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Characterising the novel activation of wt1b in the notochord damage response of zebrafish larvaeLopez Baez, Juan Carlos January 2015 (has links)
The notochord is the defining structure of all chordates. A semi-‐flexible elongated tube of cells, it forms along the central axis of the embryo and provides axial support during development. It also acts as a signalling centre during early embryogenesis, controlling the patterning of a number of tissues and establishing the early body axis of the embryo. In vertebrates, the function of the notochord expands beyond early development. It creates morphogenic gradients for the patterned formation of the vertebral bodies and, in adults, the remnants of the notochord form the nucleus pulposus, a gel-‐like structure with an integral role in the distribution of vertebral pressure in the intervertebral disc. Little is known about how the notochord copes with damage during embryogenesis, but degeneration of the nucleus pulposus can lead to debilitating spinal disorders. In this thesis, I use a zebrafish model system to present new data that describes the cellular behaviours associated with how the notochord copes with external damage and how this damage can influence the future development of the vertebrae. I have uncovered a novel damage response in the notochord of zebrafish larvae and characterised the morphogenetic changes involved in the process using transgenic fluorescent lines. I have explored the damage in the context of the Wilms’ Tumour 1 (Wt1) gene, a vertebrate-‐conserved transcription factor, which has recently been associated with several regenerative responses, and discovered that one of its zebrafish orthologues, wt1b, becomes upregulated in the notochord damage response. I have used fluorescent confocal imaging and immunohistochemistry to present new evidence that shows that upon injury, the outer notochord sheath cells upregulate the expression of wt1b. Additionally, I have used time-‐lapse microscopy to show that damage to the notochord induces novel morphological changes in the injured organ, which include the loss of cellularity of the inner vacuolated cells and the movement of the wt1b-‐positive outer sheath cells into the injured lumen. Long-‐term imaging experiments have also demonstrated the capacity of the notochord to heal the damage over time, which ultimately leads to the formation of an extra, smaller vertebra in the wounded area. Skeletal staining of these fish has revealed a previously unknown putative cartilage switch at the site of damage, which leads to the formation of the new vertebral body. This finding has been supported by the microarray analysis of the injured area, which shows the unexpected de-‐novo expression of cartilage markers at the site of damage The work in this thesis identifies for the first time an endogenous repair mechanism in the notochord of zebrafish larvae and describes the cellular, genetic and molecular processes cotrolling this novel wt1b-‐associated damage response.
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Characterization of the Interface between the Annulus Fibrosus and the Vertebral Bone.Nosikova, Yaroslavna 15 December 2011 (has links)
Replacing a diseased disc with a tissue engineered disc has the potential to restore normal spinal biomechanics. However, recreating the interface between annulus fibrosus (AF) and vertebral bone (VB) will be necessary to facilitate proper function of the implant in vivo. This study characterizes the native bovine AF-VB interface and assesses adult human discs. The AF insertion site in humans and cows is uniquely differentiated from other soft tissue-bone interfaces, as AF collagen fibers anchor into the calcified region of vertebral endplate through a zone of hyaline cartilage and have a different organization in inner and outer AF. Mineralization-associated proteins are present in this region and the chondroid tissue undergoes calcification over time. Based on these observations an in vitro AF culture system was developed and demonstrated that AF cells can induce mineralization. Understanding mechanism(s) regulating AF mineralization will help develop conditions to ensure proper integration of bioengineered AF.
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Characterization of the Interface between the Annulus Fibrosus and the Vertebral Bone.Nosikova, Yaroslavna 15 December 2011 (has links)
Replacing a diseased disc with a tissue engineered disc has the potential to restore normal spinal biomechanics. However, recreating the interface between annulus fibrosus (AF) and vertebral bone (VB) will be necessary to facilitate proper function of the implant in vivo. This study characterizes the native bovine AF-VB interface and assesses adult human discs. The AF insertion site in humans and cows is uniquely differentiated from other soft tissue-bone interfaces, as AF collagen fibers anchor into the calcified region of vertebral endplate through a zone of hyaline cartilage and have a different organization in inner and outer AF. Mineralization-associated proteins are present in this region and the chondroid tissue undergoes calcification over time. Based on these observations an in vitro AF culture system was developed and demonstrated that AF cells can induce mineralization. Understanding mechanism(s) regulating AF mineralization will help develop conditions to ensure proper integration of bioengineered AF.
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Back to the beginning: identifying lesions of diffuse idiopathic skeletal hyperostosis before vertebral ankylosisCastells Navarro, Laura, Buckberry, Jo 06 January 2020 (has links)
Yes / Objective: To better understand the pathogenesis of DISH, identifying early or pre-DISH lesions in the spine and investigating the relationship between spinal and extra-spinal manifestations of DISH.
Material: 44 skeletonized individuals with DISH from the WM Bass Donated Skeletal Collection.
Methods: For each vertebra, location, extension, point of origin and appearance of vertebral outgrowths were recorded. The size of the enthesophytes at the olecranon process, patella and calcaneal tuberosity was measured with digital callipers.
Results: At either end of the DISH-ankylosed segment, isolated vertical outgrowths arising from the central third of the anterior aspect of the vertebral body can usually be observed. These bone outgrowths show a well-organized external cortical layer, an internal structure of trabecular bone and usually are unaccompanied by or show minimal associated endplate degeneration. Analysis of the relationship between spinal and extra-spinal manifestations (ESM) suggests great inter-individual variability. No correlation between any ESM and the stage of spinal DISH was found.
Conclusions: Small isolated outgrowths represent the earliest stages of the spinal manifestations of DISH. The use of ESM as an indicator of DISH should be undertaken with great caution until the relationship between these two features is understood.
Significance: Improved accuracy of paleopathological diagnostic criteria of DISH.
Limitations: Small sample comprised of only individuals with DISH.
Future research: micro-CT analysis to investigate the internal structure of the spinal lesions. Analysis of extra-spinal enthesophytes in individuals with and without DISH to understand their pathogenesis and association with the spinal lesions in individuals with DISH. / Institute of Life Sciences Research Studentship awarded by the University of Bradford, Bradford, UK.
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