woerly

, Petrov

, Sykova

, Roitbak

, Simonova

, and Harvey

A.R.

Neural tissue formation within porous hydrogels implanted in brain and spinal cord lesions: ultrastructural, immunohistochemical, and diffusion studies. Tissue Eng, 5, 467, 1999.

129.

Hejcl

, Lesny

, Pradny

, et al. Macroporous hydrogels based on 2-hydroxyethyl methacrylate. Part 6: 3D hydrogels with positive and negative surface charges and polyelectrolyte complexes in spinal cord injury repair. J Mater Sci Mater Med, 20, 1571, 2009.

130.

Kataoka

, Suzuki

, Kitada

, et al. Alginate enhances elongation of early regenerating axons in spinal cord of young rats. Tissue Eng, 10, 493, 2004.

131.

Perale

, Rossi

, Santoro

, et al. Multiple drug delivery hydrogel system for spinal cord injury repair strategies. J Control Release, 159, 271, 2012.

132.

Ziemba

A.M.

, and Gilbert

R.J.

Biomaterials for local, controlled drug delivery to the injured spinal cord. Front Pharmacol, 8, 245, 2017.

133.

Fukunaga

, Tsutsumi

, and Mihara

Cell differentiation on disk- and string-shaped hydrogels fabricated from Ca(2+)-responsive self-assembling peptides. Biopolymers, 106, 476, 2016.

134.

H.Y.

, Fuhrmann

, Zhou

, and Dalton

P.D.

Host reaction to poly(2-hydroxyethyl methacrylate) scaffolds in a small spinal cord injury model. J Mater Sci Mater Med, 24, 2001, 2013.

135.

Grulova

, Slovinska

, Blasko

, et al. Delivery of alginate scaffold releasing two trophic factors for spinal cord injury repair. Sci Rep, 5, 13702, 2015.

136.

Yang

, De Laporte

, Zelivyanskaya

M.L.

, et al. Multiple channel bridges for spinal cord injury: cellular characterization of host response. Tissue Eng Part A, 15, 3283, 2009.

137.

Olson

H.E.

, Rooney

G.E.

, Gross

, et al. Neural stem cell- and Schwann cell-loaded biodegradable polymer scaffolds support axonal regeneration in the transected spinal cord. Tissue Eng Part A, 15, 1797, 2009.

138.

Dumont

C.M.

, Carlson

M.A.

, Munsell

M.K.

, et al. Aligned hydrogel tubes guide regeneration following spinal cord injury. Acta Biomater, 86, 312, 2019.

139.

, Yang

, Li

, et al. A therapeutic strategy for spinal cord defect: human dental follicle cells combined with aligned PCL/PLGA electrospun material. Biomed Res Int, 2015, 197183, 2015.

140.

Espinosa-Jeffrey

, Oregel

, Wiggins

, et al. Strategies for endogenous spinal cord repair: HPMA hydrogel to recruit migrating endogenous stem cells. Adv Exp Med Biol, 760, 25, 2012.

141.

Slotkin

J.R.

, Pritchard

C.D.

, Luque

, et al. Biodegradable scaffolds promote tissue remodeling and functional improvement in non-human primates with acute spinal cord injury. Biomaterials, 123, 63, 2017.

142.

Tuinstra

H.M.

, Margul

D.J.

, Goodman

A.G.

, et al. Long-term characterization of axon regeneration and matrix changes using multiple channel bridges for spinal cord regeneration. Tissue Eng Part A, 20, 1027, 2014.

143.

Makadia

H.K.

, and Siegel

S.J.

Poly lactic-co-glycolic acid (PLGA) as biodegradable controlled drug delivery carrier. Polymers (Basel), 3, 1377, 2011.

144.

Chu

D.S.

, Sellers

D.L.

, Bocek

M.J.

, Fischedick

A.E.

, Horner

P.J.

, and Pun

S.H.

MMP9-sensitive polymers mediate environmentally-responsive bivalirudin release and thrombin inhibition. Biomater Sci, 3, 41, 2015.

145.

Teng

Y.D.

, Lavik

E.B.

, Qu

, 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, 99, 3024, 2002.

146.

Kamperman

, Henke

, van den Berg

, et al. Single cell microgel based modular bioinks for uncoupled cellular micro- and macroenvironments. Adv Healthc Mater, 6, 2017.

147.

Lienemann

P.S.

, Rossow

, Mao

A.S.

, Vallmajo-Martin

, Ehrbar

, and Mooney

D.J.

Single cell-laden protease-sensitive microniches for long-term culture in 3D. Lab Chip, 17, 727, 2017.

148.

Pakulska

M.M.

, Ballios

B.G.

, and Shoichet

M.S.

Injectable hydrogels for central nervous system therapy. Biomed Mater, 7, 024101, 2012.

149.

Khaing

Z.Z.

, Ehsanipour

, Hofstetter

C.P.

, and Seidlits

S.K.

Injectable hydrogels for spinal cord repair: a focus on swelling and intraspinal pressure. Cells Tissues Organs, 202, 67, 2016.

150.

Kong

X.B.

, Tang

Q.Y.

, Chen

X.Y.

, Tu

, Sun

S.Z.

, and Sun

Z.L.

Polyethylene glycol as a promising synthetic material for repair of spinal cord injury. Neural Regen Res, 12, 1003, 2017.

151.

Horn

E.M.

, Beaumont

, Shu

X.Z.

, et al. Influence of cross-linked hyaluronic acid hydrogels on neurite outgrowth and recovery from spinal cord injury. J Neurosurg Spine, 6, 133, 2007.

152.

Park

, Lim

, Back

, Na

, Park

, and Sun

Nerve regeneration following spinal cord injury using matrix metalloproteinase-sensitive, hyaluronic acid-based biomimetic hydrogel scaffold containing brain-derived neurotrophic factor. J Biomed Mater Res A, 93, 1091, 2010.

153.

Khaing

Z.Z.

, Milman

B.D.

, Vanscoy

J.E.

, Seidlits

S.K.

, Grill

R.J.

, and Schmidt

C.E.

High molecular weight hyaluronic acid limits astrocyte activation and scar formation after spinal cord injury. J Neural Eng, 8, 046033, 2011.

154.

Gupta

, Tator

C.H.

, and Shoichet

M.S.

Fast-gelling injectable blend of hyaluronan and methylcellulose for intrathecal, localized delivery to the injured spinal cord. Biomaterials, 27, 2370, 2006.

155.

Fuhrmann

, Obermeyer

, Tator

C.H.

, and Shoichet

M.S.

Click-crosslinked injectable hyaluronic acid hydrogel is safe and biocompatible in the intrathecal space for ultimate use in regenerative strategies of the injured spinal cord. Methods, 84, 60, 2015.

156.

Kubinova

, Horak

, Hejcl

, et al. SIKVAV-modified highly superporous PHEMA scaffolds with oriented pores for spinal cord injury repair. J Tissue Eng Regen Med, 9, 1298, 2015.

157.

Tavakol

, Mousavi

S.M.M.

, Tavakol

, Hoveizi

, Ai

, and Sorkhabadi

S.M.R.

Mechano-transduction signals derived from self-assembling peptide nanofibers containing long motif of laminin influence neurogenesis in in-vitro and in-vivo. Mol Neurobiol, 54, 2483, 2017.

158.

Tukmachev

, Forostyak

, Koci

, et al. Injectable extracellular matrix hydrogels as scaffolds for spinal cord injury repair. Tissue Eng Part A, 22, 306, 2016.

159.

Zhu

, Lu

, Yu

, et al. Effect of decellularized spinal scaffolds on spinal axon regeneration in rats. J Biomed Mater Res A, 106, 698, 2018.

160.

Pawar

, Cummings

B.J.

, Thomas

, et al. Biomaterial bridges enable regeneration and re-entry of corticospinal tract axons into the caudal spinal cord after SCI: association with recovery of forelimb function. Biomaterials, 65, 1, 2015.

161.

Koffler

, Zhu

, Qu

, et al. Biomimetic 3D-printed scaffolds for spinal cord injury repair. Nat Med, 25, 263, 2019.

162.

Liu

, and Blesch

Targeted tissue engineering: hydrogels with linear capillary channels for axonal regeneration after spinal cord injury. Neural Regen Res, 13, 641, 2018.

163.

Ashammakhi

, Ndreu

, Yang

, Ylikauppila

, Nikkola

, and Hasirci

Tissue engineering: a new take-off using nanofiber-based scaffolds. J Craniofac Surg, 18, 3, 2007.

164.

Ashammakhi

, Ndreu

, Piras

A.M.

, et al. Biodegradable nanomats produced by electrospinning: expanding multifunctionality and potential for tissue engineering. J Nanosci Nanotechnol, 7, 862, 2007.

165.

Schnell

, Klinkhammer

, Balzer

, et al. Guidance of glial cell migration and axonal growth on electrospun nanofibers of poly-epsilon-caprolactone and a collagen/poly-epsilon-caprolactone blend. Biomaterials, 28, 3012, 2007.

166.

Xie

, Willerth

S.M.

, Li

, et al. The differentiation of embryonic stem cells seeded on electrospun nanofibers into neural lineages. Biomaterials, 30, 354, 2009.

167.

Jakobsson

, Ottosson

, Zalis

M.C.

, O'Carroll

, Johansson

U.E.

, and Johansson

Three-dimensional functional human neuronal networks in uncompressed low-density electrospun fiber scaffolds. Nanomedicine, 13, 1563, 2017.

168.

Ren

, Han

, Zhou

, et al. Repair of spinal cord injury by inhibition of astrocyte growth and inflammatory factor synthesis through local delivery of flavopiridol in PLGA nanoparticles. Biomaterials, 35, 6585, 2014.

169.

Carvalho-de-Souza

J.L.

, Treger

J.S.

, Dang

, Kent

S.B.

, Pepperberg

D.R.

, and Bezanilla

Photosensitivity of neurons enabled by cell-targeted gold nanoparticles. Neuron, 86, 207, 2015.

170.

Eom

, Kim

, Choi

J.M.

, et al. Enhanced infrared neural stimulation using localized surface plasmon resonance of gold nanorods. Small, 10, 3853, 2014.

171.

, Lee

S.Y.

, Wu

, et al. Neuroprotective ferulic acid (FA)-glycol chitosan (GC) nanoparticles for functional restoration of traumatically injured spinal cord. Biomaterials, 35, 2355, 2014.

172.

Ross

C.L.

The use of electric, magnetic, and electromagnetic field for directed cell migration and adhesion in regenerative medicine. Biotechnol Prog, 33, 5, 2017.

173.

Omidinia-Anarkoli

, Boesveld

, Tuvshindorj

, Rose

J.C.

, Haraszti

, and De Laporte

An injectable hybrid hydrogel with oriented short fibers induces unidirectional growth of functional nerve cells. Small, 13, 1702207, 2017.

174.

, Luciani

, Richard

, et al. A 3D magnetic tissue stretcher for remote mechanical control of embryonic stem cell differentiation. Nat Commun, 8, 400, 2017.

175.

Song

, Sun

, Liu

, et al. Polymerizing pyrrole coated poly (l-lactic acid-co-epsilon-caprolactone) (PLCL) conductive nanofibrous conduit combined with electric stimulation for long-range peripheral nerve regeneration. Front Mol Neurosci, 9, 117, 2016.

176.

Zhou

J.F.

, Wang

Y.G.

, Cheng

, Wu

, Sun

X.D.

, and Peng

Preparation of polypyrrole-embedded electrospun poly(lactic acid) nanofibrous scaffolds for nerve tissue engineering. Neural Regen Res, 11, 1644, 2016.

177.

Zhu

, Ye

, Lee

S.J.

, et al. Enhanced neural stem cell functions in conductive annealed carbon nanofibrous scaffolds with electrical stimulation. Nanomedicine, 14, 2485, 2018.

178.

Liu

, Ma

, Wu

, Lin

, and Wang

Polymeric nanofibers with ultrahigh piezoelectricity via self-orientation of nanocrystals. ACS Nano, 11, 1901, 2017.

179.

Wang

, Wang

, Zhang

, Jian

, and Zhang

Feeding single-walled carbon nanotubes or graphene to silkworms for reinforced silk fibers. Nano Lett, 16, 6695, 2016.

180.

Tavakol

, Saber

, Hoveizi

, Aligholi

, Ai

, and Rezayat

S.M.

Chimeric self-assembling nanofiber containing bone marrow homing peptide's motif induces motor neuron recovery in animal model of chronic spinal cord injury; an in vitro and in vivo investigation. Mol Neurobiol, 53, 3298, 2016.

181.

Ashammakhi

, Kaarela

, and Ferretti

Pulling and pushing stem cells to control their differentiation. J Craniofac Surg, 29, 804, 2018.

182.

Führmann

, Anandakumaran

P.N.

, and Shoichet

M.S.

Combinatorial therapies after spinal cord injury: how can biomaterials help? Adv Healthc Mater, 6, 1601130, 2017.

183.

Lin

X.Y.

, Lai

B.Q.

, Zeng

, et al. Cell transplantation and neuroengineering approach for spinal cord injury treatment: a summary of current laboratory findings and review of literature. Cell Transplant, 25, 1425, 2016.

184.

Han

, Jin

, Xiao

, et al. The promotion of neural regeneration in an extreme rat spinal cord injury model using a collagen scaffold containing a collagen binding neuroprotective protein and an EGFR neutralizing antibody. Biomaterials, 31, 9212, 2010.

185.

Anderson

M.A.

, O'Shea

T.M.

, Burda

J.E.

, et al. Required growth facilitators propel axon regeneration across complete spinal cord injury. Nature, 561, 396, 2018.

186.

Bakshi

, Fisher

, Dagci

, Himes

B.T.

, Fischer

, and Lowman

Mechanically engineered hydrogel scaffolds for axonal growth and angiogenesis after transplantation in spinal cord injury. J Neurosurg Spine, 1, 322, 2004.

187.

Lee

, McKeon

R.J.

, and Bellamkonda

R.V.

Sustained delivery of thermostabilized chABC enhances axonal sprouting and functional recovery after spinal cord injury. Proc Natl Acad Sci U S A, 107, 3340, 2010.

188.

Piantino

, Burdick

J.A.

, Goldberg

, Langer

, and Benowitz

L.I.

An injectable, biodegradable hydrogel for trophic factor delivery enhances axonal rewiring and improves performance after spinal cord injury. Exp Neurol, 201, 359, 2006.

189.

Stokols

, and Tuszynski

M.H.

Freeze-dried agarose scaffolds with uniaxial channels stimulate and guide linear axonal growth following spinal cord injury. Biomaterials, 27, 443, 2006.

190.

Agbay

, Edgar

J.M.

, Robinson

, et al. Biomaterial strategies for delivering stem cells as a treatment for spinal cord injury. Cells Tissues Organs, 202, 42, 2016.

191.

Liu

, Chen

, Liu

, et al. Acellular spinal cord scaffold seeded with mesenchymal stem cells promotes long-distance axon regeneration and functional recovery in spinal cord injured rats. J Neurol Sci, 325, 127, 2013.

192.

Aguado

B.A.

, Mulyasasmita

, Su

, Lampe

K.J.

, and Heilshorn

S.C.

Improving viability of stem cells during syringe needle flow through the design of hydrogel cell carriers. Tissue Eng Part A, 18, 806, 2012.

193.

Ritfeld

G.J.

, Rauck

B.M.

, Novosat

T.L.

, et al. The effect of a polyurethane-based reverse thermal gel on bone marrow stromal cell transplant survival and spinal cord repair. Biomaterials, 35, 1924, 2014.

194.

Williams

R.R.

, Henao

, Pearse

D.D.

, and Bunge

M.B.

Permissive Schwann cell graft/spinal cord interfaces for axon regeneration. Cell Transplant, 24, 115, 2015.

195.

Gomes

E.D.

, Mendes

S.S.

, Leite-Almeida

, et al. Combination of a peptide-modified gellan gum hydrogel with cell therapy in a lumbar spinal cord injury animal model. Biomaterials, 105, 38, 2016.

196.

Dumont

C.M.

, Munsell

M.K.

, Carlson

M.A.

, Cummings

B.J.

, Anderson

A.J.

, and Shea

L.D.

Spinal progenitor-laden bridges support earlier axon regeneration following spinal cord injury. Tissue Eng Part A, 24, 1588, 2018.

197.

Onuma-Ukegawa

, Bhatt

, Hirai

, et al. Bone marrow stromal cells combined with a honeycomb collagen sponge facilitate neurite elongation in vitro and neural restoration in the hemisected rat spinal cord. Cell Transplant, 24, 1283, 2015.

198.

Nagashima

, Miwa

, Soumiya

, et al. Priming with FGF2 stimulates human dental pulp cells to promote axonal regeneration and locomotor function recovery after spinal cord injury. Sci Rep, 7, 13500, 2017.

199.

Stephan

M.T.

, Moon

J.J.

, Um

S.H.

, Bershteyn

, and Irvine

D.J.

Therapeutic cell engineering with surface-conjugated synthetic nanoparticles. Nat Med, 16, 1035, 2010.

200.

Cao

, He

, Wang

, et al. Transplantation of ciliary neurotrophic factor-expressing adult oligodendrocyte precursor cells promotes remyelination and functional recovery after spinal cord injury. J Neurosci, 30, 2989, 2010.

201.

Iyer

N.R.

, Wilems

T.S.

, and Sakiyama-Elbert

S.E.

Stem cells for spinal cord injury: strategies to inform differentiation and transplantation. Biotechnol Bioeng, 114, 245, 2017.

202.

Zhang

, Zeng

, Zhang

, Wang

, Wu

, and Li

Co-transplantation of neural stem cells and NT-3-overexpressing Schwann cells in transected spinal cord. J Neurotrauma, 24, 1863, 2007.

203.

Tang

, Liao

, Shi

, et al. The effects of controlled release of neurotrophin-3 from PCLA scaffolds on the survival and neuronal differentiation of transplanted neural stem cells in a rat spinal cord injury model. PLoS One, 9, e107517, 2014.

204.

Angeli

C.A.

, Edgerton

V.R.

, Gerasimenko

Y.P.

, and Harkema

S.J.

Altering spinal cord excitability enables voluntary movements after chronic complete paralysis in humans. Brain, 137, 1394, 2014.

205.

Calvert

J.S.

, Grahn

P.J.

, Strommen

J.A.

, et al. Electrophysiological guidance of epidural electrode array implantation over the human lumbosacral spinal cord to enable motor function after chronic paralysis. J Neurotrauma, 36, 1451, 2019.

206.

Gad

P.N.

, Kreydin

, Zhong

, Latack

, and Edgerton

V.R.

Non-invasive neuromodulation of spinal cord restores lower urinary tract function after paralysis. Front Neurosci, 12, 432, 2018.

207.

Gill

M.L.

, Grahn

P.J.

, Calvert

J.S.

, et al. Neuromodulation of lumbosacral spinal networks enables independent stepping after complete paraplegia. Nat Med, 24, 1677, 2018.

208.

Chen

, Marsh

B.C.

, Cowan

, et al. Sequential therapy of anti-Nogo-A antibody treatment and treadmill training leads to cumulative improvements after spinal cord injury in rats. Exp Neurol, 292, 135, 2017.

209.

Bilgen

, and Al-Hafez

Comparison of spinal vasculature in mouse and rat: investigations using MR angiography. Neuroanatomy, 5, 12, 2006.

210.

Sekhon

L.H.

, and Fehlings

M.G.

Epidemiology, demographics, and pathophysiology of acute spinal cord injury. Spine (Phila Pa 1976), 26, S2, 2001.

211.

Hilton

B.J.

, Anenberg

, Harrison

T.C.

, Boyd

J.D.

, Murphy

T.H.

, and Tetzlaff

Re-establishment of cortical motor output maps and spontaneous functional recovery via spared dorsolaterally projecting corticospinal neurons after dorsal column spinal cord injury in adult mice. J Neurosci, 36, 4080, 2016.

212.

StemCells

Inc.

former management. Reaction from StemCells, Inc. to two papers in stem cell reports on the efficacy of human NSCs in mouse models of Alzheimer's disease and spinal cord injury. Stem Cell Reports, 8, 194, 2017.

213.

Tazaki

, Tanaka

E.M.

, and Fei

J.F.

Salamander spinal cord regeneration: the ultimate positive control in vertebrate spinal cord regeneration. Dev Biol, 432, 63, 2017.

214.

Mostafalu

, Lenk

, Dokmeci

M.R.

, Ziaie

, Khademhosseini

, and Sonkusale

S.R.

Wireless flexible smart bandage for continuous monitoring of wound oxygenation. IEEE Trans Biomed Circuits Syst, 9, 670, 2015.

215.

Ashammakhi

, Ahadian

, Darabi

M.A.

, et al. Minimally invasive and regenerative therapeutics. Adv Mater, 31, e1804041, 2019.

216.

Ashammakhi

, and Kaarela

Stimuli-responsive biomaterials: next wave. J Craniofac Surg, 28, 1647, 2017.

217.

, Aimetti

A.A.

, Langer

, and Gu

Bioresponsive materials. Nat Rev Mater, 2, 16075, 2017.

218.

Ashammakhi

, Ahadian

, Zengjie

, et al. Advances and future perspectives in 4D bioprinting. Biotechnol J, 13, e1800148, 2018.

219.

Nikkola

, Viitanen

, and Ashammakhi

Temporal control of drug release from biodegradable polymer: multicomponent diclofenac sodium releasing PLGA 80/20 rod. J Biomed Mater Res B Appl Biomater, 89, 518, 2009.

220.

Sivaraman

, Swaminathan

, Moore

, et al. Magnetically-responsive, multifunctional drug delivery nanoparticles for elastic matrix regenerative repair. Acta Biomater, 52, 171, 2017.

221.

Marx

, Walles

, Hoffmann

, et al. “Human-on-a-chip” developments: a translational cutting-edge alternative to systemic safety assessment and efficiency evaluation of substances in laboratory animals and man? Altern Lab Anim, 40, 235, 2012.

222.

Ahadian

, Civitarese

, Bannerman

, et al. Organ-on-a-chip platforms: a convergence of advanced materials, cells, and microscale technologies. Adv Healthc Mater, 7, 1700506, 2018.

223.

Ashammakhi

N.A.

, and Elzagheid

Organ-on-a-chip: new tool for personalized medicine. J Craniofac Surg, 29, 823, 2018.

224.

McLean

I.C.

, Schwerdtfeger

L.A.

, Tobet

S.A.

, and Henry

C.S.

Powering ex vivo tissue models in microfluidic systems. Lab Chip, 18, 1399, 2018.

225.

Wang

, Ma

, Li

, et al. Microfluidic engineering of neural stem cell niches for fate determination. Biomicrofluidics, 11, 014106, 2017.