Introduction: Spinal regenerative therapies, including cell replacement therapies (CRT), are rapidly gaining traction as viable treatments for acute and chronic spinal cord injury (SCI). Before such a treatments can effectively be translated into clinical practice, large animal data is needed to characterize effective immunosuppression protocols and long-term survival of grafted cells in the potentially inhospitable milieu of the acutely injured spinal cord. In the present study, we characterize the survival and maturation of clinical grade human spinal stem cells (hNPCs) grafted in and around the injury epicenter using a porcine L3 contusion model.

Methods: Isoflurane-anesthetized adult Gottingen-Minnesota minipigs (n=10) underwent 2-level laminectomies (L2-L5) followed by L3 spinal contusion using a 5-mm-diameter circular bar (peak force of 2.5kg at a velocity of 3cm/sec). At 24 hours post-injury, animals received 12 bilateral injections of hNPCs targeted in and around the injury epicenter. After cell grafting, animals were continuously immunosuppressed with tacrolimus (targeted blood level 50-60ng/ml) and mycophenolate mofetil (30mg/kg/day). During recovery, motor and sensory function were periodically monitored for 4 weeks. After survival, the presence of grafted cells was confirmed after staining spinal cord sections with a combination of human-specific (hNUMA, HO14, hNSE, hSYN) or non-specific (DCX, MAP2, CHAT, GFAP, APC) antibodies.

Results: In all cell-grafted animals, hNUMA-positive cells were readily identified. Numerous terminally differentiated grafted neurons with extensive axo-dendritic sprouting were seen; these exhibited hNSE and HO14 immunoreactivity. Similarly, a high density of hSYN-positive terminals derived from grafted neurons and residing in the vicinity of host neurons were also seen. A moderate degree of inflammatory change, as evidenced by the appearance of reactive astrocytes and microglia, was also identified.

Conclusions: These data demonstrate that, using this immunosuppression protocol, cells grafted into the acutely injured spinal cord can survive a minimum of 4 weeks despite the inflammatory, post-traumatic environment.

Patient Care: This study demonstrates that cell replacement therapy (CRT) can be performed within 24hrs post-injury, despite the potentially hostile, inflammatory milieu of the acutely injured cord. This demonstrates that the timing of treatment may be more flexible than previously believed, and hence that acute SCI may be amenable to cell replacement therapy. This brings us closer to realizing stem cell therapy as a viable therapeutic options for patients with acute SCI.

Learning Objectives: By the conclusion of this session, participants should be able to 1) Identify some of the potential barriers to cell replacement therapies (CRT) in patients with acute spinal cord injury (SCI), 2) Appreciate the means by which CRT may benefit SCI patients, 3) Describe some of the goals of future large animal and human trials necessary to bring CRT to fruition as an active therapeutic option.

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