published_or_final_version / abstract / toc / Paediatrics and Adolescent Medicine / Master / Master of Philosophy

published_or_final_version / Anatomy / Master / Master of Philosophy

published_or_final_version / Biochemistry / Doctoral / Doctor of Philosophy

No description available.

No description available.

The normal aging process is accompanied by changes in cognitive function. One of the brain regions known to be an early target of the aging process is the hippocampus, a medial temporal lobe structure that is critically involved in spatial learning and memory function. The formation and maintenance of memory relies on rapid and sustainable synaptic modification, which requires new gene expression. Immediate-early genes are the first genes to be induced following relevant stimuli, and include genes that encode transcription factors, such as c-fos and zif268, and effector proteins that directly influence cellular function, such as Arc (activity-regulated cytoskeletal gene) and Homer1A. Blocking the expression of any one of these genes interferes with memory function, and thus, each of these genes is thought to have a memory enhancing effect. The hypothesis tested here was that aged animals would show a reduction in the expression of memory-promoting immediate-early genes within the hippocampus, and moreover, that these changes in expression would be subregion specific, based on the finding that the dentate gyrus is most vulnerable to the aging process.Potential age-related changes in immediate-early gene expression within the hippocampus was determined under basal conditions and after induction by a simple behavioral task. Of the genes under investigation, only c-fos did not show age-related changes under basal conditions, or following behavioral induction. The remaining genes, Arc, zif268 and Homer1A, each showed subregion specific patterns of change within the hippocampus under basal conditions or following induction (or both). The coordinate expression of immediate-early genes within the hippocampus was also investigated by assessing the extent to which Arc was expressed within the same neurons as c-fos, zif268 or H1a. The coordinate transcription of these genes was not significantly altered in the aged hippocampus, even though changes in the size of Arc and zif268 neural ensembles occurs within the aged denate gyrus.Taken together, these data indicate that age-related reductions in the basal and induced levels of immediate-early genes occur within the hippocampus, and that these changes are subregion specific.

Neutrophils, key cells of the innate immune system, are responsible for preventing bacterial infections. They are rapidly recruited to sites of infection where they eliminate bacteria through killing methods that require reactive oxygen dependent processes. It has recently been established that neutrophils are capable of rapid and complex changes in gene expression during inflammatory responses. The concept that neutrophils only directly kill bacteria has been replaced by the concept that activated neutrophils can influence the immune response through the secretion of a variety of cytokines and by acting as antigen-presenting cell (APC) expressing MHC Class II, allowing for activation of T cells. Recent advances in neutrophil biology demonstrated that neutrophils also have an active regulatory role in angiogenesis and tumoral fate. It has been noted that a number of diseases including arthritis, periodontitis and acute respiratory distress syndrome (ARDS) are associated with neutrophil hyperactivity that results in significant tissue damage. Our group has previously shown that for some periodontal diseases, neutrophil hyperactivity is a key determinant of disease progression and severity. However, it remains unclear what factors are responsible for a patient developing a hyperactive neutrophil mediated disease. I hypothesize that local gene expression changes in neutrophils are responsible for the hyperactive behaviour of these cells during an inflammatory response. In order to assess this, I characterized the neutrophil gene expression profile in various compartments (bone marrow, blood and peritoneum in mice and blood and oral cavity in humans) and then characterized this genetic and phenotypic profile during an inflammatory response. I hypothesize that the neutrophil has a characteristic set of genes that are normally activated when it enters a site of inflammation from the circulation and that neutrophils can be polarized into a different functional subset under certain conditions that result in inflammation mediated diseases. To identify changes in neutrophil gene expression in the circulation and inflamed tissue I used recent advances in neutrophil isolation, RNA amplification, and microarray technologies to characterize the specific transcriptome associated with neutrophil site-specific responses.

Neutrophils, key cells of the innate immune system, are responsible for preventing bacterial infections. They are rapidly recruited to sites of infection where they eliminate bacteria through killing methods that require reactive oxygen dependent processes. It has recently been established that neutrophils are capable of rapid and complex changes in gene expression during inflammatory responses. The concept that neutrophils only directly kill bacteria has been replaced by the concept that activated neutrophils can influence the immune response through the secretion of a variety of cytokines and by acting as antigen-presenting cell (APC) expressing MHC Class II, allowing for activation of T cells. Recent advances in neutrophil biology demonstrated that neutrophils also have an active regulatory role in angiogenesis and tumoral fate. It has been noted that a number of diseases including arthritis, periodontitis and acute respiratory distress syndrome (ARDS) are associated with neutrophil hyperactivity that results in significant tissue damage. Our group has previously shown that for some periodontal diseases, neutrophil hyperactivity is a key determinant of disease progression and severity. However, it remains unclear what factors are responsible for a patient developing a hyperactive neutrophil mediated disease. I hypothesize that local gene expression changes in neutrophils are responsible for the hyperactive behaviour of these cells during an inflammatory response. In order to assess this, I characterized the neutrophil gene expression profile in various compartments (bone marrow, blood and peritoneum in mice and blood and oral cavity in humans) and then characterized this genetic and phenotypic profile during an inflammatory response. I hypothesize that the neutrophil has a characteristic set of genes that are normally activated when it enters a site of inflammation from the circulation and that neutrophils can be polarized into a different functional subset under certain conditions that result in inflammation mediated diseases. To identify changes in neutrophil gene expression in the circulation and inflamed tissue I used recent advances in neutrophil isolation, RNA amplification, and microarray technologies to characterize the specific transcriptome associated with neutrophil site-specific responses.

No description available.

Genes expressed in developing pea pods were isolated as cDNAs by differential screening techniques. The cDNAs were characterised by DNA sequencing and expression studies were used to investigate the role of isolated cDNAs in pod development. A clone isolated from a pea {Piswn sativum L.) pod cDNA library was shown to contain the complete coding sequence of a polypeptide with considerable homology to various members of the Rab subfamily of small ras-related GTP- binding proteins. Conserved sequences in the isolated clone include the GTP-buiding site, GDP/GTP hydrolysis domain and C-terminal Cys residues involved in membrane attachment. The high percentage amino acid identity suggests that this cDNA may be the product of a gene, designated Psa-rai?, which is the plant counterpart of Rab7. Rab/Ypt proteins are thought to be involved in intracellular transport from the endoplasmic reticulum to the Golgi apparatus and in vesicular transport. If Psa-ra6 is a functional counterpart of yeast YPT7 (RabT) it should be able to complement a yeast YPT7 mutant. An attempt was made to demonstrate that this was the case. Northern analysis showed invariant expression of Psa-rab in developing pods 'with different phenotypes, indicating an essential function for Psa-rab in developing pods. Hybridisation of the Psa-rab cDNA to pea genomic DNA showed that this protein is probably encoded by a single gene. Nearly isogenic pea lines were selected to investigate the genetic basis for lignification of the pea {Pisum sativum L.) pod endocarp. The development of the pod endocarp in the normal and mutant pea pod phenotypes was examined by histochemical staining and light microscopy. The effect of plant growth regulators on endocarp development was also investigated. A pea pod cDNA library representing poly (A)+ RNA purified from L59 pea pods (genotype, PV; phenotype, lignified endocarp) was differentially screened with total cDNA probes prepared from total pod RNA from L59 and LI390 (genotype, PV; phenotype, no lignification of endocarp) pods 4-6 days after flowering (DAP). Two clones, designated pLP18 and pLP19, were selected for further characterisation on the basis of hybridisation to the L59 cDNA probe, but not the LI390 cDNA probe. Northern blotting was used to show that pLP18 represented a mRNA of 0.95 kb. The predicted polypeptide from the LP18 cDNA encoded a putative blue type I copper protein. The expression pattern of LP 18 mRNA in pods and tissues of the experimental pea lines was determined using RT-PCR quantitation. Hybridisation of the cDNA to pea genomic DNA showed that this protein is probably encoded by a single gene. Clone pLP19 yielded a 1.02 kb cDNA fragment encoding the C-terminal portion of an Hsp70 homologue belonging to a highly conserved family of proteins found in a number of eukaryotic species. Northern analysis of RNA from lignified and unlignified pods showed the presence of differentially expressed LP19 transcripts of varying lengths, which may represent differently processed transcripts. Southern analysis confirmed the presence of a single hybridised band in genomic digests of L59, L58 arid LI390. Several mRNA transcripts of the LP19 gene were isolated which differ in the length of their- 3' untranslated regions.