| Synchrotron infrared (IR) microspectroscopy, a technique that allows the spatially resolved determination and mapping of multiple components in situ, was applied to the study of tissue sections. IR spectra provide information about a sample's chemical composition and conformation, such as the secondary structure of proteins. Bright synchrotron light allowed the high spatial resolution (10 microns) needed for this study. The hypothesis was that the molecular information available from IR microspectroscopy would reveal new information about Alzheimer's disease pathology and scar tissue formation.; The first project involved studying brain tissue from TgCRND8 mice, a transgenic model of Alzheimer's disease (AD), the leading cause of dementia in the ageing population. The main hallmark of AD is the deposition of plaques composed of aggregated amyloid beta (Abeta) peptide in the brain. Dense-cored and diffuse plaques in the hippocampus, cortex and caudate were IR mapped and the results correlated with histochemistry and immunostaining. Spectral analysis confirmed that congophilic plaque cores were composed of protein in a beta-sheet conformation, with elevated beta-sheet restricted to the core. The amide I maximum of plaque cores was 1623 cm-1; there was no evidence of the high frequency (1680-1690 cm-1) peak seen in in vitro Abeta fibrils and attributed to anti-parallel beta-sheet. A significant elevation in phospholipids was found around dense-cored plaques in TgCRND8 mice ranging in age from 5 to 21 months. This was due to an increase in cellular membranes from dystrophic neurites and glial cells around the core, but could also contribute to Abeta aggregation through the interaction of newly secreted Abeta with the phospholipids. In contrast, diffuse plaques were not associated with IR detectable changes in protein secondary structure or relative concentrations of other tissue components.; In addition, focally elevated deposits of creatine, a molecule with a crucial role in energy metabolism, were discovered in AD tissue with synchrotron IR microspectroscopy. Both energy metabolism and creatine kinase activity are decreased in AD. The creatine deposits may therefore be a previously undiscovered disease marker.; A second project in this thesis was part of a Natural Sciences and Engineering Research Council Collaborative Health Research Project (NSERC-CHRP) to test the hypothesis that treatment with anti-oxidants, L-2-oxo-thiazolidine-4-carboxylate (OTC) and quercetin, following spinal surgery may reduce oxidative stress, inflammation, and scarring. In humans, excessive formation of scar tissue is the main cause of back pain following spinal surgery. The effect of OTC and quercetin was studied in rats that had undergone laminectomy. Initial synchrotron IR microspectroscopy data were collected on scar tissue from OTC, quercetin and saline (control) treated animals, sacrificed at 3 and 21 days post-surgery (n = 5 per age and treatment group). Scar tissue at 3 days post-surgery was low in collagen, and high in proteoglycans and nucleic acids. The scar tissue at 21 days post-surgery was heterogeneous, containing regions at various stages of development. However, it was marked by the deposition of collagen and a decrease of lipids, due to a reduction in the number of inflammatory cells and fibroblast in the scar. The spectral differences could therefore be correlated with the stages of wound healing. Additionally, the intrinsically polarized synchrotron light was found to affect IR spectra of fibrous 21-day-old scar tissue, rich in oriented collagen. Achilles tendon was therefore studied as a model system of oriented collagen. |
