Spinal parenchymal occupation by neural stem cells after subpial delivery in adult immunodeficient rats

Abstract

Neural precursor cells (NSCs) hold great potential to treat a variety of neurodegenerative diseases and injuries to the spinal cord. However, current delivery techniques require an invasive approach in which an injection needle is advanced into the spinal parenchyma to deliver cells of interest. As such, this approach is associated with an inherent risk of spinal injury, as well as a limited delivery of cells into multiple spinal segments.Here, we characterize the use of a novel cell delivery technique that employs single bolus cell injections into the spinal subpial space. In immunodeficient rats, two subpial injections of human NSCs were performed in the cervical and lumbar spinal cord, respectively. The survival, distribution, and phenotype of transplanted cells were assessed 6-8 months after injection. Immunofluorescence staining and mRNA sequencing analysis demonstrated a near-complete occupation of the spinal cord by injected cells, in which transplanted human NSCs (hNSCs) preferentially acquired glial phenotypes, expressing oligodendrocyte (Olig2, APC) or astrocyte (GFAP) markers. In the outermost layer of the spinal cord, injected hNSCs differentiated into glia limitans-forming astrocytes and expressed human-specific superoxide dismutase and laminin. All animals showed normal neurological function for the duration of the analysis. These data show that the subpial cell delivery technique is highly effective in populating the entire spinal cord with injected NSCs, and has a potential for clinical use in cell replacement therapies for the treatment of ALS, multiple sclerosis, or spinal cord injury. K E Y W O R D S glia limitans formation from grafted neural precursors, human-specific mRNA sequencing, immunodeficient rat, neuraxial neural precursor migration, subpial stem cell injection 1 | BACKGROUND The use of spinally targeted cell-replacement therapies for the treatment of a variety of neurodegenerative disorders, including amyotrophic lateral sclerosis (ALS) and spinal cord injury (SCI), has recently reached the clinical trial stage with several phase I or phase II trials underway.Extensive preclinical studies in rodent and large animal models of ALS and SCI have tested several spinal cell delivery techniques to define the most effective and safe cell delivery approach to be used in human clinical trials.

Looking backward to move forward: a meta-analysis of stem cell therapy in amyotrophic lateral sclerosis

“…For ALS, local delivery targeting the upper motoneurons (intracortical), lower motoneurons (intraspinal), or neuromuscular junction (intramuscular) requires invasive procedures and multiple injections, each representing a risk of AEs, which, in particular, into the CNS parenchyma may have serious consequences, and therefore requires a careful risk–benefit analysis. Equally important, with local approaches the cell distribution is rather restricted to the injected segment or area (for example, with 20 intraspinal injections only about 15% (8 cm) of the spinal cord area was covered) 41 . This has to be taken into account when calculating the cell dose because increasing the cell number without increasing their spread could be detrimental.…”

“…This route makes it feasible to repeat the administration and to perform a sham procedure allowing to evaluate separately the effect of the procedure and that of the SC product (although in this case all trials that used the intrathecal route were done with BM-MSCs so it is not possible to completely separate the effect of both variables on the outcome). A more recently proposed infusion into the subpial space 41 appears promising in preclinical models, by achieving a broad distribution of infused cells, but requires surgery making more problematic to repeat the administration. Given that the effect on the clinical progression appears to be transient, an invasive procedure would not be desirable.…”

Glial-Neuronal Interactions in Pathogenesis and Treatment of Spinal Cord Injury

“…This delivery technique can be used in experimental, pre-clinical and human clinical studies to regulate genes of interest in specific spinal cord segments and/or in the projection of motor and ascending sensory axons. This novel approach is extremely effective in achieving trans-spinal occupation by grafted cells, particularly in the treatment of SCI characterized by multisegmental degeneration [154][155][156].…”

Glial-Neuronal Interactions in Pathogenesis and Treatment of Spinal Cord Injury

“…This novel subpial delivery technique can potentially be used in pre-clinical and human clinical studies to regulate genes of interest in specific spinal cord segments and/or in the projection of motor and ascending sensory axons. This novel approach is extremely effective in achieving trans-spinal occupation by grafted cells, particularly in the treatment of spinal cord injury characterized by multisegmental degeneration [64][65][66].…”