Tissue Engineered Spinal Tracts for Reconstruction After Spinal Cord Injury

Introduction: We previously demonstrated the utility of engineered neural tissue using rodent embryonic sensory neurons to re-connect healthy tissue above and below a spinal cord injury (SCI) lesion. The current study characterized the in vitro growth of tissue engineered spinal tracts (TE-STs) built using human iPSC-derived spinal motor neurons and evaluated TE-ST survival and integration in the injured rodent spinal cord.

Methods: Methacrylated hyaluronic acid (MeHA) microcolumns with an inner lumen containing collagen and laminin were fabricated (600µm outer diameter, 300µm inner diameter, 12mm length). Human motor neurons (FUJIFILM) were centrifuged in pyramidal wells to form spherical aggregates that were added to either end of the microcolumn (N=44). Neurite length was measured over 21 days followed by immunocytochemistry for STEM-121 (human marker), CHAT (motor neurons), and TUJ-1 (neurons/axons). Athymic rats underwent T8 balloon compression SCI and were survived for 6 weeks. Groups included no implant (n=5), acellular microcolumns (n=5), and TE-STs (n=10). Outcomes included functional (Basso, Beattie and Bresnahan) and histological (e.g., TE-ST survival, outgrowth, gliosis, cavitation, host regeneration) measures.

Results: Cells stained for STEM-121, CHAT, and TUJ-1, showing human origin with motor neuron phenotype. These human motor neurons extended axons to span the 12mm length of MeHA microcolumns, thereby connecting the neuronal aggregates by 21 days in vitro. Balloon compression SCI elicited severe functional deficits, cavitation, and gliosis. Following longitudinal TE-ST implantation to span the lesion, immunohistochemistry showed TUJ-1+ and STEM-121+ cells in the lumen, indicating survival of transplanted neurons/axons with maintenance of cytoarchitecture, as well as axonal extensions into host spinal cord. Additional analyses are ongoing to contextualize TE-ST survival and outgrowth with functional recovery.

Conclusion : TE-STs were developed featuring long-distance axonal growth from human motor neurons within biocompatible microcolumns. Human TE-STs survived within the injured spinal cord for several weeks with clinical implications for SCI.