Organotypic human spinal cord slice culture as an alternative to direct transplantation of human bone marrow precursor cells for treating spinal cord injury.

Abstract

To determine the possibility of differentiation of human bone marrow-derived mesenchymal precursor cells (BMDMPCs) into neuronal lineage cells using a human spinal cord organotypic slice coculture technique as an alternative to an in vivo human study. Human BMDMPCs were stained with PKH-26 dye before transplantation into 12 human spinal cord slices. In the control group, BMDMPCs were embedded into one spinal cord-free six-well plate containing media. The morphologic differentiation of the transplanted BMDMPCs were observed at 0, 3, 7, and 14 days. Neuroglial differentiation was identified with immunohistochemistry and reverse transcriptase-polymerase chain reaction (RT-PCR). Spherical cells were seen in both groups at day 0. On days 7 and 14, cells developed one or two thick, short processes and typical spindle-shaped cells in the control group and three to five thin, long processes and neuron-like cells in the experimental group. Immunohistochemistry showed double-stained cells with PKH-26 dye (positive) and vimentin (positive), PKH-26 (positive) and neuronal nuclei (NeuN) (positive), and PKH-26 (positive) and glial fibrillary acidic protein (GFAP) (positive) in the experimental group only. RT-PCR showed weak expression of tyrosine kinase A, NeuN, β-tubulin III, and GFAP in the experimental group. Organotypic human spinal cord slice culture may be a useful method to verify the neuroglial differentiation of human BMDMPCs as an alternative to a direct human study.