Mechanobiology of the nucleus pulposus: The effect of hydrostatic pressure and glucose deprivation on nucleus pulposus cell sub‐populations

The nucleus pulposus of the intervertebral disc contains two cell types: notochordal (NC) and chondrocyte‐like mature nucleus pulposus (MNP) cells; the ratio of which changes with species, age and health. Intervertebral disc (IVDs) that contain predominantly MNP cells are susceptible to degeneration, while NC cell–rich discs remain healthy. Loss of NC cells is hypothesized to be initiated by mechanical stress and/or nutrient deprivation. This study aimed to investigate the phenotypic and functional differences between NC and MNP cells in response to a range of hydrostatic pressures and glucose concentrations. In this study a novel isolation method was developed that isolated single MNP cells and NC cell‐clusters from the same nucleus pulposus. Each cell type was cultured in 3D alginate beads, and assessed for phenotypic markers and extracellular matrix (ECM) expression on days 1 and 7 of culture. A custom‐built hydrostatic pressure chamber was used to expose each cell type to low/ physiological (0.6 MPa) and high/ pathological (2 MPa) pressure for 24 hours. To study the effect of nutrient deprivation, each cell type was cultured in either normal (5.5 mM) glucose media or glucose‐ deprived (0.55 mM) media. Finally, the combined effect of glucose deprivation and high pressure was examined. Following isolation, NC cells remained in cell clusters and expressed NC markers cytokeratin‐8 and brachyury. NC cells expressed significantly more aggrecan and GAG than MNP cells over the 7 day culture period. MNP cells responded to low pressure with an increase in total GAG while high pressure resulted in a significant decrease in aggrecan gene expression. NC cells showed no response in matrix gene expression or production to either loading regime. Glucose deprivation decreased the expression of the anti‐catabolic factor TIMP‐1 in NC cells but had no effect on MNP gene expression. However, the combination of glucose deprivation and high pressure resulted in a decrease in aggrecan, collagen II and ADAMTS‐5 gene expression in MNP cells. NC cells did not alter their gene expression response to the combined pathological insult. Together these findings show that NC cells are the major producers of proteoglycans, and are resistant to mechanical stress. In contrast, MNP cells produce relatively less proteoglycan and are susceptible to mechanical stress, which is exacerbated under glucose‐deprived conditions. This study has shown that NC and MNP cells from the same IVD have dramatically different responses to two key pathological stimuli, providing insight into the differential roles these cells play in the pathogenesis of disc degeneration.

Name thesis:degree_name

PhD

Level thesis:degree_level

Doctoral

Discipline thesis:degree_discipline

Biomedical Science

Grantor dc:publisher

ResearchSpace@Auckland

Year dc:date.issued

2016

Author dc:creator

• Saggese, Taryn

Advisors dc:contributor.advisor

• McGlashan, S
• Thambyah, A

Statement dc:rights

• Items in ResearchSpace are protected by copyright, with all rights reserved, unless otherwise indicated. Previously published items are made available in accordance with the copyright policy of the publisher.

Licence dc:rights.uri

• https://researchspace.auckland.ac.nz/docs/uoa-docs/rights.htm

Handle dc:identifier.uri

https://hdl.handle.net/2292/28254

OAI identifier oai:identifier

oai:researchspace.auckland.ac.nz:2292/28254