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An investigation into the mechanism linking cartilage calcification and neovascularisation.

Di Benedetto, Giuseppe; (1991) An investigation into the mechanism linking cartilage calcification and neovascularisation. Doctoral thesis (Ph.D.), University College London. Green open access

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Although angiogenesis is an essential component of growth plate function, the mechanisms which control it are not understood. This study has demonstrated production of the angiogenic factor ESAF (Endothelial cell Stimulating Angiogenesis Factor) by cultured growth plate chondrocytes. Production of ESAF was stimulated when cells were induced to calcify their matrix by addition of substrates for alkaline phosphatase (ALP), an enzyme implicated in mineralisation. The mineral was shown by X-ray microanalysis to be deposited in a form similar to octacalcium phosphate, a potential precursor to hydroxyapatite. Mineralisation also resulted in virtually complete cessation of collagen synthesis. The degree to which ESAF production was stimulated depended on the substrate used to induce mineralisation, but was not a direct consequence of phosphohydrolytic cleavage: rather there was a requirement for de novo protein synthesis, though ESAF is not itself a protein. Inhibition of ALP resulted in decreased mineral deposidon and ESAF producdon. The use of stereospecific ALP inhibitors indicated that ESAF production was more closely related to mineral deposition than to enzyme activity, a finding which was corroborated by the observation that addition of mineral to cell cultures also stimulated ESAF production. ESAF is a potent activator of latent collagenase, and its involvement in growth plate remodelling was invesdgated in vitro using rachitic rat growth plates and high-density chondrocyte cultures. Procollagenase produced by rachitic rat growth plates was activated by ESAF in a dose-dependent manner following destruction of the tissue inhibitor of metalloproteinases. Culture of rachitic tissue in media containing ALP substrates resulted in increased mineral deposition and production of an ESAF-like component. However, mineralisation of high density chondrocyte cultures failed to stimulate collagenase activity as monitored by the release of tritiated proline from the cell layer. Addition of ESAF resulted in a 218% increase in proline release, whereas addidon of Interleukin-1 to stimulate procollagenase synthesis increased release by only 29%. There was a partial requirement for de novo protein synthesis in ESAF-mediated proline release, since cycloheximide treatment depressed release by 20%. This study has demonstrated production of a procollagenase-activating angiogenic factor by growth plate chondrocytes in response to deposition of mineral in the extracellular matrix. Furthermore, this factor is implicated in the dynamic remodelling which is a central feature of growth plate function.

Type: Thesis (Doctoral)
Qualification: Ph.D.
Title: An investigation into the mechanism linking cartilage calcification and neovascularisation.
Open access status: An open access version is available from UCL Discovery
Language: English
Additional information: Thesis Digitised by Proquest.
URI: https://discovery.ucl.ac.uk/id/eprint/10124876
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