Biomaterials, spinal cord injury, and rehabilitation: A new narrative

Synthetic-based hydrogels

Among synthetic non-biodegradable hydrogel used for SCI repair poly(N-(2-hydroxypropyl)-methacrylamide) (PHPMA) is very promising. The research group of Woerly et al. developed a hydrogel made of PHPMA, obtained by a radical polymerization of the monomer HPMA with the use of a divinyl cross-linking agent ( ), functionalized by a synthetic peptide which includes RGD sequence ( ). The implantation of the hydrogel into the neonatal and adult spinal cord reveals a good infiltration of cells and blood vessels and the following implantation of the hydrogel seeded with rat mesenchymal stem cells (MSCs) in rats results in better Basso, Beattie, Bresnahan (BBB) score than the control group without the implantation ( ). Another investigated polymer in nerve tissue regeneration is the biocompatible and hydrophilic poly(2-hydroxyethyl methacrylate) (PHEMA). As previously said, the charge and the structure of the hydrogel can influence the behavior of cells and the ingrowth of the new tissue. A study by Hejcl et al. ( ) was conducted to compare the different effects of the surface charge and structure of HPMA and HEMA hydrogels on tissue regeneration. Specifically, four different hydrogels were prepared and seeded with rat MSCs: one HPMA-RGD hydrogel by using heterophase separation (HPMA-HS-RGD), which resulted in a structure characterized by microparticles, two HPMA hydrogels by using a solid porogen, one functionalized by RGD peptide (HPMA-SP-RGD), and one without functionalization (HPMA-SP), which resulted in network structures, and the last hydrogel made by positively charged copolymers of HEMA with [2-(methacryloyloxy)ethyl]trimethylammonium chloride (MOETA +). After successful in vitro studies, hydrogels were implanted into rat SCI hemisection model. The best results in terms of in vitro adhesiveness and in vivo survival of MSC was found in the positively charged HEMA-MOETA + hydrogel, whereas the best results in terms of axonal ingrowth and vascularization was found in the HPMA-SP-RGD hydrogel demonstrating the higher efficacy of the network architecture respect to the globular ones. With respect to the influence of the RGD peptide, it increases the vascularization but has no effect the growth of axons.

Hydrogel functionalization with cell-adhesive peptides

The presence of cell-adhesive peptides on hydrogels, such as the laminin-derived peptide sequence SIKVAV [Ser-Ile-Lys-Val-Ala-Val] and fibronectin-derived peptide RGD [Arg-Gly-Asp], can enhance cell adhesion, migration on the scaffold, proliferation, and differentiation ( ). The research group of Kubinova et al. functionalized a copolymer-based hydrogel of HEMA and 2-aminoethyl methacrylate (AEMA) with the laminin-derived Ac-CGGASIKVAVS-OH peptide by disulfide bridges ( ). The functionalization with SIKVAV and RGD ( ) was made on the same type of hydrogel also through the maleimide-thiol coupling reaction. All the functionalized hydrogels guarantee the adhesion and proliferation of rat MSCs maintaining their multi-lineage potential. In addition, in vivo studies results shown a higher connective tissue and vascularization on fibronectin-modified HEMA hydrogel compared to the non-functionalized one ( ). Other molecules such as serotonin can be used as neurotransmitter and can improve neuronal differentiation of implanted or endogenous neuronal progenitor precursors. Despite promising in vitro results, in vivo model studies on implanted PHEMA functionalized with serotonin showed a migration of seeded neural progenitor out from the polymer leading to a fail in proving a long-term effect on nerve tissue reconstruction ( ).

Porosity orientation

The orientation of fibers is important for tissue regeneration because it provides preferential lines along with the cells growth and proliferation. In addition, adequate porosity and mechanical properties have to support the movements of the organ and the regeneration of the new tissue. Hence, in the case of scaffolds for nervous tissue regeneration, the best one has to be characterized by parallel guiding channels and pores. The group of developed SIKVAV-modified PHEMA hydrogels with parallel oriented pores prepared by a salt-leaching method with ammonium oxalate needle-like crystals, and added 8%, 4%, and 0% (wt%) of [2-(methacryloyloxy)ethoxy]acetic acid (MOEAA) obtaining three hydrogels with 57%–77% porosity, pore diameter of ~ 60 mm, and an elastic modulus of 6.7, 27.4, and 45.3 kPa along the pore axis and 2.9, 3.6, and 11 kPa in a perpendicular direction. After 2 months of implantation the results showed that the softest hydrogel collapses because of the thinness of walls causing a sparse axonal growth inside the hydrogel, whereas the stiffest hydrogel supported axonal ingrowth into the pore guides but cyst formed at the tissue-scaffold interface because of difference in mechanical properties between the two components. The best results in terms of axonal ingrowth, presence of blood vessels and Schwann cells are obtained using the hydrogel with the moderate elasticity modulus of 27.4 kPa along the pores. Unfortunately, the use of the moderate scaffold seeded with MSCs was not able to promote a sufficient axonal growth.

Indeed, the rate of axonal growth resulted very slow and after 6 months from the implantation only few axons were be able to cross the hydrogel and infiltrate the caudal stump ( ). Therefore, other factors are necessary in order to promote axonal regeneration. For example, the presence of MSCs overexpressing of an NT-3 receptor ( ) or brain-derived neutrophic factor (BDNF) ( ) on the scaffold can be added in order to enhance axonal growth and recovery of motor functions.

Natural-based hydrogels

This type of hydrogels may be made by ECM derived components such as collagen or hyaluronic acid. Hyaluronic acid (HA) is a natural biocompatible polymer, biodegradable and non-toxic, but it is does not favor the attachment of cells. A possible overcoming solution is represented by the use the hydroxyphenyl derivative of HA which is able to covalently crosslink in situ, forming a hydrogel in presence of horseradish peroxidase enzyme and hydrogen peroxidase ( ). Moreover, the RGD peptide can be linked to the HA-PH derivative ( ) in order to favor the attachment of cells on (HP-HA) hydrogel. Human Wharton’s jelly derived mesenchymal stem cells (hWJ-MSCs) were encapsulated in the hydrogel, which then was injected in the sub-acute spinal cord hemisection. In situ crosslinking had no cytotoxic effect or negative effect on cells. HA-PH-RGD hydrogel was able to favor axonal ingrowth and the presence of hWJ-MSCs increases the effect. However, there were no improvements of motor function probably due to the low quantity of cells incapsulated inside the gel.