• Taryn Saggese, Prutha Redey, and Susan R McGlashan.
• Department of Anatomy with Radiology, Faculty of Medical and Health Sciences, University of Auckland, Private Bag 92019, Auckland, 1021, New Zealand.
• Eur Spine J. 2015 Sep 1;24(9):1976-85.

PurposeThe ratio of notochordal (NC) cells to mature nucleus pulposus (MNP) cells in the nucleus pulposus varies with species, age and health. Studies suggest that loss of NC cells is a key component of intervertebral disc degeneration. However, few studies have examined the phenotypes of these two cell populations. Therefore, this study aimed to isolate NC and MNP cells from the same intervertebral disc and study phenotypic differences in extracellular matrix production and cell morphology in 3D culture over 7 days.MethodsSequential mechanical dissociation and enzymatic digestion were used to isolate NC cell clusters and single MNP cells from bovine caudal discs. Cells were cultured in alginate beads and subsequently analysed for viability, cytokeratin-8 expression, GAG production and extracellular matrix gene expression.ResultsMechanical dissociation allowed NC cells to be extracted as intact cell clusters. NC cells represented 8% of the NP cell population and expressed both cytokeratin-8 and vimentin. MNP cells expressed vimentin, only. Both cells types were viable for 7 days. In addition to morphological differences, NC cells produced up to 30 times more total proteoglycan than MNP cells. NC cells had significantly higher aggrecan and brachyury expression.ConclusionsNC and MNP cells can be isolated from the same bovine disc and maintain their distinct phenotypes in 3D culture.

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