| dc.description.abstract | Lower Back Pain (LBP) is known to be a major health issue worldwide with a
significant socioeconomic impact and it is widely accepted that LBP is associated with
degeneration of the intervertebral disc (IVD). IVD degeneration is thought to initiate within
the nucleus pulposus (NP), a highly hydrated gel-like matrix rich in proteoglycans (PG) and
glycosaminoglycans (GAG). Being an avascular structure, the nutrient supply within the IVD
is limited, resulting in a compromised microenvironment with limited oxygen and glucose, a
low pH and the presence of inflammatory factors which are responsible for cell death and loss
of structural integrity. Cell-based therapies have sought to restore the heathy tissue composition
by implanting healthy cells to the degenerated site. However, several limitations are associated
with cell-based techniques, including poor cell retention and the need of extensive 2D
expansion to obtain the required cell numbers.
The overall aim of this thesis was to develop an injectable and biomimetic biomaterial
that can aid nasal chondrocytes (NC) in the regeneration of the NP region of the IVD.
Specifically, this thesis set out to explore: i) the development of an injectable and biomimetic
biomaterial system to permit cell delivery to the area of interest, ii) the potential of NCs as an
alternative cell source due to their ability to synthesise NP-like tissue, iii) the influence of the
harsh microenvironmental conditions typical of the degenerated disc niche on the ability of
NCs to remain viable and deposit de novo matrix.
This thesis began with the development of an injectable biomaterial derived from
decellularised and solubilised disc extracellular matrix (ECM) to be employed as a cell carrier.
In an attempt to meet the initial requirements of biomimicry, functionalised chondroitin sulfate
(fCS) was added to the material composition. The addition of fCS to the gel composition was
found to enhance key physical characteristics of the hydrogels such as high swelling capacity
and short gelation time and to influence NC matrix deposition capacity by enhancing the
synthesis of sGAG.
The thesis continued by investigating how microenvironmental conditions may affect
viability and matrix production of NCs and NP cells under varying pH (7.1, 6.8 and 6.5) and
with the presence of physiologically relevant levels of inflammatory cytokines such as IL-1β
and TNF-α. It was found that acidity of the media is the main factor influencing the biological
behaviour of cells in culture, while the presence of inflammatory cytokines had negligible
effects in 3D culture conditions. Moreover, it was found that culture in degenerative conditions
is more detrimental to NP cells than NCs, causing increased cell death and poor matrix
synthesis.
Further, the effects of culturing NCs at different seeding densities (1x106, 2.5x106,
5x106 and 10x106 cells/ml) were investigated and results demonstrated a correlation between
higher seeding density and both decreased cell viability and matrix accumulation.
Finally, a disc explant model was developed with the aim of testing the potential of the
developed decellularised ECM biomaterial and NCs in simulated degenerative disc conditions
in comparison to NP cells. Results from this study, demonstrated that NCs possess a more
robust and higher resilience to degenerated disc-like culture conditions compared to NP cells,
which translated into higher viability and the synthesis of NP-like matrix.
In conclusion, this work provides early stage validation for a two-step regenerative
approach that involves the use of NCs isolated from a nasal biopsy, expanded in vitro and
injected in situ in low numbers in a NP ECM-derived hydrogel previously functionalised with
CS. Moreover, it also provides important insights on the influence of environmental acidity
and physiological inflammatory conditions on the biological behaviour of NCs and NP cells,
which may be critical in determining the optimal opportunity for surgical intervention. | en |
