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Isolation and Characterization of Adult Spinal Disc Stem Cells from Healthy Human Spinal Disc Tissues

Christopher Duntsch, Erika Dillard and Umar Akbar

This report details the isolation, culture, and characterization of spinal disc stem cells derived from human adult spinal disc tissue specimens. Using stem cell suspension culture methods and biology, human adult spinal disc stem cells were isolated and monoclonally cultured into multicellular sphere-like clusters (discospheres). Discospheres from the first culture series were collected, processed, and replated as single stem cells for serial expansion studies using suspension culture, demonstrating linear expansion was possible. Discospheres and adult spinal disc stem cells were plated on matrix coated culture surfaces in stem cell media for several hours to allow fixation, and assayed for the stem cell biomarkers. Discospheres and adult spinal disc stem cells were plated on laminin-coated culture surfaces in chondrogenic media and culture conditions for 14 days to differentiate them into NP cells. NP cells cultured from these experiments demonstrated NP morphology and phenotype; NP biomarker expression, secretion of extracellular matrix, and the ability to be serially passaged with large volume expansion possible. Tissue engineering studies using the “burst kinetic assay”, demonstrated that discospheres have remarkable intrinsic developmental and tissue engineering biology that is robust and organized. In summary, adult disc stem cells and NP cells have been isolated, cultured, and characterized, from healthy spinal disc tissues. These findings demonstrate the important potential to be explored for using stem cell based tissue engineering for the treatment of degenerative disc disease (DDD).