Intraspinal Cell Transplantation for Targeting Cervical Ventral Horn in Amyotrophic Lateral Sclerosis and Traumatic Spinal Cord Injury

Abstract

Respiratory compromise due to phrenic motor neuron loss is a debilitating consequence of a large proportion of human traumatic spinal cord injury (SCI) cases 1 and is the ultimate cause of death in patients with the motor neuron disorder, amyotrophic laterals sclerosis (ALS) 2 .ALS is a devastating neurological disorder that is characterized by relatively rapid degeneration of upper and lower motor neurons. Patients ultimately succumb to the disease on average 2-5 years following diagnosis because of respiratory paralysis due to loss of phrenic motor neuron innnervation of the diaphragm 3 . The vast majority of cases are sporadic, while 10% are of the familial form. Approximately twenty percent of familial cases are linked to various point mutations in the Cu/Zn superoxide dismutase 1 (SOD1) gene on chromosome 21 4 . Transgenic mice 4,5 and rats 6 carrying mutant human SOD1 genes (G93A, G37R, G86R, G85R) have been generated, and, despite the existence of other animal models of motor neuron loss, are currently the most highly used models of the disease.Spinal cord injury (SCI) is a heterogeneous set of conditions resulting from physical trauma to the spinal cord, with functional outcome varying according to the type, location and severity of the injury 7 . Nevertheless, approximately half of human SCI cases affect cervical regions, resulting in debilitating respiratory dysfunction due to phrenic motor neuron loss and injury to descending bulbospinal respiratory axons 1 . A number of animal models of SCI have been developed, with the most commonly used and clinically-relevant being the contusion 8 .Transplantation of various classes of neural precursor cells (NPCs) is a promising therapeutic strategy for treatment of traumatic CNS injuries and neurodegeneration, including ALS and SCI, because of the ability to replace lost or dysfunctional CNS cell types, provide neuroprotection, and deliver gene factors of interest 9 .Animal models of both ALS and SCI can model many clinically-relevant aspects of these diseases, including phrenic motor neuron loss and consequent respiratory compromise 10,11 . In order to evaluate the efficacy of NPC-based strategies on respiratory function in these animal models of ALS and SCI, cellular interventions must be specifically directed to regions containing therapeutically relevant targets such as phrenic motor neurons. We provide a detailed protocol for multi-segmental, intraspinal transplantation of NPCs into the cervical spinal cord ventral gray matter of neurodegenerative models such as SOD1 G93A mice and rats, as well as spinal cord injured rats and mice 11 . Video LinkThe video component of this article can be found at

Degeneration of Phrenic Motor Neurons Induces Long-Term Diaphragm Deficits following Mid-Cervical Spinal Contusion in Mice

“…Several contusion models at mid-cervical levels (C3, C4, C5) that show chronic abnormalities in respiratory parameters have been developed in the rat. [22][23][24][25] The extension of a contusion injury model to the mouse provides opportunities for manipulating genes of interest and for evaluating their role in relevant outcome measures affected by cervical spinal contusion such as forelimb motor function, respiratory function, and neuropathic pain.…”

“…The dorsal skin and underlying muscle layers were incised along the midline between the spinous processes of C2 and T1 to expose the cervical region of the spinal cord. 24 The dorsal muscle layers were retracted, and the paravertebral muscles overlying C4-C6 were removed. Following unilateral laminectomy on the right side at C4 and C5 levels, mice were subjected to a double spinal contusion injury at both C4 and C5 levels using the Infinite Horizon (IH) impactor (Precision C5 double contusion 3 3 6 3 6 9…”

Umbilical cord-derived mesenchymal stem cell therapy for neurological disorders via inhibition of mitogen-activated protein kinase pathway-mediated apoptosis

“…Several studies have demonstrated that the transplantation of UC-MSCs is a potential therapeutic strategy for the treatment of central nervous system diseases, including cerebrovascular disease, nerve degenerative diseases, spinal cord injury and cerebral palsy. The transplantation path in cell therapy is diverse (14)(15)(16) and mainly includes the intervention path, local implantation, intravenous route and lumbar puncture way. Migration toward pathology is the first critical step in UC-MSCs engagement during regeneration and it is hypothesized that the…”

Immunohistochemical Toolkit for Tracking and Quantifying Xenotransplanted Human Stem Cells

“…The dorsal skin and underlying muscle layers were incised along the midline between the spinous processes of C2 and T1 [25–27]. Following right unilateral laminectomy, mice were administered 100,000 GRP in 2 μl in the ventral horns of both C4 and C5 levels, at a depth of 0.75 mm.…”