An injectable gelatin-based conjugate incorporating EGF promotes tissue repair and functional recovery after spinal cord injury in a rat model

Abstract

Spinal cord injury (SCI) is a devastating condition drastically reducing the quality of life that affects about 300,000 patients in the USA. As a result of the injury, sensory perception and motor functions are lost. Current treatments do not address the root cause - degeneration and loss of neural tissue. The overall goal of this pre-clinical work was to evaluate a novel gelatin-based conjugate (gelatin-hydroxyphenyl propionic acid; Gtn-HPA) capable of undergoing covalent cross-linking in vivo after being injected as a liquid. Gtn-HPA incorporating epidermal growth factor (EGF) and/or stromal cell-derived factor - 1ɑ (SDF-1ɑ) was evaluated for promoting tissue healing and functional recovery using a standardized 2-mm hemi-resection SCI rat model, four weeks after injection. Injection of Gtn-HPA/EGF immediately after the surgical excision injury significantly improved motor functional recovery, compared to gel alone and non-treated controls.

Bladder function was also improved in Gtn-HPA/EGF-treated animals. Functional improvement correlated with the amount of spared tissue. The volume of gel in the defects was quantified by a newly developed MRI-based method employing T1-weighted inversion recovery to unambiguously image Gtn-HPA in the injury site in a non-destructive manner. Histological analysis showed the presence of multiple islands of Gtn-HPA in the injury site after four weeks. There was a significantly greater number of cells migrating into the Gtn-HPA/EGF, compared to the gel alone, and these cells displayed neural progenitor cell markers: nestin, vimentin, and Musashi. The cells infiltrating Gtn- HPA were negative for glial fibrillary acidic protein (GFAP), a marker for astrocytes. Injection of the gel reduced the reactive astrocytic presence at the border outlining the injury site indicating the reduction of glial scar.

There was no notable inflammatory response to the Gtn-HPA gel, reflected in the number of CD68-positive cells, including macrophages. Of note was the demonstration by immunohistochemistry that the Gtn-HPA remaining at 4 weeks post-injection contained EGF. MMP2 was found to be playing a role in in vivo degradation of the Gtn-HPA gel. Additional behavioral and histological results were acquired injecting Gtn-HPA/EGF in 2-mm complete resection SCI rat model. Collectively, the findings signaled that injury sites injected with Gtn-HPA/EGF had greater potential for regeneration. In summary, this work commends an injectable, covalently cross-linkable formulation of Gtn-HPA incorporating EGF for further investigation in promoting functional recovery and potential regeneration for treatment of SCI and thereby improve the quality of life of SCI patients.