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

Abstract

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. Significance statement This article describes a novel subpial spinal cell delivery technique that does not require direct spinal tissue needle penetration and is associated with robust spinal cord occupation by subpially injected cells. The effectiveness of this cell delivery technique was validated in long-term studies in immunodeficient rats receiving subpial injection of human neural precursors. This is the first report to demonstrate the successful spinal cell occupation by neural stem cells, which do not require invasive central nervous system or spinal cord cell delivery. Because of the simplicity of this approach, the use of this technique can substantially improve current clinical protocols aimed at spinal delivery of therapeutic cells in the treatment of spinal neurodegenerative disorders.