This work used <i>Pax6</i><sup>+/-</sup> mice as a model for Pax6-realted corneal diseases and assessed the roles of oxidative stress and epithelial injury in the aetiology of ARK. Histological investigation revealed epithelial lesions in <i>Pax</i>6<sup>+/-</sup> corneas. Proteomic analysis demonstrated reduced levels of protective enzymes, transketolase, aldehyde dehydrogenase 3A1 and glutathione S-transferase α4, and cytoskeletal proteins. Keratin 5 and 12 in <i>Pax</i>6<sup>+/-</sup> adult mouse corneas. These physical/structural and chemical defects imply that <i>Pax</i>6<sup>+/-</sup> corneas may be in a chronic ‘stressed and wounded’ state. Using a DNPH/protein oxidation assay, <i>Pax</i>6<sup>+/-</sup> corneal protein oxidation was found to be consistently higher than that in <i>wild-type</i> (WT), and to get worse with age, in parallel with the development of corneal opacity. H<sub>2</sub>O<sub>2</sub> was used to induce oxidative stress in mouse corneas and this was found to activate the Ca<sup>2+</sup> (protein kinase C/ phospholipase C) → p38/p42/p44 mitogen activated kinase signalling pathway. Oxidative stress-induced Pax6 exclusion form cell nucleus led to abnormal expression of non-corneal epithelial markers, indicating a metaplasia process that may cause normally transparent epithelial cells to become opaque. This report for the first time describes cytoskeleton architectures <i>in vivo</i> using flat-mount mouse corneal epithelial by fluorescent staining and confocal microscopy, which is potentially applicable to studies interested in cytoskeleton <i>in vivo</i>. Keratin, desmoplakin and actin cytoskeletal structures were found to be heterogeneous and defective in <i>Pax</i>6<sup>+/-</sup> cells. Twenty-one hours after wounding WT corneal epithelia <i>in vivo</i>, healing cells developed desmoplakin and actin structural features, intercellular gaps, interdigitated filopodia-like processes and vesicles similar to the unscratched <i>Pax</i>6<sup>+/-</sup> corneal epithelia. These data support the hypothesis of a ‘chronic wounded’ state in apparently uninjured <i>Pax</i>6<sup>+/-</sup> corneal epithelia and reveal the cytoskeletal origins of poor adhesion and cellular structure.
| Identifer | oai:union.ndltd.org:bl.uk/oai:ethos.bl.uk:499697 |
| Date | January 2008 |
| Creators | Ou, Jingxing |
| Publisher | University of Aberdeen |
| Source Sets | Ethos UK |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
| Source | http://digitool.abdn.ac.uk:80/webclient/DeliveryManager?pid=25200 |
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