Thesis (Ph. D.)--University of Hong Kong, 2006. / Title proper from title frame. Also available in printed format.

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published_or_final_version / Orthopaedics and Traumatology / Doctoral / Doctor of Philosophy

Thesis (MAppSc in Physiotherapy) -- University of South Australia

Herniation of the intervertebral disc is a pathological condition characterized by protrusion of the tissue posterio-laterally, often impinging on the spinal chord or nerve roots. The disease is accompanied by a reduction in the average molecular weight and viscosity of the disc proteoglycans, in excess of that which normally occurs with increasing age. The proteoglycans of disc however have not been examined in terms of the modern concepts of cartilage matrix structure. Bovine nasal cartilage, has been shown to contain proteoglycan aggregates, trapped in the intersticess of a collagen network, which can be dissociated with 4M guanidine hydrochloride into diffusible proteoglycan subunits and a multicomponent "linking" fraction. A similar system was thought to occur in the intervertebral disc. It seemed possible that, if such a system was present in the disc, the reduction in the molecular weight and viscosity of the proteoglycans with increasing age and herniation could be due to a decrease in proteoglycan aggregation. The present study showed that proteoglycan aggregates similar to those of bovine nasal cartilage are found in the human intervertebral disc, but that they only represent 5% of the total proteoglycans in the tissue. In contrast, bovine nasal cartilage contained 70% of the proteoglycans in the aggregated form. A novel modification of the extraction procedure was devised by which it was possible to assess the degree of proteoglycan aggregation. Sequential extraction of the tissue with a weak and strong electrolyte (0.4 M and 4M guanidine hydrochloride) selectively removed non-aggregated and aggregated proteoglycans respectively. This procedure provides a new and rapid method for assessing the degree of proteoglycan aggregation in a variety of connective tissues. The small proportion of aggregate in the disc was almost exclusively located in the annulus fibrosus. Re-aggregation studies suggested that both disc and cartilage contain two proteoglycans, only one of which is capable of forming aggregates. Examination of the proteoglycans in a limited number of discs suggested that the degree of aggregation did not change with increasing age. Since, in addition, aggregates represent only a small proportion of the disc proteoglycans, it appeared unlikely that a decrease in the degree of aggregation could account for the decrease in molecular weight and viscosity of the disc proteoglycans observed with increasing age and/or degenerative disc disease. / Medicine, Faculty of / Pathology and Laboratory Medicine, Department of / Graduate

published_or_final_version / Orthopaedics and Traumatology / Doctoral / Doctor of Philosophy

abstract / Orthopaedics and Traumatology / Doctoral / Doctor of Philosophy

published_or_final_version / Orthopaedics and Traumatology / Doctoral / Doctor of Philosophy

abstract / Orthopaedics and Traumatology / Doctoral / Doctor of Philosophy

Back pain is a prevalent musculoskeletal disorder affecting populations worldwide. Degeneration of the intervertebral discs (IVD) is indicated to be one of the main etiologic factors of back pain. The loss of proteoglycan and consequently dehydration and reduction of swelling of the intervertebral disc core, the nucleus pulposus (NP), is one of the earliest degenerative events. The replacement of large vacuolated notochordal cells by the smaller chondrocyte-like cells in the NP coincide with the onset of IVD degeneration, the loss of the notochordal cells was therefore postulated to be a cause of the degeneration and associated with ageing . To date, the true origin of the smaller chondrocyte-like cells was still under dispute. In this study, Egfp reporter was constitutively and permanently activated in the notochord via specific Cre expression by the Foxa2mNE enhancer under the recombination system in the Foxa2mNE-cre; Z/EG double transgenic mice. The notochord descendent cells were therefore labeled with green fluorescent protein (GFP) and their localization and expression characteristic were tracked during development, maturation, ageing of mouse tail IVD. To further investigate the cellular changes during degeneration, disc degeneration in the murine disc was induced by puncture. The GFP labeled notochord cells were demonstrated to be entrapped and remained in the NP of the murine disc. With the validation of degeneration by the multichromatic FAST staining, a grading system based on altered sulfated glycosaminoglycan content and cellular organization specialized to murine disc was proposed. The segregation of the notochordal NP cell mass segregation was found to be a common pathway of both age-related and puncture-induced degeneration. While apoptosis and loss of NP cells was presented in puncture-induced disc degeneration, invasion of lamellae cells was only found in severely degenerated discs. Consistent with the findings of the murine puncture model, increased expression of Col2a1 and Col1a1 were demonstrated in punctured discs by ISH, which the notochordal NP cells were showing a remarkable elevation in expression. The data supports a chondrogenic differentiation of the endogenous notochordal cells instead of invasion of chondrocyte-like cells into the NP. Three NP markers identified as a common genes from microarray of rat and human NP, (Carbonic anhydrase3 (Car3), UDP-N-acetyl-alpha-D-galactosamine:polypeptide; N-acetylgalactosaminyltransferase 3 (Galnt3) and HOP homeobox (Obl)), were found specifically co-localized with the notochord descendent cells in all ages and different time-points after puncture, suggesting them as novel but reliable markers for notochordal NP cells. This study provides direct evidence of the origin of NP cells in aged and degenerated IVDs using transgenic mice, and thus enhances the understanding of the role of notochordal cells in the progression of IVD degeneration. / published_or_final_version / Orthopaedics and Traumatology / Doctoral / Doctor of Philosophy