Biomaterial-based strategies: a new era in spinal cord injury treatment

Abstract

Enhancing neurological recovery and improving the prognosis of spinal cord injury have gained research attention recently. Spinal cord injury is associated with a complex molecular and cellular microenvironment. This complexity has prompted researchers to elucidate the underlying pathophysiological mechanisms and changes and to identify effective treatment strategies. Traditional approaches for spinal cord injury repair include surgery, oral or intravenous medications, and administration of neurotrophic factors; however, the efficacy of these approaches remains inconclusive, and serious adverse reactions continue to be a concern. With advancements in tissue engineering and regenerative medicine, emerging strategies for spinal cord injury repair now involve nanoparticle-based nanodelivery systems, scaffolds, and functional recovery techniques that incorporate biomaterials, bioengineering, stem cell, and growth factors as well as three-dimensional bioprinting. Ideal biomaterial scaffolds should not only provide structural support for neuron migration, adhesion, proliferation, and differentiation but also mimic the mechanical properties of natural spinal cord tissue. Additionally, these scaffolds should facilitate axon growth and neurogenesis by offering adjustable topography and a range of physical and biochemical cues. The three-dimensionally interconnected porous structure and appropriate physicochemical properties enabled by three-dimensional biomimetic printing technology can maximize the potential of biomaterials used for treating spinal cord injury. Therefore, correct selection and application of scaffolds, coupled with successful clinical translation, represent promising clinical objectives to enhance the treatment efficacy for and prognosis of spinal cord injury. This review elucidates the key mechanisms underlying the occurrence of spinal cord injury and regeneration post-injury, including neuroinflammation, oxidative stress, axon regeneration, and angiogenesis. This review also briefly discusses the critical role of nanodelivery systems used for repair and regeneration of injured spinal cord, highlighting the influence of nanoparticles and the factors that affect delivery efficiency. Finally, this review highlights tissue engineering strategies and the application of biomaterial scaffolds for the treatment of spinal cord injury. It discusses various types of scaffolds, their integrations with stem cells or growth factors, and approaches for optimization of scaffold design.

Introduction

Spinal cord injury (SCI) is a form of spinal cord damage that can result in a loss of function, sensation, or motion. It can be caused by trauma such as car accidents, sports injuries, or falls or by nontraumatic factors such as infection or disease. SCI can lead to paralysis, muscle weakness, and impaired bowel and bladder control (Easthope et al., 2018; Golestani et al., 2022; Lu et al., 2024). SCI can be categorized as complete and incomplete on the basis of its severity. Moreover, SCI can also be classified as primary or secondary on the basis of the pathophysiological mechanisms. Primary SCI typically refers to mechanical trauma to the spinal cord, and encompasses injuries such as compression, contusion, laceration, or stretching. These injuries directly disrupt the nerve fibers and vascular structures within the spinal cord, leading to malfunction of the blood–spinal cord barrier (BSCB). Secondary SCIs encompass the persistent harm and pathophysiological alterations that occur in the spinal cord subsequent to the initial injury. Thus, secondary SCIs involve an intricate series of cellular and molecular events, including oxidative stress, excitotoxicity, ion dysregulation, lipid peroxidation, inflammation, neuronal apoptosis, demyelination, and scar formation. These processes can result in additional tissue harm and neurological impairment (Squair et al., 2019).

At present, motor nerve axonal regeneration and endogenous nerve bridging are widely acknowledged as viable approaches for treatment of SCIs. Spinal cord function rehabilitation relies on restructuring and integrity of neural circuits. Neuronal circuit dysfunction can occur as a result of neuronal axonal disruption and neuronal death following SCI. The corticospinal tract is a motor axon that frequently undergoes regeneration at injury sites. Rehabilitation of the corticospinal tract may be impaired following severe SCI, limiting the ability to regain motor function in the limbs (Han et al., 2020). Thus, the traditional principles of SCI repair include promoting the regeneration and extension of the corticospinal tract and reconstructing connections with distal neurons (Jo and Perez, 2020). However, due to the large distances, regenerated axons cannot readily connect with the distal neurons. The endogenous neural bridge hypothesis, which proposes that exogenous or endogenous neurons can connect two lesion sites, has gained popularity in recent years. This mechanism helps create new axonal connections and neural circuits, improving motor function after SCI. Primitive spinal cord interneurons, or cells differentiated from transplanted stem cells and indigenous neural stem cells (NSCs), can both become interneurons to facilitate recovery from SCI (Li et al., 2021b).

In the event of SCI, the responsiveness of the local microglia and the destruction of the BSCB create a favorable milieu for blood immune cells to infiltrate the spinal cord tissue, thereby initiating an immune response. Upon activation, these immune cells invade the site of injury and release proinflammatory or immunomodulatory substances to engage in the immune response. The ongoing immune and inflammatory cascade leading to secondary SCI that impedes neural regeneration is considered to be a devastating pathological mechanism influencing the prognosis of SCI. Anti-inflammatory therapies based on this mechanism can inhibit neutrophil infiltration with specific immune depletion or neutrophil baits, which are composed of nanoparticles (NPs) covered by neutrophil membranes, prevent extensive activation and infiltration of immune cells, promote neuronal survival, and thus significantly boost functional recovery after SCI (Chen et al., 2022; Dai et al., 2022a).

In addition, the microenvironment at the SCI site inhibits axon regeneration. Without timely and effective intervention, damaged axons cannot spontaneously extend into the injured area or connect to the intended target. On the other hand, both stem cells and transplanted biomaterial scaffolds play a “seed” role in the reorganization of neural circuits following SCI. Thus, the optimal “soil” or “fertilizer” is essential for creating a favorable regeneration milieu and facilitating the regulation of the cell fate. Currently, a significant body of research has focused on the utilization of biomaterials within the injured regulatory microenvironment. This includes the integration of biomaterials with neurotrophic factors (NFs), stem cells, and NPs. Combining these components can help mitigate the local immune response and promote neural regeneration and repair following SCI through synergistic enhancement of biological effects. Thus, regulation of resident and recruited immune cell activation and regulation of the immunological milieu in SCI holds considerable importance in mitigating secondary injury and facilitating the process of repair and regeneration following SCI (Figure 1).

Figure 1.

Schematic depiction of the implementation of innovative therapeutic strategies grounded in the pathophysiology of SCI.

Early SCI is characterized by M1 microglial infiltration and neuronal apoptosis due to a rapid inflammatory response. After NP injection, the damaged spinal cord shows decreased M1 microglial infiltration and increased formation of M2 microglia, inhibiting neuroinflammation and neuronal apoptosis. Biomaterial scaffolds also limit astrocyte proliferation surrounding the lesion site, preventing glial scarring. High ROS levels cause oxidative stress and inflammation in subacute SCI. The combination of hydrogels combined with NSCs and NP implantation can scavenge ROS, reduce mitochondrial damage, decrease neuroinflammation, and promote neuronal differentiation at the damaged spinal cord. On the other hand, the combination of 3D-printed scaffolds with NSC implantation can fill the wounded site cavity and increase NSC differentiation, neuronal survival, and axonal connection. A composite biomaterial scaffold with NT-3, NPs, and hydrogel can also boost angiogenesis at the injury site in addition to ensuring neuronal survival and M2 microglial infiltration while inhibiting inflammatory cell infiltration. 3D: Three-dimensional; NP: nanoparticle; NSC: neural stem cell; NT-3: neurotrophin-3; ROS: reactive oxygen species; SCI: spinal cord injury.

Conventional approaches for SCI restoration encompass surgical intervention (Ter Wengel et al., 2019; Zhang and Liu, 2022; Wan et al., 2024a) and the administration of oral or intravenous medications (Sandhu et al., 2019; Zhou et al., 2019a; Table 1). Surgical fixation primarily aims to mitigate the compression of the wounded spinal cord by the surrounding tissues, thereby preventing the occurrence of secondary injury (Yao et al., 2020). Methylprednisolone is frequently used to enhance SCI healing by suppressing lipid peroxidation and inflammatory reactions. Nevertheless, the effects of methylprednisolone are not precise, and the potential for severe consequences resulting from high dosage, such as femoral head necrosis and infection, necessitates careful consideration when employing methylprednisolone (Bracken et al., 1990).

Table 1.

Clinical trials of traditional treatment strategies for SCI

Method	Patient condition	Route of administration	Quantity and times	Sample size	Outcome	Phase	Reference
Drug
MPS	Acute SCI	Intravenous bolus and Intravenous infusion	Twice; 30 mg/kg bolus dose and 5.4 mg/kg maintenance dose	160	Administration of the drug within 8 h after SCI facilitated motor and sensory rehabilitation.	Phase I	Bracken et al., 1990
Nimodipine	Acute SCI	Intravenous infusion	0.015 mg/kg/h for 2 h followed by 0.03 mg/kg/h for 7 d	100	The drug did not improve the progression of SCI. Thus, the authors could not prove that this drug treatment provided a benefit.	N/A	Pointillart et al., 2000
Surgery
Posterior decompression + internal fixation	SCI with thoracolumbar spinal fractures	N/A	After admission	40	Stabilized fractures, relieved spinal cord compression, and preserved anatomy with little invasiveness, but failed to relieve anterior spinal cord compression, resulting in a poor prognosis	N/A	Stahel et al., 2013
Real-time anterior decompression + internal fixation	SCI with thoracolumbar spinal fractures	N/A	After admission	40	Made surgery more effective, helped patients restore neurological function, improved quality of life, and reduced complications, but difficult to expose the surgical site and safeguard the segmental blood supply	N/A	Yao et al., 2020

MPS: Methylprednisolone; N/A: not applicable; SCI: spinal cord injury.

Current advancements in SCI repair encompass several approaches, such as the administration of neurotrophic agents, the utilization of biomaterials, bioengineering techniques, three-dimensional (3D) bioprinting, and stem cell research (Table 2). NFs play a crucial role in facilitating neural repair by suppressing neuronal apoptosis following damage and enhancing the regeneration of axons and the differentiation of NSCs. However, NFs are easily degraded in vitro, necessitating repeated administration (Gao et al., 2022a). Due to rapid advancements in tissue engineering, successful therapies for treating SCI have been developed to achieve tissue repair while minimizing the creation of glial scars. With the advent of regenerative medicine, the use of biomaterial implants has become prevalent in the restoration or substitution of impaired tissues by means of bioactive materials that encompass cells and/or growth factors. These implanted scaffolds have the potential to deliver bioactive substances and/or cells to modify the microenvironment, while also serving as stabilizing agents in neural repair. The use of bioactive scaffolds can also reduce the occurrence of secondary damage following SCI. The implantation of these scaffolds serves the dual purpose of safeguarding the lesion and minimizing the development of glial scar tissue. Additionally, these scaffolds provide contact-mediated guidance to facilitate aligned axon regeneration from damaged sites to the distant host tissues. Thus, biomaterial-based approaches exhibit considerable promise and hold significant potential for the prospective therapeutic management of SCI (Figure 2).

Table 2.

Clinical trials of emerging treatment strategies for SCI

Method	Application	Patient condition	Duration of follow-up	Sample size	Phase	Outcome	Reference
Biomaterial	Collagen-hydroxyapatite matrix	Patients with a multilevel anterior cervical discectomy and fusion and/or ACCF	18 mon	60	N/A	The patients’ pain, numbness, and motor function were improved, and successful radiographic fusion indicated that this composite biomaterial could be used as a safe and effective adjuvant material to facilitate multilevel anterior cervical discectomy fusion and ACCF fusion.	Khoueir et al., 2007
Tissue engineering	Collagen scaffold with hUMSCs (NeuroRegen scaffold)	Acute complete SCI patients	1 yr	2	N/A	Significant improvements in motor function and sensory function of the bladder and bowel were observed after implantation, which may be attributable to scaffold-induced differentiation of endogenous NSCs and restoration of interrupted nerve conduction.	NCT02510365
Stem cells	Human spinal-cord-derived NSCs	Chronic SCI patients	27 mon	4	Phase I	Improved sensory and motor function were observed after transplantation, which may be attributed to the better remyelination and/or the formation of fresh synaptic connections with host neurons and downstream motor tracts.	NCT01772810
Growth factors	Human hepatocyte growth factor	Acute SCI patients	168 d	43	Phase I/II	Significantly boosted axon regeneration and motor function recovery were observed, which may be attributable to the ability of HGF to encourage neuron survival, exert neuroprotective effects, and enhance angiogenesis.	NCT02193334

ACCF: Anterior cervical corpectomy and fusion; HGF: hepatocyte growth factor; hUMSC: human umbilical cord mesenchymal stem cell; N/A: not applicable; NSC: neural stem cell; SCI: spinal cord injury.

Figure 2.

Evolutionary timeline of SCI treatment strategies.

The development of treatment strategies for SCI can be roughly divided into four stages: traditional treatment represented by drugs and surgery, cell molecular therapy represented by stem cells and growth factors, nanodelivery systems represented by nanoparticles, and scaffold implantation represented by biomaterials. Traditional treatment is effective but has a poor long-term prognosis. With the advent of cell molecular therapy, neural regeneration has become possible, but it is restricted by the low cell survival and success rates. The subsequent use of nanodelivery systems has brought SCI treatment into the targeted era, but the rapid circulatory clearance of these systems has made them short-lived. In the era of tissue engineering, the excellent physical and chemical properties of biomaterial scaffolds address most of these previous shortcomings. However, more complexity and uncertainty make clinical translation difficult. AT-MSC: Adipose tissue-derived MSC; BDNF: brain-derived neurotrophic factor; BMSC: bone marrow-derived mesenchymal stem cell; ESC: embryonic stem cell; iPSC: induced pluripotent stem cell; MnO₂: manganese dioxide; MSC: mesenchymal stem cell; NGF: nerve growth factor; NP: nanoparticle; NPC: neural progenitor cell; NSC: neural stem cell; NT: neurotrophin; OPC: oligodendrocyte progenitor cell; PCL: ε-polycaprolactone; PDGF: platelet-derived growth factor; PEG: polyethylene glycol; PLA: polylactic acid; PLGA: poly lactic-glycolic acid; UCMSC: umbilical cord mesenchymal stem cell; VEGF: vascular endothelial growth factor.

The primary emphasis of this review pertains to various bioactive scaffolds and functional restoration strategies that are grounded in the fields of diverse biomaterials, bioengineering, stem cell research, and 3D bioprinting. Our primary aim is to elucidate the underlying mechanisms of neural regeneration and determine the optimal design pathway for a future ideal biomaterial scaffold while considering its safety and stability, all of which are prerequisites for successful clinical translation.

Search Strategy

The studies cited in this review were identified using a PubMed search and were mostly published from January 2019 to December 2024. The following MeSH terms were used for the PubMed search: spinal cord injuries, nerve regeneration, nanoparticles, tissue engineering, tissue scaffolds, stem cells, and nerve growth factor. Furthermore, the following title/abstract search words were used: biomaterial scaffold, therapy strategies. The included articles underwent screening based on their titles and abstracts. Articles that did not meet our inclusion criteria were excluded from further analysis.

Pathophysiological Mechanisms and Treatment Strategies

After the occurrence of primary SCI, severe subsequent injury-induced changes occur around the injury site, including persistent oxidative stress, prompt and pronounced inflammatory responses, rapid influx of glutamate and calcium ions into cells, heightened cell permeability, activation of proapoptotic signaling pathways and ischemic injury as a result of microvascular damage. Moreover, glial scars are formed as a result of inflammatory cell infiltration, axonal demyelinating lesions, and the gigantic cystic cavity that forms after SCI. The regeneration of damaged axons is hindered by the cystic cavities and glial scarring, posing a significant challenge to neural healing following SCI (Xue et al., 2024). Additionally, both the axon and vascular networks are disrupted during injury. Notable axonal regeneration at and beyond the injury area in the adult central nervous system (CNS) is usually scarce. This difficulty arises not only from the limited inherent regenerative ability of axons, but also from various external barriers that hinder the process of axonal regeneration. Vascular dysfunction is the primary factor responsible for the unfavorable milieu that results in the demise of neurons and impedes axonal regeneration. Therefore, angiogenesis, which is necessary for restoring circulation and oxygen supply, plays a crucial role in tissue-engineering therapy. Thus, overcoming this hurdle and boosting the angiogenesis are keys in the treatment of SCI.

Neuroinflammation

A rapid and severe inflammatory response and BSCB destruction promote spinal cord swelling and damage in the early stages of injury. Neuroinflammation is mediated by the upregulation of cytokines or chemokines, including tumor necrosis factor (TNF-α), interleukin (IL)-1β, and IL-6, which are usually produced by resident microglia, astrocytes, peripherally derived immune cells, and endothelial cells (ECs). Inflammatory cytokines and chemokines cause the proliferation of immune cells and induce the production of more inflammatory mediators (Kitade et al., 2023). On the other hand, neurons and glial cells show disorders in intracellular calcium regulation, resulting in the activation of calpain. This stimulation causes mitochondrial malfunction and cell death (Li et al., 2022d). Microglia are vital to the inflammatory response. Microglial response is rapid and has an initial protective effect. However, the shift to proinflammatory cells and the release of cytokines initiate a sequence of steps that lead to peripheral immune cell infiltration. The expression of inflammatory cytokines IL-1β and TNF-α has been detected in microglia and astrocytes within 30 minutes of injury, peaking after 1 hour (Poulen et al., 2021).

In the chronic phase, glial and supportive cells form a thick scar surrounding the damage. Glial scar boundaries limit inflammation, oxidative stress, and cellular excitotoxicity at damage sites and protect adjacent tissues. Elongated astrocytes, microglia that release chondroitin sulfate proteoglycans (CSPGs), dense fibroblasts, and extracellular matrix (ECM) form the scar’s physicochemical barrier. Immune cells, macrophages, and neural stem/progenitor cells constitute the rest of the scar. After an injury, cellular signals are sent to the adjacent region to initiate glial scar development, a process that lasts for months. Resident astrocytes and other glial cells enter reactive gliosis in response to these stimuli (Gu et al., 2019). However, although the barrier retains viable neural tissue, infiltrating immune cells can cause inflammation and delay neural regeneration by inhibiting neuronal function and preventing axogenesis by secreting molecules such as CSPGs, Nogo A, and myelin-associated glycoprotein (Maynard et al., 2023).

Gao et al. (2022b) synthesized NPs by combining retinoic acid (RA) and curcumin (Cur) with bovine serum albumin (BSA), and they utilized these composite NPs to treat SCI and examined their anti-inflammatory effects on the SCI repair (Figure 3A). Lipopolysaccharide (LPS)-treated macrophages injected intravenously into the tails of completely transected SCI rats showed the M1 phenotype with high CD86 levels, while RA@BSA@Cur NP-injected macrophages showed the M2 phenotype with high CD206 levels (Figure 3B). After 24 hours of NP interaction, flow cytometry showed a reduction in the number of CD80⁺/F4/80⁺ macrophages and a modest enhancement in the percentage of CD206⁺/F4/80⁺ macrophages, demonstrating the ability to decrease M1 polarization and boost M2 polarization (Figure 3C). Western blotting analyses showed that the LPS-treated group had higher levels of CD86, phosphorylated IκBα (an inhibitor of NF-κB), and phosphorylated p65 than the control group, indicating an inflammatory state. The treatment with NPs dramatically reduced CD86 levels and increased the total IκBα level, with no significant changes in the expression of phosphorylated P65. Thus, RA@BSA@Cur NPs may reduce inflammatory responses by modulating the NF-κB pathway, suppressing M1 macrophages, and enhancing M2 macrophages (Figure 3D). Furthermore, low levels of TNF-α/IL-6 in the LPS-treated group could cause inflammation and worsen tissue and neural damage, while RA@BSA@Cur NPs could raise IL-4/IL-10 levels to boost anti-inflammatory effect, tissue regeneration, and neural repair (Figure 3E).

Figure 3.

RA@BSA@Cur NPs exert anti-inflammatory effects by modulating macrophage phenotypes.

(A) Schematic representation of BSA loaded with RA and Cur forming optimized RA@BSA@Cur NPs and regulating macrophage polarization under the effect of LPS. (B) Immunofluorescence and quantitative analysis results of CD86⁺ M1 and CD206⁺ M2 macrophages in different treatment groups after LPS treatment. CD86⁺ M1 macrophages were marked by yellow arrows, CD206⁺ M2 macrophages by white arrows, and nuclei were shown in blue by DAPI. Scale bar: 50 μm. (C) Results of flow cytometric analysis and quantitative analysis of CD80⁺ M1 and CD206⁺ M2 macrophages cultured in different treatment groups in the presence of LPS. (D) Western blotting results of different treatment groups after treatment with LPS. (E) Quantitative analysis of the levels of inflammatory factors, including IL-6, TNF-α, and IL-4, after RA@BSA@Cur NP treatment. Reprinted from Gao et al. (2022b). *P < 0.05, **P < 0.01, ***P < 0.001. a.u.: Arbitrary unit; BSA: bovine serum albumin; Cur: curcumin; DAPI: 4′,6-diamidino-2-phenylindole; IL: interleukin; LPS: lipopolysaccharide; NP: nanoparticle; ns: not significant; PBS: phosphate-buffered saline; p-IκBα: phosphorylated IκBα; p-P65: phosphorylated p65; RA: retinoic acid; ROS: reactive oxygen species; TNF-α: tumor necrosis factor α.

Oxidative stress

During the subacute phase of SCI, large amounts of reactive oxygen species (ROS) are produced and damage cells in vivo, subsequently leading to oxidative stress (Jiang et al., 2023). Li et al. (2019) demonstrated that low-dose ROS supported the survival of cells, whereas high doses triggered apoptosis, potentially causing malfunction in ECs and the initiation of inflammatory responses. This process could subsequently facilitate the movement and growth of inflammatory cells. Moreover, the increase in antioxidant enzymes resulted in a decline in the production of adhesion molecules and a decrease in the attachment of leukocytes to the endothelium during inflammation (Stewart et al., 2022). In addition, the occurrence of oxidative stress could also result in fragmentation of DNA and damage to mitochondria, stimulating the generation and release of apoptotic factors (Stewart et al., 2021). Thus, oxidative stress may expedite neuroinflammation and neuronal apoptosis, which have a significant impact on secondary damage (Liu et al., 2020b).

Nevertheless, the human body possesses three levels of antioxidant systems to protect itself from such harm. The initial defense mechanism involves the utilization of small-molecule antioxidants, such as uric acid and glutathione, to eliminate ROS and hinder subsequent oxidative harm. When this initial defense is breached, the body initiates a secondary defense mechanism involving antioxidant enzymes such as superoxide dismutase, catalase, and glutathione peroxidase. These enzymes transform ROS into smaller and less harmful molecules. When the second line of defense fails, the harmful effects of different oxides trigger the activation of the third line of defense. This line consists of a range of damage-repair mechanisms and enzymes that can restore injured cells and tissues. For instance, DNA repair enzymes, such as apurinic/apyrimidinic (AP) endonuclease and 8-hydroxyguanine DNA glycosylase, reverse DNA damage resulting from oxidative stress. In summary, antioxidant activity is primarily mediated through this triple line of defense. However, an imbalance between the oxidative and antioxidant defense systems can result in significant oxidative stress, causing harm to cells and tissues.

Cui et al. (2024) developed a bioactive metal ion-based injectable hydrogel by integrating ultrathin and antioxidant MgMn-based layered double hydroxides with silk fibroin (SF). This composite hydrogel could continuously scavenge ROS and release oxygen via Mn³⁺ to alleviate oxidative stress and hypoxia, thereby improving the pathological microenvironment and enhancing SCI repair. Li et al. (2019) constructed a hydrogel by dispersing manganese dioxide (MnO₂) NPs in hyaluronic acid (HA) hydrogel that was modified with a peptide named PPFLMLLKGSTR. They used this hydrogel to explore oxidative stress in a pathological microenvironment and to ameliorate the oxidative microenvironment in injured areas by removing ROS, thereby improving the survival, integration, and differentiation of transplanted stem cells and facilitating nerve tissue growth (Figure 4A). Both in vitro (Figure 4B–D) and in vivo (Figure 4E) experiments proved that the presence of MnO₂ NP-dotted hydrogel contributed to a considerable reduction in the level of ROS in cultured cells in comparison with the control group. Additionally, the cell viability and number of mesenchymal stem cells (MSCs) significantly increased, indicating that the MnO₂ NP-dotted hydrogel effectively protected against oxidative damage and reduced cell death caused by oxidative stress.

Figure 4.

Evaluation of the antioxidant effects of MnO₂ NP-dotted hydrogel in vitro and in vivo.

(A) Schematic diagram of MnO₂ NP-dotted hydrogel prepared with MSCs and implanted into a model of severe long-span spinal cord transection with a lesion gap to resist the oxidative microenvironment. (B) Schematic representation of the protective effect of hydrogels on 3D cultured MSCs in an in vitro oxidative microenvironment. (C) DCFH-DA staining map and quantitative analysis of blank hydrogel and MnO₂ NP-dotted hydrogel after 24 hours of culture. Green, DCFH-DA; blue, DAPI. Scale bars: 100 μm. (D) The viability of MSCs was quantitatively analyzed by live/dead assays after 24 hours of culture. Green, live cells; red, dead cells. Scale bars: 100 μm. (E) DHE staining and fluorescence intensity histogram of peroxide products and staining and percentage of stained area of 4-HNE and the oxidative DNA damage marker 8-OHdG in injured spinal cord tissue 7 days after MnO₂ NP-dotted hydrogel implantation. Scale bars: 100 μm. **P < 0.01, vs. blank group. Reprinted with permission from Li et al. (2019). Copyright 2019 American Chemical Society. 3D: Three-dimensional; 4-HNE: 4-hydroxynonenal; 8-OHdG: 8-hydroxy-2′-deoxyguanosine; DAPI: 4′,6-diamidino-2-phenylindole; DCFH-DA: 2′,7′-dichlorodihydrofluorescein diacetate; DHE: dihydroethidium; MnO₂: manganese dioxide; MSC: mesenchymal stem cell; NP: nanoparticle; ROS: reactive oxygen species.

Axonal regeneration

Neurons undergo axon and dendrite extension to establish connections with other neurons throughout neural development. Axons need to elongate to a greater extent to reach their targets than dendrites. Axonal elongation occurs on the growth cone, which is situated at the distal end of the developing axon and possesses the ability to perceive and interpret external signals to make suitable decisions regarding the growth direction (Giandomenico et al., 2019). The movement of growth cones and the extension of axons are governed by the neuronal cytoskeleton, which consists of microtubules and actin filaments (Sekine et al., 2022). Microtubules serve as the foundational support structures of axons, and their ongoing construction and modification are crucial for controlling the advancement of growth cones and subsequent regeneration of axons. The dynamics of actin in growth cones are crucial for both pathfinding and axon regeneration (Stern et al., 2021). After SCI, the destabilization of microtubules and the disintegration of actin result in the collapse of growth cones and the creation of retraction bulbs (Kong et al., 2024). Axonal regeneration begins with the restoration of growth cone-like structures from severed axon stumps; most interventions that have attempted to enhance axonal regeneration primarily focus on this first phase. Furthermore, the processes of rearranging the cytoskeleton and reorganizing growth cones in injured axons require substantial amounts of energy in the form of adenosine triphosphate, which is primarily generated within mitochondria (Han et al., 2020). SCI-induced mitochondrial dysfunction and energy deprivation worsen the failure of axonal regeneration (Zheng et al., 2024).

Boosting the dynamics of the cytoskeleton in growth cones can promote the growth of axons, whereas disrupted cytoskeletal dynamics following injury are a significant barrier to the regeneration of axons (Sen et al., 2022). The failure of axonal regeneration is mostly attributed to the restricted inherent development capacity of neurons in the adult CNS. Axonal regenerative capacity substantially declines with age and is completely inhibited at maturity (Zheng and Tuszynski, 2023). Another hindrance to axonal regeneration following SCI is the presence of an unfavorable microenvironment (Chen et al., 2023). The absence of exogenous growth inhibitors and NFs additionally impedes neuroplasticity and the regeneration of axons (Ma et al., 2024). Myelin-associated inhibitors and CSPGs are the primary inhibitors that hinder axonal regeneration following SCI, and Nogo A, myelin-associated glycoprotein, and oligodendrocyte myelin glycoprotein are three typical myelin-associated inhibitory molecules (Francos-Quijorna et al., 2022; Schwaiger et al., 2023). NFs are a group of proteins that play crucial roles in promoting the survival of neurons, regulating synaptic function, and facilitating the growth of axons in the mature nervous system (Shen et al., 2019). During maturity, growth factors (GFs) persistently influence several activities of the nervous system, such as the survival of neurons, the generation and release of neurotransmitters, and the adaptability of synapses (Cao et al., 2022). Consequently, the absence of GFs or NFs produced at the proper temporal and spatial gradients remains an obstacle to the regrowth of axons after SCI (Liu et al., 2024a). Thus, to acquire a more profound understanding of the pathological processes that occur after SCI and undertake thorough investigations into the internal and external mechanisms that regulate the growth of axons, innovative tactics that can yield more advantageous outcomes are essential (Shen et al., 2019).

Liu et al. (2024c) synthesized a double-crosslinked conductive hydrogel containing black phosphorus nanoplates, which showed excellent biocompatibility. Moreover, this hydrogel could inhibit the activation of microglia and enhance the anti-inflammatory effect. Injection of the double-crosslinked conductive hydrogel in mice with complete transection SCI in a rotating magnetic field promoted the differentiation of endogenous NSCs into functional neurons and synapses, leading to behavioral and electrophysiological recovery. Liu et al. (2021b) developed a novel biocompatible bioink that consisted of functional chitosan, HA derivatives, and matrix gel, which was used to fabricate NSC-loaded scaffolds through 3D bioprinting at 37°C. They then explored axon regeneration after scaffold implantation and evaluated the underlying mechanisms (Figure 5A). Their in vivo studies demonstrated substantial presence of neurons near the lesion site three months after implantation of the scaffold. Additionally, the regeneration of nerve fibers was notably greater than that in the control group, as shown in Figure 5B. Thus, in vivo implantation of 3D-printed hydroxypropyl chitosan/HA/matrigel scaffolds filled with NSCs could greatly enhance neural and axonal regeneration at the injury site.

Figure 5.

Applications of biomaterial scaffolds to promote axonal regeneration and neuronal differentiation as well as angiogenesis.

(A) Schematic representation of 3D bioprinted neural tissue constructed for SCI repair. (B) DAPI staining and quantitative histogram analysis of Tuj1⁺ and NF⁺ areas in longitudinal sections of the damaged spinal cord at 3 months after composite scaffold implantation. Scale bars: 1 mm. A and B were reprinted from Liu et al. (2021b). (C) Schematic of the preparation process of the BP@TA-LAMC hydrogel and its application in SCI repair. (D) Immunofluorescence images and quantitative histograms of VEGF and CD31 expression in longitudinal spinal cord sections implanted with different conductive hydrogels, as well as Western blotting results of the corresponding labeled proteins. Scale bars: 1 mm (upper) and 100 μm (lower). *P < 0.05, **P < 0.01, ***P < 0.001. C and D were reprinted with permission from Liu et al. (2024b). Copyright 2024 Wiley‐VCH GmbH. 3D: Three-dimensional; BP: black phosphorus; DAPI: 4′,6-diamidino-2-phenylindole; HA: hyaluronic acid; HA-SH: thiolated hyaluronic acid; HA-VS: vinyl sulfonated hyaluronic acid; HBC: hydroxypropyl chitosan; LAMC: lipoic acid-modified chitosan; MA: matrigel; NF: neurotrophic factor; NSC: neural stem cell; SCI: spinal cord injury; TA: tannic acid; VEGF: vascular endothelial growth factor; β-CD-AOI: β-cyclodextrin modified with 2-isocyanatoethyl acrylate.

Revascularization

ECs, basement membranes, pericytes, and the terminal foot processes of astrocytes constitute the BSCB and normally protect the brain parenchyma (He et al., 2023). The junctional complexes among these elements, including tight junctions and adherens junctions, are the main factors determining the permeability of the BSCB (Xie et al., 2023). Recent studies have demonstrated extensive necrosis of ECs and substantial loss of blood vessels at the center of injury within 2–3 days following SCI. Additionally, the junctional complexes mentioned above were destroyed, along with complete mechanical damage to the blood vessels. These factors can contribute to the disruption of barrier integrity, allowing the proliferation of immune cells and neurotoxic substances and resulting in neuronal death and long-lasting neurological dysfunction (Ran et al., 2023; Zuo et al., 2023).

Severe SCI disrupts the integrity of neuronal circuits and causes changes in the microvasculature and insufficient blood flow to the injured area (Zhou et al., 2019b). The availability of nutrients and oxygen in regenerating blood is important for axon regeneration. Furthermore, axonal fibers in the spinal cord often show an orderly arrangement with blood vessels. Consequently, the impairment of vascular integrity after SCI is partially responsible for cellular demise, inflammatory reactions, and inadequate regeneration of axons (You et al., 2023). Microvascular disruption reduces the transport of oxygen, nutrients, and GFs to the injury site, and causes vascular ischemia, hemorrhage, and elevated vascular permeability, leading to an influx of inflammatory cells, which significantly limits spinal cord regeneration (Li et al., 2022a). The vascular bed at the injury site after SCI is disorganized, functionally inefficient, and rapidly opened, leading to neuronal and non-neuronal apoptosis and necrosis (Tran et al., 2020).

Taking into account the environment of ischemia, hypoxia, and limited neural regeneration, which are not suitable for SCI repair, Liu et al. (2024b) explored a novel injectable hydrogel system containing conductive black phosphorus nanoplates coated with lipoic acid-modified chitosan and introduced tannic acid to significantly enhance the conductivity of the hydrogel (Figure 5C). The biodegradable composite hydrogel was implanted into the damaged spinal cord region, and it significantly enhanced the endogenous angiogenic stimulation of the NSC differentiation pathway, indicating that the composite biomaterial significantly promoted angiogenesis and neural differentiation (Figure 5D).

Nanodelivery Systems

In the field of SCI repair and regeneration research, a wide range of natural and composite biomaterials, including NPs, HA, collagen, and poly(lactic-glycolic acid) (PLGA), have been rapidly developed. Nanotechnology has yielded significant advancements in SCI treatment by enabling targeted drug delivery, prolonging drug circulation time, and enhancing bioavailability, thereby providing a new direction for spinal cord regeneration research (Andrabi et al., 2020). Nanotechnology aims to capitalize the unique physical and chemical properties of NPs, such as their nanostructure and large surface areas. It has been successfully applied to drug-delivery systems and tissue engineering, among other applications (Vismara et al., 2020; Lin et al., 2021a; Zhou et al., 2024). A series of dynamic pathophysiological processes occurring after SCI, including vascular leakage and neuroinflammation leading to BSCB dysfunction, result in enhanced permeability and temporary BSCB opening. Nanomaterial-based delivery systems can directly transport to the injury site through improved penetration and retention effects, thus enabling efficient loading and significant improvements in drug, gene, and protein pharmacokinetic profiles. Therefore, intravenous infusion of a single dose of NPs loaded with bioactive molecules could prolong the residence time at the injury site, thereby modulating cellular activity after SCI (Sun et al., 2019; Ciciriello et al., 2022; Jaffer et al., 2023). Furthermore, microenvironment-responsive nanomaterial delivery systems have shown great potential in improving therapeutic efficiency by dispensing bioactive drugs in a controlled manner specific to the microenvironment conditions (Yuan et al., 2023). Additionally, leveraging the electronic properties of nanomaterials along with redox catalytic properties exhibited by inorganic NPs like carbon nanotubes or MnO₂ can stimulate neural repair and alleviate oxidative stress after SCI, thereby avoiding secondary damage and increasing neuronal longevity (Li et al., 2022b; Jaffer et al., 2023; Table 3).

Table 3.

Various types of nanodelivery systems applied for SCI treatment

Type of nano-drug–delivery system		Combination	Delivery method	Injury model	Function & mechanism	Reference
Biodegradable organic materials	Chitosan NPs	Valproic acid-labeled	Intravenous injection	Adult rats with laminectomy and weight-drop hitting	Facilitated tissue regeneration and functional recovery, and reduced the amount of microglia after SCI Significantly improved the amount of Tuj1+ cells in the spinal cord and the expression of NFs after SCI, indicating enhanced differentiation of NSCs after SCI	Wang et al., 2021a
	PLGA NPs	ChABC	Intrathecal injection	Adult rats with laminectomy and weight-drop hitting	ChABC could relieve the inhibition of axon elongation, neuron growth, and CNS plasticity by removing the chondroitin sulfate side chain, thus facilitating the regrowth of nerve fibers, the development of new branches, and the restoration of function following SCI PLGA NPs could protect ChABC from rapid degradation and control their release to overcome the barrier of SCI treatment	Azizi et al., 2020
Inorganic nonmetallic materials	Silica NPs	PMMSN	Intravenous injection	SD rats with SCI	As a sustained-release agent resisted oxidative stress, allowing successful delivery of the drugs to the spinal cord, and resulting in a decrease in cell death and inflammation, and a notable improvement in function recovery Addressed the difficulties in vehicle toxicity, targeting, degradability, and poor solubility of drugs	Jiang et al., 2022
	Hydroxyapatite NPs	Chitosan-hydroxyapatite NPs	Intravenous injection	Adult rats with laminectomy and weight-drop hitting	Significantly boosted the efficiency of targeted delivery to the injured spinal cord, reduced the volume of the injured cavity, and improved the distribution structure, thereby promoting functional recovery and tissue regeneration following SCI	Ma et al., 2021
Metal materials	CeO2 NPs	CeO2 NPs coated on a gelatin-PCL polymer scaffold	Implanted into the spinal cord lesion	Rats with hemisection SCI	Improved motor function and pain relief, which may be attributable to the decreased expression of Iba-1 and GCSF, and increased expression of Tau and MAG	Rahimi et al., 2023
	MnO2 NPs	MnO2 NPs-dotted hydrogel modified with the PPFLMLLKGSTR peptide	Implanted into the lesion gap	Rats with complete transection SCI	Enhanced the adhesion and proliferation of MSCs, and the bridging of neural tissue Modulated the oxidative microenvironment, substantially boosting the survival capacity of MSCs Notably improved motor function and neural regeneration	Li et al., 2019

CeO₂: Cerium oxide; ChABC: chondroitinase ABC; CNS: central nervous system; GCSF: granulocyte colony-stimulating factor; Iba-1: ionized calcium-binding adapter molecule 1; MAG: myelin-associated glycoprotein; MnO₂: manganese dioxide; MSC: mesenchymal stem cell; NF: neurotrophic factor; NP: nanoparticle; NSC: neural stem cell; PCL: ε-polycaprolactone; PLGA: poly lactic-glycolic acid; PMMSN: plasma complex component-functionalized manganese-doped silica NPs; SCI: spinal cord injury; SD: Sprague–Dawley.

Recent studies have investigated the integration of hydrogels with functional NPs to prevent injury and stimulate regrowth. Hydrogels and NPs can function as vehicles for medicines and bioactive substances. Hydrogels fill damaged cavities after SCI, creating a milieu that closely resembles the ECM of the spinal cord to promote the regeneration of neural tissue. On the other hand, NP-based medications can easily enter the BSCB and target the pathophysiological mechanisms underlying SCI. Thus, the integration of hydrogels with NPs holds significant potential for neuroprotection through reduction of inflammation and promotion of neural regeneration (Lin et al., 2021b; Wang et al., 2023b). Neuronal apoptosis and demyelination are frequently observed with secondary SCI. Hydrogels and NPs have been demonstrated to hinder neuronal apoptosis following SCI by regulating signaling pathways. Additionally, they have the potential to stimulate neural regeneration, angiogenesis, and remyelination in the injured area (Wan et al., 2024b). Yin et al. (2024) proposed a multimodal treatment strategy using hydrogels and nanomedicines to promote SCI repair. In their study, cerium oxide (CeO₂) @BSA NPs, an NSC-loaded hydrogel with albumin biomimetic CeO₂ NPs, could scavenge ROS and reduce oxidative stress to promote NSC differentiation and M2 polarization of microglia, thereby facilitating neural regeneration and the recovery of motor function as well as suppressing inflammation at the site of injury and improving the survival of encapsulated NSCs (Figure 6A). Furthermore, plasma complex component-functionalized manganese-doped silica NPs, which exhibit a redox response, could serve as a specialized vehicle for delivering drugs to the reticuloendothelial system. These NPs showed the ability to eliminate ROS and malondialdehyde, while also enhancing the activity of superoxide dismutase and glutathione peroxidase. This led to a reduction in the levels of inflammatory cytokines such as TNF-α, IL-1β, and IL-6, as well as apoptotic cytokines such as cleaved caspase-3. Plasma complex component-functionalized manganese-doped silica NPs have been shown to regulate oxidative stress, inflammation, cell death, as well as the formation of glial scarring after SCI, ultimately promoting the recovery of neurological function (Figure 6B). Similarly, gold NPs injected directly into the injured spinal cord region of SCI rats and locally treated with near-infrared irradiation have been shown to reduce the size of the lesion cavity and decrease the levels of inflammatory cytokines (Figure 6C). Oxidized iron NPs that imitate nanovesicles in mimic exosomes promoted angiogenesis, reduced inflammation, and inhibited cell apoptosis (Figure 6D). Moreover, chitosan-coated hollow MnO₂ nanocarriers significantly mitigated oxidative stress by reducing the levels of ROS, malondialdehyde, and superoxide dismutase while boosting the levels of glutathione peroxidase, thereby improving brain inflammation and apoptosis (Figure 6E).

Figure 6.

Application of hydrogel and inorganic NP-based multimodal treatment strategies to promote SCI repair.

(A) Preparation process and application of the NSC-loaded ROS-scavenging hydrogel containing albumin biomimetic CeO₂ NPs (CeO2@BSA NPs). (B) Schematic representation of PMMSNs with a redox response as a specialized vehicle for delivering drugs to the RES. (C) Schematic of the treatment process of the SCI rat model and the anti-inflammatory mechanism of gold NPs. (D) Preparation and mechanism of action of iron oxide NPs. (E) Design process of chitosan modified hollow MnO₂ (CM) NPs and their application for the efficient delivery of drugs. Reprinted with permission from Yin et al. (2024). Copyright 2023 Wiley‐VCH GmbH. BSA: Bovine serum albumin; Ce: cerium; CeNPs: cerium nanoparticles; CeO₂: cerium oxide; Cl caspase-3: cleaved caspase-3; CM: chitosan-coated hollow manganese dioxide nanocarriers; GelMA: gelatin methacryloyl; GNP: gold nanoparticle; IL: interleukin; iNOS: inducible nitric oxide synthase; MMSN: complex component-functionalized manganese-doped silica nanoparticle; MnO₂: manganese dioxide; NIR: near-infrared; NP: nanoparticle; NSC: neural stem cell; PEG: polyethylene glycol; PMMSN: plasma MMSN; RES: reticuloendothelial system; ROS: reactive oxygen species; SCI: spinal cord injury; TNF-α: tumor necrosis factor α; UDCA: tauroursodeoxycholic acid; UV: ultraviolet.

The efficiency of nanodelivery systems is significantly influenced by the size, shape, and surface charge characteristics of NPs (Pink et al., 2019). Cruz et al. (2016) demonstrated that 100-nm polyethylene glycol (PEG)-polylactic acid-glycolic acid NPs exhibited the deepest penetration and highest accumulation in nerve tissue in injury models in comparison with 200- and 800-nm particles, indicating a negative correlation between NP size and BSCB permeability. Spherical NPs are common due to their rapid internalization, while other shapes such as rods and cubes exhibit superior internalization and drug-loading efficiency (Pink et al., 2019). Zhu et al. (2019) also observed that anisotropic particles were internalized less efficiently than spherical particles, potentially due to reduced clathrin- and reticulin-mediated phagocytosis leading to prolonged retention in vivo. Under static or flow conditions, rod-shaped NPs include more adhesive particles than spherical particles, and they show enhanced internalization in vascular ECs, which might be attributable to their low aspect ratio and ligand density and high surface area, which influence specific area accumulation. In terms of surface charge, Elci et al. (2016) observed that positively charged NPs exhibited prolonged accumulation in the spleen and liver, while neutral particles demonstrated extended circulation and evaded rapid clearance. The electrostatic interactions between particles with positive charge and BSCB EC membranes with negative charge promoted internalization; however, the high positive charge of these NPs may increase the amount of ROS produced, leading to immediate oxidative stress, mitochondrial damage, and disruption of BSCB integrity. Additionally, charge influences the formation of surface protein crests, which subsequently influences receptor-mediated endocytosis. Thus, precise regulation of surface charge could enhance BSCB transport and control cytotoxicity (Monahan et al., 2022).

In summary, NPs can be used as carriers for targeted drug delivery and increased circulation time to improve drug bioavailability, and the use of nanostructures in biomaterial scaffolds can also influence cell fate during SCI repair, making nanotechnology a promising option for spinal cord regeneration.

Scaffolds

Tissue-engineering technology is an advanced technique used to manipulate the growth, differentiation, and organization of cells. It can accurately mimic the immature spinal cord in vivo (Lin et al., 2019). Langer and Vacanti (1993) first posited that tissue engineering involves the use of bioactive chemicals to rebuild or mend organs and tissues via in vitro cultivation or building methods. Regenerative medicine and tissue engineering have led to the exploration of various innovative approaches through multidisciplinary and multifaceted research. These include combining biomaterial scaffolds with stem cells or GFs, using cell scaffolds to mimic growth and support the cellular microenvironment via physical or chemical methods, and fostering collaboration between cells and bioactive molecules (Roh et al., 2023).

SCI often leads to tissue damage and swelling, necessitating structural support to prevent tissue collapse. Electrospun nanofiber scaffolds and hydrogel scaffolds offer physical support for this purpose, facilitating easy implantation and showing great promise as biomaterials (Cnops et al., 2020). Hydrogels consist of either natural or synthetic polymer materials with a hydrophilic network structure. Covalent crosslinked polymer hydrogels are often made from natural materials such HA, silk, chitosan, alginate, and collagen. These hydrogels contain unique cell adhesion molecules and can undergo biodegradation (Liu et al., 2023b). Electrospun nanofiber scaffolds possess an elevated surface area/volume ratio, rendering them suitable for tissue-regeneration purposes. Their exceptional physical and chemical properties have been harnessed in tissue vascular repair applications. Through the adjustment of polymer proportions, hierarchical structural arrangement of electrospinning fibers, and electrospinning conditions, researchers have successfully optimized the mechanical characteristics of these scaffolds to facilitate cell adhesion and growth (Madhavan et al., 2018; Wan et al., 2022). Furthermore, novel heparin-functionalized scaffolds have been developed to address the issue of limited pore size and low porosity associated with electrospun scaffolds. These scaffolds enhance cellular proliferation and infiltration while exhibiting a commendable antithrombotic effect and promoting EC growth (Tan et al., 2016). Ultimately, to optimize the biological impact of biomaterial scaffolds and minimize any negative effects, studies should prioritize the selection of suitable scaffolds for clinical application in patients with SCI, considering their strengths and limitations.

Examples of biodegradable and highly biocompatible natural bioactive materials that promote cell adhesion and proliferation include collagen scaffolds, gelatin scaffolds, ECM-derived scaffolds, and polysaccharide scaffolds (Fan et al., 2022). Collagen is the primary component of the ECM, and collagen scaffolds have attracted significant interest in tissue engineering because of their favorable characteristics such as low immunogenicity, high biocompatibility, permeability, and biodegradability. Moreover, collagen can be fabricated into diverse scaffolds, such as collagen hydrogels, linearly arranged collagen scaffolds, electrospun collagen fibers, aligned collagen sponges, and collagen tubes. Collagen scaffolds can potentially be enhanced by cross-linking reactions and noncovalent interactions to regulate the mechanical qualities and biochemical functions of the scaffolds, thereby enhancing neurogenesis and promoting neurite development (Ghane et al., 2020). Gelatin is derived from collagen. Its abundant arginine-glycine-aspartic acid sequences and integrin binding motifs promote cell adhesion and multiplication by undergoing hydrolytic breakdown (Echave et al., 2017). Gelatin scaffolds, such as gelatin hydrogels, gelatin sponges, and electrospun gelatin fibers, have shown the ability to improve the formation of fresh neurons in a laboratory setting and facilitate the regrowth of neurons in vivo (Yao et al., 2021). Moreover, these scaffolds have been shown to hinder the growth of reactive astrocytes and glial scars while decreasing the inflammatory reaction after implantation in wounded regions (Ma et al., 2020). Decellularized ECM also has the potential to serve as a substrate for neural regeneration due to its ability to maintain the mechanical integrity, bioactivity, and 3D structure of the original tissue matrix (Kim et al., 2020). Decellularized ECM scaffolds have shown beneficial effects on the formation of neurites in cortical and hippocampal neurons, as well as the axonal regeneration and remyelination in damaged areas (Harris et al., 2017). Moreover, these agents can diminish the cavity volume and stimulate macrophage polarization toward the M2 phase while mitigating macrophage invasion (Hong et al., 2020). Matrigel, an extract of ECM consisting of laminin, type IV collagen, entactin, heparan sulfate proteoglycans, and multiple GFs, can efficiently enhance the viability and development of NSCs both in vitro and in vivo (Wang et al., 2020). The linear polysaccharide HA, which consists of d-glucuronic acid and N-acetyl-d-glucosamine disaccharide repeats, plays a vital role in the formation of the ECM. HA scaffolds have the capacity to facilitate the extension of neurites, hinder further harm to both upper and lower neurons, and restrict the extent of the lesion. Consequently, they have been considered as potential substrates for nerve tissue engineering (Li et al., 2022c). Chitosan, another linear polysaccharide, has been demonstrated to reduce fibrous scar formation, promote axonal regeneration and remyelination, and modulate oxidative metabolism and inflammatory responses (Yang et al., 2015). Furthermore, the alginate scaffold can be immediately crosslinked to significantly decrease fibrous scarring while promoting axonal regeneration beyond the site of injury. It also enhances motor recovery and can further boost therapeutic efficiency through biochemical functionalization (Sitoci-Ficici et al., 2018; Liu et al., 2022b). In summary, these natural biomaterials and their derivatives are designed to mimic ECM properties with excellent biocompatibility, biodegradability, and low cytotoxicity; however, the lack of physical rigidity limits their potential for medical applications.

Synthetic scaffolds, including both synthetic peptide scaffolds and synthetic polymer scaffolds, exhibit exceptional mechanical properties and remarkable stability. Synthetic peptide scaffolds, which are typically composed of a hydrophobic tail, a β-sheet forming unit, a charged group, and a bioactive epitope, are synthesized through solid-phase peptide synthesis. Through hydrophobic interactions, these scaffolds may self-assemble, filling cavities in the damaged spinal cord and integrating with host tissues (Holmes et al., 2000). Certain amphipathic molecules, such as immobilized laminin-derived peptide (IKVAV) and binding peptide amphipathic, have been studied for their potential in establishing ideal conditions for regeneration. In addition, they can enable local administration of GFs and stem cells to treat SCI (Tysseling et al., 2010; Iwasaki et al., 2014; Zweckberger et al., 2016; Tran et al., 2020). Ultimately, peptide scaffolds that self-assemble and mimic the ECM, especially when paired with bioactive signals, have the ability to encourage the growth of neurites while also facilitating the attachment, movement, and differentiation of nerve cells and enhancing cell survival. Furthermore, these scaffolds can downregulate inflammatory responses and reduce glial scarring (Luo et al., 2022). In contrast, biocompatible synthetic polymers are frequently used in spinal cord tissue engineering because of their unique degradation design, mechanical qualities, and ease of functionalization. These include biodegradable polymers such as PEG, PLGA, and nondegradable polymers such as poly(2-hydroxyethyl methacrylate), as well as conductive polymers. PEG is a linear polymer that dissolves in water and has the ability to gel in situ or undergo in vivo cross-linking. PEG scaffolds serve several functions, including guiding the development and expansion of axons, promoting the differentiation of neural cells, facilitating myelination, protecting damaged neuronal cell membranes, reducing the formation of glial scar tissue by glial cells, and facilitating the recovery of normal behavior (Luo and Shi, 2007). PLGA is a polymer with adjustable kinetic degradation that has been employed as a scaffold to boost the process of neural differentiation, axon regrowth, and restructuring of the spinal cord by transplanting NSCs, MSCs, and SCs (Han et al., 2019a). Polypyrrole hydrogel scaffolds, which are made of conductive polymers, have the ability to offer electrical stimulation to cells (Zhang et al., 2023). These hydrogels have demonstrated the ability to enhance neural repair and functional restoration. Nevertheless, in comparison with natural biomaterials, further exploration is required to fully understand the biocompatibility of synthetic polymeric biomaterials for improved SCI repair (Table 4).

Table 4.

Natural and synthetic biomaterial scaffolds used for the treatment of SCI

Biomaterial	Application	Injury model	Mechanisms & function	Reference
Natural
Collagen	CBD-LP-miR21-EXO-CoI	Adult rats with complete transection SCI	The combination of Lamp2b and CBD peptide on the surface of exosomes could steadily attach exosomes to the type I collagen scaffold, allow preservation of exosomes loaded with miR21 at the site of injury, and facilitate the ongoing release of miR21 into cells, thereby inhibiting glial scar formation at the site of SCI, reducing cell apoptosis, encouraging neuronal survival, and better exerting neuroprotection	Liu et al., 2022c
Gelatin	3D-GS	Fascicularis monkeys with hemisection SCI	No deterioration of the existing neuroinflammation or astrocyte reaction was observed at the injured site after the implantation of the scaffold, indicating good biocompatibility The number of α-SMA+ cells decreased significantly after implantation, leading to the reduction of fibrotic stress on the remaining spinal cord tissue A significant quantity of cells migrated into the implant and secreted rich ECM, forming a microenvironment that was favorable for the regrowth of nerve fibers, remyelination, vascularization, neurogenesis, and electrophysiology	Zeng et al., 2023
SF	F-SAP/SF	Adult rats with complete transection SCI	The scaffold possessed strong mechanical properties due to the conformational modification of SF by F-SAP, and the controlled release of NT-3 improved the motor and electrophysiological properties by providing a permissive environment for axon regeneration, inflammation regulation, and myelin regeneration	Feng et al., 2023
Chitosan	Chitosan/NT-3	Rhesus monkeys with hemisection SCI	Effective synaptic connections between neurons were found after implantation, and the motor axons in CST not only entered the injured site within the biomaterial but also grew across the injured area and extended to the distal spinal cord, indicating that the scaffold was capable of achieving powerful neural regeneration, along with recovery of motor and sensory function	Rao et al., 2018
Alginate	Alginate saline gels	Adult rats with hemisection SCI	After implantation, the rats were found to show significant motor recovery and reduced fibrous scarring in the spinal cord The scaffold may improve functional recovery after SCI by mechanical stabilization of the wound, reducing secondary injury and inhibiting fibrous scar formation	Sitoci-Ficici et al., 2018
Synthetic
PCL	Aligned electrospun PCL fibers@Nap-E7-YIGSR	SD rats with ischemic SCI	The functional self-assembling peptide demonstrated distinct properties of self-assembly on the surface of PCL fibers and elevated SSC adhesion Effectively differentiated SSC into diverse types of neurons by modulating integrin β1/GSK3β/β-catechin signaling pathway, which possessed functions, and could rebuild neural circuits, participate in nerve electrical signal transmission, and promote the restoration of function	Wang et al., 2022
PLA	PLA/DHA-CSNM	Adult rats with hemisection SCI	Possessed sufficient mechanical properties, and sustainably released DHA to fully exert biological effects Significantly improved neurological function on behavioral assessment after implantation Histological analysis showed effective reduction of neuronal loss and increased sprouting of serotonergic nerves after implantation	Liu et al., 2020c
PLGA	PLGA MS mixed with Laponite hydrogel	SD rats with ischemia SCI	Possessed high loading efficiency and biocompatibility, and facilitated and prolonged melatonin delivery to the injured spinal cord by in situ in vivo injection Enhanced the inhibitory effect of melatonin on macrophage/microglia polarization to the M1 phenotype, thereby preventing the destruction of the biomaterial for better neuroprotection and suppression of oxidative stress and inflammatory responses	Zhang et al., 2021
PEG	Embryonic spinal progenitors@PEG tubes	Adult mice with hemisection SCI	Implantation provided guidance for the injured area of new axons and supported remyelination of these newly formed axons The pro-regenerative effects contributed to the better survival of transplanted stem cells while promoting axonal extension, neurogenesis, and functional recovery	Ciciriello et al., 2020

3D: Three-dimensional; 3D-GS: gelatin sponge with 3D structure; CBD: collagen-binding domain; CBD-LP-miR21-EXO-CoI: collagen-binding domain-fused lysosome-associated membrane glycoprotein 2b-miR21-exosomes-collagen-I; CST: corticospinal tract; DHA: docosahexaenoic acid; DHA-CSNM: docosahexaenoic acid core-shell nanofiber membrane; ECM: extracellular matrix; F-SAP: functional self-assembling peptide; GSK3β: glycogen synthase kinase 3β; Lamp2b: lysosome-associated membrane glycoprotein 2b; MS: microspheres; Nap-E7-YIGSR: Nap-FFGEPLQLKMCDPGYIGSR; NT-3: neurotrophin-3; PCL: ε-polycaprolactone; PEG: polyethylene glycol; PLA: polylactic acid; PLGA: poly(lactic-co-glycolic acid); SCI: spinal cord injury; SD: Sprague–Dawley; SF: silk fibroin; SSC: spermatogonial stem cell; α-SMA: alpha smooth muscle actin.

Despite their multiple advantages, the mechanical strength of biomaterials used as scaffolds is usually unsatisfactory. Moreover, constructing scaffolds using natural materials is challenging due to the lack of suitable methods for achieving optimal conditions (Zhai et al., 2020). Furthermore, the synthetic materials used in biomaterial scaffolds show limitations in clinical applications because of inadequate cell adhesion and affinity, inaccurate degradation rates, dangerous degradation byproducts, and difficulties in engineering properties like porosity and 3D structure (Li et al., 2020). Therefore, the correct selection and application of scaffolds is a promising clinical goal for improving recovery after SCI. With advancements in research based on locally released 3D polymeric scaffolds, these techniques can bypass the BSCB, allowing improved therapeutic efficiency without compromising the biological activity of the molecule (Echave et al., 2017). Furthermore, synthetic polymeric biomaterials are used alongside natural macromolecules by means of chemical cross-linking or chemical modification, thereby improving the characteristics of the transplanted scaffolds. In comparison with single materials, modified engineered structures may perform better on complicated biological systems (Thomas and Shea, 2013; Liu et al., 2020a). Therefore, the modified composite scaffold is gradually becoming a promising biomaterial scaffold in the field of spinal cord tissue engineering because it combines the advantages of synthetic materials, natural materials and nanomaterials, making it an ideal therapeutic strategy for SCI repair (Liu et al., 2023b). However, these techniques are still in their infancy, and their effective and efficient translation to clinical practice will require time (Dai et al., 2022b).

Collagen

Collagen is a predominant and extensively distributed protein within the human body. Each individual pre-collagen is composed of three polypeptide chains that intertwine to create a highly coiled structure resembling a rope. The fibrous structure of collagen facilitates cellular functions such as adhesion, proliferation, and reproduction. Additionally, collagen has demonstrated its suitability as a natural polymeric material for repairing SCI due to low immunogenicity, excellent biocompatibility, and biodegradability, as well as its optimal porosity and mechanical strength (Yang et al., 2021).

In comparison with simple biomaterials, functionalized biomaterials can synergistically enhance the biological effects of each component and compensate for the shortcomings of individual components to maximize benefits. Therefore, functionalized biomaterial scaffolds have been widely used for neural repair after SCI. Using genetic engineering, Suo et al. constructed a collagen-binding domain (CBD)-fused lysosome-associated membrane glycoprotein 2b-miR21-exosomes-type I collagen scaffold (CBD-LP-miR21-EXO-Col) to repair SCI. In their study, miR21 expression after SCI was upregulated in a time-dependent manner to regulate astrocyte hypertrophy and glial scar development (Liu et al., 2022c). In addition, miR21 inhibited its targeted pro-apoptotic genes, including programmed cell death protein 4, FAS ligand, and phosphatase and tensin homolog, thereby reducing apoptosis and exerting neuroprotective effects (Kang et al., 2019). Nevertheless, the development of miRNA-based therapeutics has been impeded by the instability of miRNAs and their limited capacity to enter cells in vivo (Kang et al., 2019). Exosomes are crucial for intercellular or interorgan communication. Exosomes are regarded as optimal vehicles for delivering miRNAs because of their low immunogenicity, inherent stability, and their ability to penetrate tissues and cells (Mu et al., 2021). Additionally, their bilayer membrane structure protects their components from decomposition and facilitates the transfer of these components to recipient cells. However, exosomes show limitations such as off-target effects, limited capability for particular miRNAs, and brief duration in vivo (Ran et al., 2023). Type I collagen scaffolds may occupy the voids formed after SCI, offering a substantial surface area for the concentration and preservation of modified exosomes, as well as sufficient space for cellular proliferation and migration. This, in turn, promotes the interaction between exosomes and cells (Liu et al., 2022c). CBD is a polypeptide composed of the amino acids TKKTLRT derived from collagenase that can attach to biomaterials based on type I collagen. Collagen, as a result of CBD modification, is an optimal medium for transporting medicines to the damaged location of SCI (Han et al., 2019b). Thus, the CBD-LP-miR21-EXO-CoI scaffold shows exceptional efficacy in repairing SCI by allowing the continuous release of miR21 into cells, preventing cell death, enhancing the survival of neurons, and minimizing the formation of glial scars at the SCI site (Liu et al., 2022c).

Zhao et al. (2024) developed a dual-network porous collagen fiber (PCFS) scaffold for neurogenesis using a combination of biomimetic plasma ammonia oxidase catalysis and conventional amide cross-linking. After plasma ammonia oxidase oxidation of the activity of collagen chain aldehyde by a Schiff base formed the first network, the residual functional groups were coupled to construct a dual stable covalent network via 1-ethyl-3-(3′-dimethylaminopropyl) carbodiimide and N-hydroxysuccinimide amidation cross-linking. The rapid prototyping ability and shear-thin nature were used to induce the formation of PCFS using an oriented wet-spinning process equipped with a microinjector pump. In in vitro experiments, the calcium-binding protein receptor of NSCs was activated by PCFS, which induced a series of protein kinase B/Yes-associated protein (AKT/YAP) mechanical signaling pathways, promoting cell orientation and neuron differentiation and adhesion (Figure 7A). In vivo PCFS bundles loaded with CBD-neurotrophin-3 (CBD-NT-3) were transplanted into completely transected SCI rats, resulting in effective neural connections and angiogenesis (Figure 7B–D).

Figure 7.

Application of PCFS scaffolds loaded with CBD-NT-3 to boost functional neuronal growth and axonal regeneration for the treatment of SCI.

(A) Schematic representation of SCI repair using PAO to create a biomimetic collagen scaffold with unique physical properties. (B, C) Immunofluorescence staining maps of Tuj-1⁺ and Map-2⁺ in the injured sites of the individual treatment groups. Scale bars: 1 mm (left) and 50 μm (right and below). (D) Quantitative analysis of the area of Tuj-1⁺ and Map-2⁺ cells at the injury site in the different treatment groups. *P < 0.05, **P < 0.01, ***P < 0.001. Reprinted with permission from Zhao et al. (2024). Copyright 2024 Wiley‐VCH GmbH. CBD-NT-3: Collagen-binding domain-neurotrophin-3; DAPI: 4′,6-diamidino-2-phenylindole; EDC: N-ethyl-N′-(3-(dimethylamino)propyl)carbodiimide; LOCS: linear ordered collagen scaffolds; Map-2: microtubule-associated protein-2; NHS: N-hydroxysuccinimide; NSC: neural stem cell; PAO: plasma amine oxidase; PCFS: porous collagen fibers; SCI: spinal cord injury.

In clinical trials, Xiao et al. (2018) reported a clinical case of collagen scaffold application in acute SCI in which the patients had sustained complete injuries at the thoracic 11 or cervical 4 levels. After transplantation of collagen scaffolds filled with huMSCs to the site of damage, they tracked the progress of these patients. At the 1-year follow-up, they observed recovery of sensory and motor function as well as bowel and bladder function after treatment. Patients with thoracic injury could walk with the aid of braces, and patients with cervical injury could lift their lower legs and shake their toes under gravity. This report ignited research enthusiasm for functional biomaterial scaffolds, indicating the potential effectiveness of these scaffolds for treating patients with SCI. Subsequently, to improve the structure and function of the reconstructed spinal cord after SCI surgery, Chen et al. (2020) cleaned necrotic spinal cord tissue from seven patients with acute complete SCI, implanted the NeuroRegen scaffold in combination with a biogel and autologous bone marrow mononuclear cells into the cleaned site, and followed up the patients for 6 months for rehabilitation and at least 3 years for clinical follow-up. No adverse reactions related to stem cell or functional scaffold implantation were observed during the 3-year follow-up. In addition, some improvements in superficial sensory and autonomic function were observed in some patients, but no recovery in motor function was observed. Postoperative follow-up magnetic resonance imaging showed an improvement in the continuity of the injured spinal cord with no significant cystic cavitation after the NeuroRegen scaffold implantation. Thus, the use of such scaffolds may be a safe clinical treatment approach for acute complete SCI. Moreover, functionalized biomaterials have the ability to rebuild the regenerative milieu and counteract the suppressive effects of myelin proteins on the regrowth of nerves. Consequently, this encouraged the transformation of endogenous or transplanted NSCs into neurons. The newly generated neurons established neural bridges across the damaged region and sent neural impulses, boosting the restoration of neurological function in individuals with transected SCI. These neural bridges generated by neurons derived from either internal or external NSCs are regarded as the primary method for repairing completely transected SCI using functional biomaterials (Yang et al., 2021).

Gelatin is derived from collagen, and gelatin scaffolds have been shown to have excellent ability to promote neural regeneration and inhibit scar formation and neuroinflammation (Echave et al., 2017). Zeng et al. (2023) developed a scaffold made of a gelatin sponge with a 3D structure (3D-GS) to transport cells and NFs for the treatment of SCI. The researchers used a non-human primate SCI model, which demonstrated superior neuroanatomical and functional resemblance to previous animal models. This was done to more accurately assess the effectiveness and safety of this scaffold for repairing the spinal cord. CD68⁺ macrophages and Iba-1⁺ microglia, the number of CSPGs secreted by reactive astrocytes, and the morphological changes in migrating glial fibrillary acidic protein-positive astrocytes were observed by immunohistochemistry to reflect the neuroinflammation and astrocyte response after implantation. The findings demonstrated that the 3D-GS scaffold exhibited favorable neurocompatibility and facilitated both cell migration and nerve fiber development by serving as a bridge. Moreover, the dimensions of the regenerated nerve tissue, which consisted of newly formed neurons, stromal cells, myelinated Schwann cells, and regenerated nerve fibers, along with a substantial amount of pro-regenerative ECM, exhibited an increase following the implantation of 3D-GS scaffolds into the spinal cord of non-human primates with injuries. This indicated that 3D-GS scaffolds have the capacity to function as biomaterials for repairing damaged spinal cords in primates.

Silk fibroin

Fibrin, a prominent constituent of the natural ECM, has been shown to have advantageous effects in facilitating neuronal connections and enhancing the regrowth of axons in both the central and peripheral nervous systems (Tsai et al., 2006). SF is a type of fibrin isolated from the cocoon of silkworm. SF has good biocompatibility and low immunogenicity, and has broad application prospects in biomedicine. Hydrogels, as biomaterials, are capable of imitating the soft tissue environment. They possess an ideal chemical composition to integrate ECM molecules and other binding proteins, thereby facilitating axon regeneration for the healing of SCI in a very efficient manner (Liu et al., 2023c). Feng et al. (2023) reported the use of a hybrid hydrogel prepared from a small functional self-assembling peptide and a large SF. Owing to the conformational transition of SF from a random helix to a β-sheet structure triggered by hydrophobic and hydrogen bonding interactions, the small peptides possessed strong mechanical properties. Besides, the combined controlled release of NT-3 offered a nanofiber substrate for axonal regeneration, inflammation regulation, and myelin regeneration, as well as a permissive environment for neural regeneration, leading to enhanced motor and electrophysiological characteristics. Thus, the combination of functionalized peptides with SF provides a viable approach to boost biological activity for tissue regeneration, and the functional self-assembling peptide/SF hybrid hydrogel could act as a good long-term scaffold for the treatment of SCI (Man et al., 2021).

Polysaccharides

The structure of chitosan is similar to that of the ECM, in which glycosaminoglycans are the major components. Chitosan is nontoxic and inexpensive, can be easily prepared, and yields absorbable degradation products, making it an excellent scaffold material (Wang et al., 2023a). Nevertheless, researchers have identified several limitations of chitosan, including inadequate mechanical strength, poor surface selectivity, and a rate of degradation that is not commensurate with the process of tissue regeneration (Chedly et al., 2017). Zhang et al. (2024b) proposed a hydrogel with immunoregulatory properties made of carboxymethyl chitosan and gallic acid to hinder inflammation and facilitate neural regeneration in acute SCI by combining noncovalent and covalent interactions between the two components. This type of multifunctional hydrogel offers numerous benefits, including biodegradability and biocompatibility, self-healing properties, tissue adhesion-promoting activity, and resistance to oxidation and inflammation. These traits enabled the hydrogel to effectively cover irregular tissue damage or lesions, minimizing invasiveness and interference with surrounding tissues (Figure 8A). In addition, according to the results of the histological analysis of immunofluorescence staining of injured spinal cord tissue in the chronic phase, applying hydrogel composed of chitosan and gallic acid directly to the affected area effectively prevented the infiltration of macrophages and microglia, and promoted the polarization of M2 macrophages. This led to considerable improvements in neural regeneration, reduced scar formation, and ultimately improved neurological function and electrophysiological recovery after SCI (Figure 8B and C). In addition, Rao et al. (2018) established a chitosan scaffold containing NT-3 and implanted it into the thoracic spinal cord of adult rhesus monkeys that had undergone hemisectioning and excision. The effects of this procedure were evaluated by a combination of functional magnetic resonance imaging, electrophysiology, kinematics-based quantitative walking behavior analysis, and immunofluorescence staining of longitudinal spinal cord sections. The findings indicated that NT-3 chitosan scaffolds were effective at promoting robust neural regeneration (Figure 8D–F). Thus, chitosan scaffolds show excellent biological potential for promoting nerve axon connections, repairing damaged tissues, and restoring nerve function. However, the insufficient physicochemical characteristics of chitosan scaffolds limit their application.

Figure 8.

Histological improvement and axonal regeneration of polysaccharide scaffolds.

(A) Schematic diagram of the use of a multifunctional CSGA hydrogel for the treatment of SCI by regulating the inflammatory microenvironment. (B) Immunofluorescence staining and percentages of cells positive for NeuN (representing mature neurons) and GFAP (astrocyte marker) in the chronic SCI model in different treatment groups. Green, NeuN; red, GFAP; blue, DAPI. Scale bars: 500 μm (upper) and 50 μm (lower). (C) Immunofluorescence staining and quantitative analysis histograms showing the percentages of Tuj-1⁺ cells (neuronal microtubule marker) and NF200⁺ cells (neurofilament marker) in the chronic SCI group among the different treatment groups. Green, NF200; red, Tuj-1; blue, DAPI. Scale bars: 500 μm (left) and 50 μm (right). A–C were reprinted from Zhang et al. (2024b). Copyright 2024, with permission from Elsevier B.V. (D) fMRI assessment of thermal sensation mediated by the spinothalamic tract in a rhesus monkey model of right thoracic SCI and functional recovery and BOLD signal changes after NT-3 chitosan treatment. (E) Schematic representation of the unilateral injection of BDA into uninjured and NT-3-chitosan-treated monkeys. (F) Immunofluorescence staining and magnified images of GFAP and BDA in longitudinal segmental spinal cord sections from the different treatment groups at 11 weeks after BDA injection. Blue, DAPI; red, BDA; green, GFAP. White small arrows mark the regenerated BDA-positive fibers. D–F were reprinted from Rao et al. (2018). *P < 0.05, **P < 0.01, ***P < 0.001. BDA: Biotinylated dextran amine; BOLD: blood oxygenation level-dependent; CS: chitosan; CSGA: chitosan and gallic acid; DAPI: 4′,6-diamidino-2-phenylindole; fMRI: functional magnetic resonance imaging; GFAP: glial fibrillary acidic protein; IL: interleukin; NF200: neurofilament-200; ns: not significant; NT-3: neurotrophin-3; ROI: region of interest; SCI: spinal cord injury; TNF-α: tumor necrosis factor α.

Alginate is a polysaccharide extracted from algae. Alginate has the capacity to augment the neural differentiation of stem cells and the expression of neural markers, including MSCs, pluripotent stem cells, and NSCs. Consequently, alginate has become an optimal medium for stem cell treatment. However, the rapid degradation rate of alginate limits its application (Liu et al., 2022a). Cağli et al. (2005) made soft alginate saline gels and injected them into rat SCI models with 2- or 4-mm SCI areas. Behavioral tests and histological examinations showed that the soft alginate saline gel improved motor recovery in rats with 2 mm of SCI. Alginate has the capacity to augment the neural differentiation of stem cells and the manifestation of neural markers, including MSCs, pluripotent stem cells, and NSCs. Consequently, alginate may be an optimal medium for stem cell treatment. Nevertheless, the rapid pace at which alginate deteriorates restricts its potential uses. Sitoci-Ficici et al. (2018) manufactured alginate saline gels with a soft consistency and inserted them into rat models with SCIs measuring either 2 or 4 mm. Behavioral tests and histological examinations demonstrated that the application of soft alginate saline gels resulted in enhanced motor recovery in rats with a 2-mm SCI. Nevertheless, the therapeutic impact was not evident in rats with a 4-mm SCI.

Composite scaffolds

Presently, the primary synthetic components employed in SCl tissue engineering include ε-polycaprolactone (PCL), PLA, PLGA, and PEG. Composite polymer scaffolds with biodegradable properties have been designed to address the dual objectives of facilitating axonal regeneration and preventing further injury to the spinal cord (Zhang et al., 2019).

PCL, an aliphatic polyester, is widely utilized in several medical applications, including tissue-engineering scaffolds, drug/gene carriers, and antiadhesion membranes, due to its biocompatibility and biodegradability. PCL is a bioinert and nontoxic polymer that undergoes degradation over a period of 24 months both in vivo and in vitro, and has the ability to enhance the development of oligodendrocytes and the formation of myelin around axons. Thus, PCL is a useful substance for repairing SCI (Babaloo et al., 2019). It can be formed into different scaffold structures and is typically appropriate for modifying the surface of various biological proteins or peptides (Madhavan et al., 2018). Zhao et al. (2023) created a biomimetic composite hydrogel by mixing acellularized spinal cord matrix with double-crosslinked gelatin acrylated-β-cyclodextrin-polyethylene glycol diacrylate hydrogel. The hydrogel was also infused with WAY-316606 to stimulate the conventional Wnt/β-catenin signaling pathway. In addition, the hydrogel was strengthened by integrating a cluster of 3D-printed aligned PCL microfibers. The acellularized spinal cord matrix was rich in polysaccharides, proteins, and signaling molecules, providing an optimal microenvironment for the rehabilitation of neural tissue and the regulation of NSC cellular behavior. Furthermore, the gelatin acrylated-β-cyclodextrin-polyethylene glycol diacrylate hydrogels possessed the ability to self-heal and included dynamic double-crosslinked networks. These hydrogels were able to preserve excellent structural stability while undergoing degradation, and they also provided a favorable area for the migration of endogenous NSCs and the extension of axons. Moreover, the drug WAY-316606, which has a slow-release mechanism, could stimulate the classical Wnt/β-catenin signaling pathway by suppressing the activity of secreted frizzled-related protein-1. As a result, this drug facilitated the growth and multiplication of neurons (Figure 9A and B). PCL microfibers, which were aligned and had excellent biocompatibility and mechanical stability, connected the two ends of the SCI region. These microfibers acted as a topographic guide, promoting the directed elongation of axons and neurites. Consequently, they reduced cell aggregation and prevented chaotic proliferation of axons (Figure 9C). Thus, this composite hydrogel scaffold effectively filled the void caused by SCI and guided the formation of new neurons. Additionally, it could load and release effective medications to enhance the local microenvironment and facilitate the regeneration of nerve tissue (Figure 9D).

Figure 9.

Application of dual-crosslinked biomimetic composite hydrogels with topographic cues and WAY-316606 to facilitate neural tissue regeneration and functional recovery following SCI.

(A) Immunofluorescence staining and fluorescence intensity histograms of β-catenin and Wnt/β-catenin signaling downstream proteins, including TCF4 and LEF1, during NSC differentiation. Scale bars: the large icons on the left side indicate 50 μm, and the small icons indicate 20 μm. (B) Immunofluorescence staining and fluorescence intensity histograms of β-catenin and LEF1 in the 3D-printed WAY-316606-loaded composite hydrogel scaffold after co-culture with NSCs for 5 days. Scale bars: the large icons on the left side indicate 50 μm, and the small icons indicate 20 μm. (C) Immunofluorescence and quantitative histogram analyses of the expression of IBA1 (inflammatory response marker), HB9 (motor neuron marker), and SYN (synaptic marker) in the central and surrounding areas after SCI. (D) Schematic diagram of the preparation and implantation process of PCL microfiber-reinforced WAY-316606 composite scaffolds. *P < 0.05, **P < 0.01, ***P < 0.001. Reprinted with permission from Zhao et al. (2023). 3D: Three-dimensional; DAPI: 4′,6-diamidino-2-phenylindole; LEF1: lymphoenhancer factor 1; NSC: neural stem cell; PCL: ε-polycaprolactone; PEGDA: polyethylene glycol diacrylate; SCI: spinal cord injury; TCF4: transcription factor 4; UV: ultraviolet; β-CD: β-cyclodextrin.

PLA is a lactic acid polymer that is usually of biogenic origin and has low toxicity. Multiple iterations of PLA have received approval from the U.S. Food and Drug Administration, indicating its potential as a favorable material for medicinal purposes. Furthermore, studies have shown that PLA and its cleavage products are biocompatible with Schwann cells and spinal cord tissue, indicating their potential as vehicles for delivering medicines and biomolecules to repair SCl (Zhang et al., 2024a). Liu et al. (2020c) developed a fiber pad containing a medicine by using electrospinning. The pad was formed of core-shell nanofibers, with a PLA shell around docosahexaenoic acid (DHA) in the core. This design allowed for the direct administration of DHA. The findings indicated that the PLA/DHA core-shell nanofiber membrane could serve as a nanostructured drug-delivery system for DHA. This method had the potential to improve neuroprotection and induce neuroplasticity changes following SCI. Thus, PLA can be considered one of the most valuable and widely studied and reported biodegradable materials due to its excellent low-toxicity, biodegradability, and biocompatibility as well as slow degradation.

PLGA is synthesized via random copolymerization of lactide and glycine. By manipulating the ratio of lactate to glycyrrheic acid, the degradation kinetics of PLGA can be accurately controlled. The ratio also influences the physical and chemical characteristics of PLGA, including bending, permeability, deformation, and swelling. Consequently, PLGA can be synthesized as a flexible material that may serve as a matrix for tissue-engineering scaffolds. PLGA may also serve as a carrier for medications and therapeutic gene delivery (Han et al., 2019a). Zhang et al. (2021) designed and synthesized Lap/MS@Mel, a combination of PLGA MS mixed with Laponite hydrogel, with the aim of achieving high loading efficiency in diverse clinical conditions. They found that the administration of melatonin to the damaged spinal cord could be promoted and prolonged by in situ injection, and that the novel composite hydrogel not only had neuroprotective effects, but also inhibited oxidative stress and inflammation. The neuroprotective benefits of this delivery system have great potential when combined with clinical treatment strategies.

PEG is a polymer that dissolves in water and has the capacity to prevent the release of free radicals, withstand lipid peroxidation, and counteract the increased permeability of cell membranes in tissue-engineering applications (Ciciriello et al., 2020). Shi et al. (1999) injected PEG into guinea pigs that had undergone complete spinal cord transection and observed that PEG scaffolds recapitulated the anatomy of the spinal cord and facilitated the restoration of normal function after injury. Subsequently, the researchers inserted PEG scaffolds into guinea pigs with induced SCl and noticed that both skin and trunk muscle reflexes were restored in the majority of the subjects, with different extents of enhancement. Finally, analysis of evoked potentials demonstrated improved conduction function in all animals, and the PEG scaffolds administered either immediately or 7 hours after damage could effectively decrease cystic cavitation and limit the extent of the lesion. Shi et al. (1999) further proved that the use of PEG resulted in a decrease in caspase-3 activity, leading to a reduction in apoptosis. Additionally, PEG may further strengthen mitochondrial function, thereby decreasing the production of apoptotic factors such as cytochrome c. For this reason, the authors hypothesized that PEG can play a crucial role in the healing process after SCl by preventing apoptosis via its interaction with mitochondria and by repairing broken cell membranes (Luo and Shi, 2007).

Stem cells

Theoretical foundation and benefits of cellular therapy

The pluripotent nature of stem cells enables them to undergo self-replication and differentiation, allowing for their transformation into various types of nerve cells. Through the replacement of damaged cells or the regulation of the microenvironment, stem cells can facilitate axon regeneration, fill cavity lesions, and promote functional recovery. The use of stem cell transplantation has recently gained prominence for the treatment of SCI, and various kinds of stem cells have been employed for this purpose in clinical practice. The transplanted cells have the capacity to undergo differentiation into several cell types, while simultaneously releasing diverse cytokines and GFs that modulate inflammatory responses, offer nutritional assistance, and boost axonal regeneration and neural repair (Curt et al., 2020; Albu et al., 2021; Shang et al., 2022). The combination of cell therapy and biomaterial scaffolds can provide structural support for the transplanted cells, while also creating a conducive microenvironment for cellular infiltration and differentiation. These scaffolds can assist the surrounding neural tissue and serve as substrates promoting cell growth, neurite formation, and axonal regeneration. Additionally, the scaffolds can be loaded with bioactive molecules to establish a relatively stable, permeable, and nutrient-rich regenerative environment. Nevertheless, the biomaterials used for cell culture have different effects on cell adhesion, differentiation, and growth due to differences in charge, hardness, and surface morphology (Guo et al., 2021; Liu et al., 2023a; Mou et al., 2023).

Liu et al. (2023a) developed an NSC-loaded conductive hydrogel scaffold (named ICH/NSCs) consisting of amino-modified gelatin and aniline tetramer grafted oxidized HA, and used this scaffold to fill the damaged cavity (Figure 10A). Six weeks after implantation, the injured area in the group that underwent implantation of ICH/NSCs was lighter in color than the control group, indicating better damage repair and less scar formation. Moreover, immunofluorescence staining of longitudinal sections of the spinal cord revealed a considerably higher number of axons in the ICH/NSC group than in the control group, and the axons had passed through the injury center, with obvious connections between the injured cephalic and caudal nerves. In addition, the larger neurofilament 200-positive area also implied that the NSCs loaded in the hydrogel at the injury site had successfully differentiated into neurons and grown into new nerve axons (Figure 10B). Liu et al. (2023a) designed an extremely permeable DNA supramolecular hydrogel with sufficient and flexible mechanical strength, rapid thixotropic properties, and good biocompatibility. They attempted to establish regenerative neural networks by utilizing endogenous stem cells as well as transplanted stem cells to proliferate and differentiate, which promoted functional recovery (Figure 10C). Two weeks after transplantation, the lesion site and the anterior/caudal host tissues were observed to be full of green fluorescent protein (GFP)-positive cells (implanted NSCs), and many GFP^– cells (host cells) were also observed in the implant material at the injured site, indicating that the implanted material could provide an appropriate cell survival environment (Figure 10D and E). In addition, because DNA hydrogels are pure supramolecular systems containing dynamic networks, they enable bidirectional migration between the implanted GFP⁺ cells and host cells. To prove this point, the researchers labeled the apoptosis rate of GFP⁺ cells by using terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick-end labeling staining and performed quantitative analysis by flow cytometry. Their results demonstrated that the system had excellent mechanical properties and biocompatibility similar to those of the natural ECM, facilitated intercellular communication, and rebuilt the extracellular environment at the lesion site (Figure 10F). Finally, to explore the proliferation and migration of implanted cells, the researchers identified active replicating cells using 5-bromodeoxyuridine labeling and co-visualized them via GFP signaling (Figure 10G) and distinguished NSCs from other cell types by nestin staining to investigate the proliferation and differentiation behaviors of NSCs implanted at the lesion site (Figure 10H; Yuan et al., 2021). These results suggested that the DNA supramolecular hydrogel-NSC system could provide long-term and effective repair and gradually differentiate as implanted cells infiltrate the host tissue, potentially eradicating the obstacle between the renowned tissue and the host and thereby enhancing the restoration of function.

Figure 10.

Application of hydrogels carrying NSCs for SCI repair.

(A) Schematic diagram of the preparation process and effects of ICH scaffolds. (B) Gross observation of the degree of repair and immunofluorescence staining and quantitative histogram of the percentage of the NF200⁺ area in longitudinal sections of the injured spinal cord in individual treatment groups at 6 weeks after scaffold implantation. *P < 0.05, **P < 0.01. A and B were reprinted with permission from Liu et al. (2023a). Copyright 2022 American Chemical Society. (C) Schematic representation of the application of a DNA supramolecular hydrogel carrying homologous NSCs to form a renascent neural network at the lesion site to repair the injured spinal cord cavity. (D) GFP/DAPI immunostaining and local magnified images of sagittal spinal cord sections at 2 weeks after transplantation. White dotted lines show the roughly estimated edge indicating boundaries between the host spinal cord and the lesion area. Scale bars: 1000 μm in D; 50 μm in D1–3. (E) Flow cytometric analysis of GFP⁺ cells in the rostral/caudal host spinal cord of the NM group at 1 and 8 weeks after transplantation. (F) Flow cytometric analysis of the apoptotic rates of GFP⁺ cells immunostained with TUNEL at 3, 7, and 14 days after transplantation. (G) Colocalization images of lesion samples using GFP⁺ and BrdU⁺ immunostaining at 1, 2, and 4 weeks after transplantation and quantitative analysis histograms of the proliferation rate of implanted cells. Scale bar: 20 μm. (H) Immunofluorescence images of NSC migration using GFP/Nestin/DAPI immunolabeling at 1 week after transplantation. Scale bars: 1000 μm in H; 50 μm in H1–H3. C–H were reprinted with permission from Yuan et al. (2021). Copyright 2021 Wiley‐VCH GmbH. AT-OHA: Aniline tetramer grafted oxidized hyaluronic acid; DAPI: 4′,6-diamidino-2-phenylindole; GFP: green fluorescent protein; ICH: an injectable, biodegradable, and self-healing conductive hydrogel; NM: NSC-carrying DNA hydrogel; NSC: neural stem cell; SCI: spinal cord injury; TUNEL: terminal deoxynucleotidyl transferase dUTP nick-end labeling.

Factors influencing cellular behavior

Cells adhere to biomaterial surfaces by a coating of adsorbed ECM proteins such as immunoglobulins, vitronectins, fibrinogen, and fibronectin. These proteins stimulate cell adherence to biomaterials and tissue regeneration (He et al., 2022). Biomaterial surface properties, including charge, functional groups of the surface, hydrophilicity, and topography, impact the nature, amount, conformation, or activity of proteins that are absorbed, which in turn influences cell adhesion, migration, and proliferation (Sun et al., 2019). Zhang et al. (2009) investigated single-wall carbon nanotube surface functional groups, charges, and hydrophilicity. Hydrophilic and positively charged surfaces with -OH groups stimulated cell proliferation more effectively than hydrophobic and negatively charged surfaces with -COO- radical and -NH₂ groups. These factors together showed that electrically neutral and hydrophilic amylose-single-wall carbon nanotube scaffolds with only -OH groups exhibited the greatest cell viability.

The rigidity of the matrix material is a crucial factor regulating cell adhesion and differentiation. Stem cells can sense mechanical cues and translate them into biological signals, generating a cascade of biological reactions that affect function and differentiation (Yao et al., 2016). Woods et al. (2022) created an optimal type IV collagen and fibronectin mix to promote axonal elongation in neuronal cell lines (SHSY-5Y and NSC-34) while triggering the typical morphological traits associated with quiescent astrocytes. The optimum mixture was inserted into an HA scaffold with aligned pores and varied stiffness. Biomimetic scaffolds functionalized with type IV collagen and fibronectin modulated primary astrocyte behavior and induced stiffness-dependent production of IL-10. Seeded SHSY-5Y neurons formed neural networks, while softer biomimetic scaffolds stimulated the development of implanted neurons and axons. This study showed that stiffness and biomaterial composition play crucial roles in cellular responses and that combining physical and matrix cues in a biomimetic manner could greatly enhance neural guiding conduit performance.

The mold method, which yields a regular and uniform shape, has been used to make most scaffolds. However, in contrast to the smooth cuts made in animal models during transection research, the uneven SCI lesions in humans necessitate scaffolds that can adapt to the damaged area without exerting pressure on healthy tissue. Unfortunately, the mold method cannot easily provide the topographical alterations and accurately fabricated microstructures necessary to fulfill these needs (Ju and Dong, 2024). Numerous studies have examined how surface form affects cell growth on material surfaces. Cell behavior can be greatly affected by material roughness. With advancements in manufacturing technology, 3D bioprinting has emerged as an appealing strategy for custom-designed scaffolds. This method allows printing materials with specific sizes and morphologies to effectively mimic the natural spinal cord (Li et al., 2023b). Lesion filling with injectable scaffolds has also been shown to be a viable approach. These liquid-based scaffolds flow into the lesion site with minimal invasiveness and rapidly solidify into a physical support structure (Santi et al., 2021).

Growth factors

Numerous reports have demonstrated that various GFs, including NF (Gao et al., 2022a), platelet-derived growth factor (Wu et al., 2023), brain-derived neurotrophic factor (BDNF) (Liu et al., 2021c), and vascular endothelial growth factor (VEGF) (Xu et al., 2020), exert diverse effects on neural regeneration, inhibition of apoptosis, inflammation suppression, angiogenesis promotion, and glial scar formation inhibition. The synergistic action of GFs at the site of injury holds great promise as a treatment for SCI.

As discussed previously, modulation of neural differentiation and inflammatory responses at the injury site is vital to encourage spinal cord regeneration following stem cell implantation. However, the presence of a niche at the lesion site poses challenges to cell survival, thereby impacting positive outcomes. To address this issue, Gao et al. (2022a) developed a bioactive niche for neural tissue regeneration using aligned SF nanofiber hydrogel scaffolds immobilized with nerve growth factor (NGF). This innovative approach incorporated multiple physical and biological cues to construct a novel bioactive niche for neural tissue regeneration. This approach also promoted neuronal/astrocyte differentiation of embryonic stem cells by regulating the amount of NGF while ensuring excellent biocompatibility of the hydrogel (Figure 11A). Furthermore, along with increased angiogenesis, these bioactive hydrogels facilitated endogenous NSC migration and differentiation, creating an optimal milieu that guided scar-free spinal cord regeneration in vivo. The resulting microstructure of the regenerated spinal cord closely resembled that of normal spinal cord tissue. In animal studies conducted on SCI rats, motor function recovery was observed after implantation of these aligned SF nanofiber hydrogel scaffolds containing NGF, highlighting their potential as promising substrates for scar-free spinal cord regeneration (Figure 11B). Liu et al. (2021c) constructed a scaffold made of collagen and chitosan. They integrated the 3D-printed collagen/chitosan composite scaffold with BDNF by low-temperature extrusion to artificially control its release, retain the biological activity of BDNF, and prolong its release to treat SCI (Figure 11C). In addition, immunofluorescence staining of NF⁺ nerve fibers (Figure 11D) as well as magnetic resonance imaging (MRI) and diffusion tensor imaging (DTI) using different sequences after implantation (Figure 11E) revealed that the presence of BDNF further promoted neural regeneration and reduced the formation of cavities and glial scars at the injured site when the 3D-printed collagen/chitosan composite scaffold with BDNF by low-temperature extrusion scaffold was transplanted to the injured site.

Figure 11.

Implantation of a scaffold loaded with GFs to promote neural regeneration for the treatment of SCI.

(A) Schematic representation of the use of arrayed SFN hydrogels loaded with NGF for long-span spinal cord repair. (B) Immunofluorescence staining of astrocytes and neurons for β-III tubulin and GFAP, as well as quantitative analysis histograms of the total area of the cavity and the percentage of positive cells in regenerated nerve tissue in the injured area at 6 weeks after implantation. A and B are reprinted with permission from Gao et al. (2022a). Copyright 2022 American Chemical Society. (C) Schematic diagram of the preparation and implantation of 3D-CC-BDNF scaffolds by low-temperature extrusion. (D) Immunofluorescence staining and magnified images of NF⁺ nerve fibers in the rostral region, injury/graft site, and caudal region of the spinal cord in different treatment groups at 8 weeks after SCI. Green and white arrow, NF-positive nerve fibers. Scale bars, 500 μm in the left D; 50 μm in the right enlarged images. (E) Representative T2WI, DTT, and DTT-reconstructed images of different treatment groups at 8 weeks after SCI. Red arrows, the injury/graft site located at the T10 spinal cord segment. C–E were reprinted with permission from Liu et al. (2021c). 3D: Three-dimensional; 3D-CC-BDNF: 3D-printed collagen/chitosan composite scaffold with BDNF by low-temperature extrusion; BDNF: brain-derived neurotrophic factor; Dapi: 4′,6-diamidino-2-phenylindole; DTT: diffusion tensor tractography; GFAP: glial fibrillary acidic protein; NF: neurotrophic factor; NGF: nerve growth factor; NSC: neural stem cell; SCI: spinal cord injury; SFN: silk fibroin nanofibers; T2WI: T2-weighted imaging.

The effects of different neurotrophins vary, and the stimulatory effects of biological factors are crucial for axonal regeneration in the CNS. Thus, ensuring consistent and effective transport of factors within a specific microenvironment represents an optimal approach for SCI repair (Xi et al., 2020). The NF family consists of protein molecules produced by innervated tissues and astrocytes, which are essential for neuronal growth. BDNF is important in promoting axon regeneration and neurogenesis, facilitating remyelination, reorganizing synapses, and enhancing synaptic transmission among various neuronal populations following SCI (Crowley et al., 2019). NT-3, a pivotal member of the neurotrophin family, has recently gained extensive attention due to its specific binding to high-affinity tropomyosin receptor kinase C. Studies on NT-3 have demonstrated its ability to enhance neuron differentiation and survival, regulate neuronal synaptic activity, and accelerate axonal regeneration after injury. However, NT-3 is easily diffused and eliminated through tissues and cerebrospinal fluid. Consequently, long-term regulated release of NT-3 after SCI has proven to be difficult (Lai et al., 2016; Li et al., 2021a). Sha et al. (2023) constructed a synthetic biomimetic hydrogel scaffold composed of SF, HA, and polydopamine coatings that was mixed with NT-3 to release NT-3 in a controlled manner at a specific location. The composite hydrogel scaffold exhibited favorable physical and chemical properties, while the polydopamine coating significantly enhanced the loading capacity and release duration of NT-3. In vitro studies investigated the pattern of NT-3 release from the scaffold as well as its biological effects on bone marrow-derived MSC (BMSCs). Subsequently, the scaffold was inserted into rats to assess its effectiveness in encouraging the regrowth of axons and the return of motor function. The in vivo trials demonstrated that implantation of the hydrogel scaffold together with the release of NT-3 could successfully reduce the inflammatory reaction and the development of cavities in the injured cord area, paving the way for neural regeneration.

Microvascular injury, hyperinflammation, and gliosis are other crucial pathophysiological alterations associated with functional conditions following SCI. The sustained release of VEGF into lesion sites has been demonstrated to promote vascular remodeling; however, its impact on reducing inflammation and gliosis remains unknown (Wang et al., 2018). To investigate the synergistic effects of VEGF and NT-3 in promoting angiogenesis, anti-inflammation activity, and neural repair, Xu et al. (2020) utilized an acellular spinal cord scaffold combined with VEGF165, NT-3, and BMSCs to treat a rat model of spinal cord hemisection injury. The findings revealed that VEGF/NT-3-NP-coupled acellular spinal cord scaffold transplantation drastically elevated the levels of VEGF/NT-3 (measured by enzyme-linked immunosorbent assay) and angiogenesis at the site of the injury, regardless of whether VEGF/NT-3-NPs were combined with BMSC transplantation. The findings indicated that the co-delivery of VEGF/NT-3 could mitigate inflammation and gliosis while promoting axon outgrowth after spinal cord hemisection injury, leading to improved motor function recovery. Conversely, the effect of BMSC transplantation alone was limited; furthermore, evidence supporting the differentiation of transplanted BMSCs into neurons or their integration into host tissues was insufficient.

Characteristics and Optimization Directions of Idealized Biomaterial Scaffolds

As mentioned previously, biomaterial scaffolds can facilitate the establishment of a favorable microenvironment for SCI treatment by suppressing local cellular apoptosis or necrosis, diminishing local inflammatory responses and cell excitotoxicity, and inhibiting glial scar formation. The functional scaffold strategy can effectively reconstruct the regenerative microenvironment by bridging the injury gap, significantly enhancing axon regeneration and neurogenesis, and accelerating synaptic connection formation and neural relay development. Consequently, this approach can improve functional recovery and exert beneficial effects on SCI injuries. Thus, to create an effective biomaterial scaffold that can connect the damaged cavities and promote the regrowth of nerve fibers, some key factors, such as maintenance of structural integrity and accurate reproduction of the surrounding milieu as well as capacity for cell and biomolecule delivery, require consideration (Cao et al., 2021).

The molecular and cellular environment following SCI is highly intricate. An optimal biomaterial scaffold should offer structural foundation for the migration, adhesion, maturation, and differentiation of neurons while mimicking the physical characteristics of native spinal cord tissue. This can facilitate axon growth and neurogenesis by providing adjustable topographic, physical, and biochemical cues. To promote effective functional recovery, the spatial arrangement of cells within the biomaterial scaffold as well as the generation of stimulating factors that facilitate the formation of specific neural circuits require attention (Liu et al., 2021a). Additionally, an ideal scaffold should possess excellent biocompatibility and biodegradability with minimal immunogenicity. It should also reduce cavity formation and glial scar formation while showing an appropriate degradation time and yielding non-cytotoxic degradation products without hindering the growth cones of axons or its role as a scaffold for axonal regeneration. Once the repair has been finished, the optimal scaffold components should undergo progressive degradation and be subsequently absorbed by the body (Liu et al., 2023a). Incorporating NGFs and other nutrients necessary for neural regeneration within the scaffold and releasing them at optimal intervals can optimize the delivery of these nutrients and further enhance regeneration processes. Thus, scaffolds with a suitable morphology that can be modified to achieve on-demand release behavior of bioactive molecules can help establish an optimal milieu at and around the injured site (Sha et al., 2023). Moreover, multifunctional scaffolds designed with spatially arranged sequential drug-delivery systems based on heterogeneity, dynamics, and cellular interactions within the SCI microenvironment can specifically regulate and reconstruct this microenvironment, thereby dynamically opening up new perspectives for effective precise spinal cord regeneration (Wang et al., 2021b).

The spinal cord tissue possesses innate softness, which facilitates the efficient transmission of signals between the brain and various sections of the spinal cord. Since the spinal cord is surrounded by cerebrospinal fluid and protected from the bony structure of the spinal canal, development of biologically optimized scaffolds tailored to the characteristics of the spinal cord is a crucial consideration (Bartlett et al., 2020). First, superior mechanical properties and microstructures are essential for promoting SCI repair. Optimal biocompatibility can promote neuron adhesion, while high water content can meet cellular metabolic requirements. Furthermore, good flexibility can enable resistance against deformation under various stresses (Carone and Hasenwinkel, 2006). Second, an ideal scaffold for neural repair should incorporate 3D bioprinting technology with a well-connected porous structure and desirable physicochemical traits such as pore size, rigidity, elasticity, and toughness to effectively enhance SCI treatment using biomaterials. The 3D porous structure can address the limitations faced by hydrogel scaffolds when accurately adapting to irregular or complex lesion sites. With advancements in polymer manufacturing technology, studies on porous structures and alignment matrices will become a prominent area of research in the future (Li et al., 2023a).

The CNS scaffold must closely mimic the native tissue it replaces, while maintaining its integrity and mechanical stability to facilitate axonal regeneration (Koffler et al., 2019). 3D bioprinting technology can mimic the native ECM, providing cell connections that influence cell survival and growth potential. Optimization of scaffold performance can minimize scarring while ensuring an unobstructed space for promoting cellular activity within the scaffold; moreover, an elevated ratio of surface area to volume encourages cell adhesion (Liu et al., 2021b). Koffler et al. (2019) described the fabrication of a 3D-printed biomimetic scaffold using a microscale continuous projection printing method to establish intricate CNS structures for the purpose of regenerating the spinal cord in medical applications (Figure 12A). The scaffold, which was made of PEG-gelatin methacrylate loaded with neural progenitor cells, could be rapidly customized to match rodent spinal cord dimensions in just 1.6 seconds and could also be scaled up to human spinal cord dimensions and lesion geometry (Figure 12B). After 4 weeks of implantation, the channels and solid core of the 3D bionic scaffold maintained their original structure without any fractures or deformations. These scaffolds were easily customized and adaptable to fit any patient-specific lesion shape and length. Moreover, damaged host axons were able to regrow into the 3D bionic scaffolds and form synapses with neural progenitor cells within the implanted device, which then allowed the axons to extend beyond the scaffold and into the spinal cord of the host below the site of injury. This feature ensured restoration of synaptic transmission and yielded significant improvements in functional outcomes (Figure 12C–H). Thus, this study demonstrated the feasibility of utilizing rapid 3D printing technology for creating bionic CNS structures and provided a precision medicine approach to enhance CNS regeneration.

Figure 12.

The application of 3D biomimetic-printed scaffolds loaded with NPC to promote SCI repair.

(A) Schematic diagram of the μCPP layer-free 3D printing process and the hypothesis of axonal arrangement and guidance in the spinal cord. Green, motor systems; blue, sensory systems. (B) The human complete SCI cavity and computer-assisted 3D-printed scaffold models that accurately fitted the shape and size of the injury cavity, as well as the mechanical dynamic analysis curves of the elastic modulus of the scaffold. Red arrow, the lesion site. (C) Immunofluorescence of GFP-expressing NSCs filling scaffold channels with extended linear axons and NF200-labeled axons. (D) Immunofluorescence of 5-HT-labeled host serotonergic axons entering a 3D biomimetic scaffold loaded with NPCs from the rostral side of the lesion (left) and exiting the tail of the channel and regenerating to the host spinal cord distal to the lesion (white arrows). (E) Immunofluorescence of host serotonergic axons regenerated into the scaffold channel versus the dendrites (MAP2-labeled) of NPC-derived (GFP-labeled) neurons (white arrow), and quantification of the mean number of 5-HT axons reaching the caudal area of the scaffold. (F) Ultrastructure of axons of different diameters in the channel (asterisks) and display of myelinated axons (M). (G) Ultrastructural diagram of the process of oligodendrocyte (green) myelination and axon ensheathment (red). (H) Ultrastructure of synapses (arrows) formed between axons within the channel and dendrites of implanted NPCs. Reprinted from Koffler et al. (2019). 3D: Three-dimensional; 5HT: 5-hydroxytryptamine; C: corticospinal tract; DC: dorsal column sensory axons; GFP: green fluorescent protein; MAP2: microtubule-associated protein 2; NF: neurotrophic factor; NPC: neural progenitor cell; NSC: neural stem cell; Pr: propriospinal tract; Ra: raphespinal tract; Ret: reticulospinal tract; Ru: rubrospinal tract; ST: spinothalamic tract; UV: ultraviolet; μCPP: microscale continuous projection printing method.

With the ongoing advancements in scaffold technology, this field of research has now reached the optimization stage. Scaffolds utilized for treating SCI have been meticulously designed to accurately imitate mechanical characteristics of the original spinal cord structure. Future research directions for biomaterial scaffolds may encompass various aspects, including streamlining the manufacturing process of biomaterial scaffolds, identifying more effective comprehensive treatment strategies, and exploring innovative approaches to enhance nervous system regeneration while minimizing secondary injury in SCI (Sousa et al., 2023). However, due to disparities between animal models and human models as well as the limited clinical trials conducted thus far, the clinical translatability of the definitive efficacy observed in rodent models remains inconclusive. Therefore, further studies utilizing large animal models are warranted to bridge this gap and expedite the clinical translation of biomaterial scaffolds (Han et al., 2019b).

Challenges and Perspectives

The pathological mechanisms of SCI exhibit spatiotemporal characteristics that are interconnected and interactive, which limit the scope for a clear explanation. Because the nervous system has limited regenerative ability and shows difficulties in clinical translation, SCI remains a clinically challenging pathophysiological condition with a poor prognosis. A better understanding of the molecular pathological mechanisms that occur after SCI has led to the identification of various intervention strategies, including controlling neuroinflammation and oxidative stress, reducing cell death and scar formation, as well as promoting axonal regeneration and angiogenesis. These strategies have demonstrated safety and efficacy in animal models. Nevertheless, while many interventions have demonstrated encouraging outcomes in animal studies of SCI, such investigations often include highly controlled injury and recovery circumstances that may not completely mirror the natural variations observed in acute SCI cases among humans in relation age, weight, sex, race, or genetic background. Therefore, further optimization of clinical research methods for treating SCI is necessary. Because of the extensive diversity of SCI and notable variations between individuals, researchers can aptly narrow the inclusion criteria and utilize objective biological indicators of injury severity as well as more accurate systems for analyzing SCI prognosis to recruit eligible populations with greater precision.

Nanomaterial-based delivery systems can be employed to provide targeted medicine delivery to the injury site by improving the effects of penetration and durability, thereby prolonging circulation time and improving drug bioavailability. These carriers possess excellent biofilm-transport properties, high biocompatibility, and specific targeting mechanisms. Through structural modification and transformation of the carriers, they can retain the physicochemical properties of nanomaterials while inheriting advantageous functions from the source cells. These advantages will enable effective targeting of diagnostic reagents, small-molecule drugs, nucleic acids, and genes to achieve the desired outcomes in spinal cord regeneration research. Furthermore, in comparison with hydrogels, NPs exert minimal additional pressure on surrounding tissues while exhibiting superior distribution ability and easier contact with the injury site. Researchers have explored the organic combination of hydrogel scaffolds with functional NPs as a promising direction for inhibiting damage and promoting regeneration. In addition, prioritizing ligand selection, carefully considering the physicochemical properties of ligand density and carrier materials, and gaining a comprehensive understanding of the surface interactions between NPs and complex biological media are important considerations for facilitating more rational central targeting design.

Strategies for the use of bioactive scaffolds and the restoration of function in SCI repair have been developed on the basis of biomaterials, bioengineering, stem cell investigations, and 3D bioprinting. Natural biomaterials and their derivatives possess excellent biocompatibility, biodegradability, and low cytotoxicity; nevertheless, the lack of rigidity of these biomaterials and discrepancy between the rate at which natural materials degrade and the rate at which tissue regenerates limit their potential for medical applications. On the other hand, synthetic scaffolds exhibit outstanding mechanical properties and significant stability. Biocompatible synthetic polymers are widely utilized in SCI engineering due to their special pre-degradation design, mechanical properties, and ease of functionalization. Nevertheless, further exploration is required to fully comprehend the biocompatibility of synthetic polymeric biomaterials to enhance SCI repair in comparison with that performed using natural biomaterials. Additionally, the mechanical strength of conventional biomaterials as scaffolds remains unsatisfactory. Scaffolds composed of natural materials pose challenges in achieving an ideal state through appropriate preparation methods, while biomaterial scaffolds made from synthetic materials show limited scope for clinical translation due to the inadequate cell adhesion and connection, ambiguous degradation rates, the generation of harmful degradants, and intricate engineering characteristics including porosity issues and complex 3D structures. Therefore, accurate selection and application of scaffolds represent a promising clinical objective for enhancing recovery after SCI. Synthetic polymeric biomaterials can be combined with natural macromolecules through chemical cross-linking or modification processes to improve the properties of the implanted scaffolds. In comparison with individual materials, these modified engineered structures may exhibit superior performance in complex biological systems.

Stem cell technology is an advanced method for promoting neural regeneration. Combination therapy of stem cells and/or GFs implanted through polymeric scaffolds has gained significant interests. Transplanted cells at the site of SCI can replicate, differentiate, and release different types of cytokines and GFs, modulate inflammatory responses, provide nutrition, boost axonal regeneration and neural repair, and change the injury microenvironment. However, stem cell therapy still shows some challenges, including unsatisfactory efficacy, ambiguous time windows, risk of oncogenicity, immune response, and the cell survival rate. Moreover, the presence of heterogeneity will necessitate more detailed preclinical investigations to evaluate the security and effectiveness. Future therapies should be based on spinal cord pathology and stem cell characteristics to facilitate neural circuit reconstruction or microenvironment repair to fulfill the conditions for a simpler clinical transition. Furthermore, the most suitable cell type for SCI regeneration and repair should be screened, and social, policy, and legal support should be obtained for the use of this cell type. GFs, on the other hand, exhibit diverse regulatory effects on neural regeneration and angiogenesis, neuroinflammation, apoptosis, and glial scar formation; however, a single injection cannot accurately localize the site of nerve injury, and prolonged treatment has not yielded satisfactory outcomes. Therefore, future clinical interventions may require multiple collaborative strategies. For example, synergizing GFs with hydrogel scaffolds at the lesion site to achieve continuous and effective delivery within the local microenvironment represents an optimal strategy for SCI repair.

As mentioned previously, the molecular and cellular microenvironment following SCI is highly intricate. Therefore, the development of combination strategies based on scaffolds that are tailored for specific stages of SCI treatment is essential. Moreover, multifunctional scaffolds designed for spatial and sequential drug-delivery systems possess the ability to precisely modulate and reconstruct the microenvironment in accordance with its heterogeneity, dynamics, and cellular interactions within the SCI microenvironment, allowing efficient and accurate regeneration of the spinal cord. With the continuing optimization of scaffold technology, one of the key concerns regarding the use of scaffolds in SCI repair is their ability to accurately mimic native tissue structure and biological function, in terms of both neuronal and axonal growth processes as well as glial cells and ECM. In addition, a key aspect of CNS scaffolds requires consideration, i.e., its ability to closely mimic the native tissue it replaces, while maintaining mechanical stability that allows for elongation of regenerated axons. Therefore, when designing an effective biomaterial scaffold for connecting injured cavities and stimulating axonal regeneration, the essential parameters should encompass structural stability, simulation of the local microenvironment, and the capacity to deliver cells and bioactive molecules. An optimal biomaterial scaffold should provide a structural foundation for the migration, adhesion, maturation, and differentiation of neurons while mimicking the physical characteristics of native spinal cord tissue to facilitate axon growth and neurogenesis through adjustable topographic, physical and biochemical cues. On the other hand, to enhance SCI treatment using biomaterials effectively, an optimal neural repair scaffold should incorporate 3D bioprinting technology with a 3D interconnected porous structure and appropriate physical and chemical properties. This can address the limitations faced by hydrogel scaffolds in accurately adapting to irregular or complex lesion sites. With advancements in polymer manufacturing technology driving innovative developments, porous structures and alignment matrices will be at the forefront of future research. 3D biomimetic scaffold technology serves two purposes: providing structural foundation for grafts and enabling controlled and localized release of bioactive molecules, thereby significantly advancing the field of tissue engineering and regenerative medicine. However, these technologies are still in their nascent stages, and achieving effective and efficient clinical translation will require considerable time.

The clinical condition provides endless inspiration for scientific research, while clinical application remains the ultimate goal of scientific research; thus, the two aspects complement each other. Looking ahead to the future, biomaterial-based tissue-engineering strategies hold immense potential and a promising outlook for translational research in SCI clinical treatment. However, as mentioned above, biomaterial-based treatment strategies for SCI are still mostly in the laboratory stage, and literature from clinical trials is limited. Consequently, our review primarily covers in vivo and/or in vitro experiments with vague significance for treatment in humans. Successful clinical translation of these findings will require a thorough grasp of the neural regeneration mechanisms, sophisticated biomimetic designs, and meticulous attention to safety and stability considerations. Similarly, future investigations on biomaterial scaffolds should emphasize streamlining the manufacturing process of biomaterials to achieve more efficient and rapid production. Furthermore, exploring innovative strategies to enhance nervous system regeneration while minimizing the secondary damage caused by SCI is imperative to identify superior comprehensive treatment strategies. However, due to the disparities between animal models and humans as well as the insufficient clinical trials conducted thus far, confidence in translating this treatment strategy into clinical practice remains limited. Therefore, further studies using larger primate or non-primate animal models are essential to establish greater clinical relevance for the research findings and expedite the clinical translation of biomaterials. In short, the era develops so rapidly, and novel biomaterials and therapeutic methods are constantly emerging. Although this review refers to almost the original research in the past five years around the world, it is still difficult to cover all the latest research results. In addition, the effect of many biomaterials in the treatment of SCI requires long-term observation to draw definite conclusions, so we look forward to the effect of biomaterials after they go out of the laboratory and are widely used in clinical treatment.

Funding Statement

Funding: This work was supported by the Sichuan Science and Technology Program, No. 2023YFS0164 (to JC); the National Natural Science Foundation of China, No. 82401629 (to XL); the Natural Science Foundation of Sichuan Province, No. 2024NSFSC1646 (to XL); and the China Postdoctoral Science Foundation, Nos. GZC20231811 (to XL) and 2024T170601 (to XL).

Data availability statement:

Not applicable.