The molecular microenvironment of the injured spinal cord does not support survival and differentiation of either grafted or endogenous NSCs, restricting the effectiveness of the NSC-based cell replacement strategy. Studying the biology of NSCs in in vivo usually requires a considerable amount of time and cost, and the complexity of the in vivo system makes it difficult to identify individual environmental factors. The present study sought to establish the organotypic spinal cord slice culture that closely mimics the in vivo environment. The cultured spinal cord slices preserved the cytoarchitecture consisting of neurons in the gray matter and interspersed glial cells. The majority of focally applied exogenous NSCs survived up to 4 weeks. Pre-exposure of the cultured slices to a hypoxic chamber markedly reduced the survival of seeded NSCs on the slices. Differentiation into mature neurons was severely limited in this co-culture system. Endogenous neural progenitor cells were marked by BrdU incorporation, and applying an inflammatory cytokine IL-1β significantly increased the extent of endogenous neural progenitors with the oligodendrocytic lineage. The present study shows that the organotypic spinal cord slice culture can be properly utilized to study molecular factors from the post-injury microenvironment affecting NSCs in the injured spinal cord.

Keywords: spinal cord injury, organotypic slice culture, neural stem cells, hypoxia, inflammatory cytokine