Diss. (sammanfattning) Uppsala : Univ., 2003. / Härtill 7 uppsatser.

Thesis (Ph.D.). / Written for the Dept. of Pharmacology and Therapeutics. Title from title page of PDF (viewed 2008/05/09). Includes bibliographical references.

Thesis (Ph.D.). / Written for the Dept. of Pharmacology and Therapeutics. Title from title page of PDF (viewed 2008/05/09). Includes bibliographical references.

Thesis supervisor: Stefan Strack. Includes bibliographic references (p. 87-100).

Thesis (Ph. D.)--University of Virginia, 1999. / Includes bibliographical references (p. 237-268). Also available online through Digital Dissertations.

Transmissible spongiform encephalopathies (TSEs) are rare, uniformly fatal neurodegenerative disorders that can affect many mammalian species, including humans. A hallmark of these diseases is the conversion of cellular prion protein (PrPC) into an abnormally folded form. This misfolded PrPC is infectious, since it can provide a template for pathogenic conversion of PrPC in a new host. In addition to any toxicity of the misfolded protein, loss of normal PrPC function could be involved in the neurodegenerative processes. However, the physiological role of PrPC is still poorly understood and this project has aimed to address that lack of knowledge. Out of the many putative functions ascribed to PrPC, the most commonly proposed is that it protects cells from stress. In contrast, I have found that stable transfection of the prion protein gene into SH-SY5Y neuroblastoma cells increases cell death in response to serum removal from the culture medium. Following treatment with several chemical toxins, two out of four stably transfected clones did, generally, display greater viability than untransfected cells that do not express detectable levels of PrPC. However, knockdown of PrPC expression by RNA interference had no effect on this stress resistance, indicating that it may not have been mediated directly by PrPC. Given the lack of robust stress protection afforded by PrPC transfection, proteomic analyses of the cells were carried out to identify alternative processes that were perturbed as a result of PrPC expression. The results obtained suggested roles for PrPC in cytoskeletal organisation and cell cycle regulation. Various proteins involved in cytoskeletal organisation were confirmed by western blotting to be differentially expressed in some or all of the stably transfected clones. Additionally, the expression changes to proteins involved in cell cycle regulation resulted in slower proliferation of the clones compared with untransfected cells, a difference that was reduced following RNA interference-mediated knockdown of PrPC. Taken together, these data suggested that specific growth factor-activated pathways were differentially regulated in the stably transfected clones. One candidate pathway was nerve growth factor (NGF) signalling, which promotes neuronal survival and differentiation as well as regulating various processes outside of the nervous system. PrPC-transfection resulted in altered expression of receptors for NGF, suggesting that the stably transfected clones were, indeed, responding differently to NGF stimulation. However, the molecular mechanism responsible for these expression changes remains to be determined, since co-immunoprecipitation experiments did not identify any physical interactions between PrPC and the NGF receptors. Nonetheless, a role for PrPC in modulating NGF signalling has the potential to explain many of the diverse phenotypic observations in PrPC-null mice and might indicate that loss of PrPC function is an important part of TSE pathogenesis.

The formation of axon collateral branches from the pre-existing shafts of axons is an important aspect of neurodevelopment and the response of the nervous system to injury. Both the actin filament and microtubule components of the cytoskeleton are required for the formation of axon branches. Recent work has begun to shed light on how these two elements of the cytoskeleton are integrated by proteins that functionally or physically link the cytoskeleton. While a number of signaling pathways have been determined as having a role in the formation of axon branches, the complexity of the downstream mechanisms and links to specific signaling pathways remain to be fully determined. Neurotrophins are growth factors that have a multitude of roles in the nervous system. In sensory neurons nerve growth factor (NGF) induces branching through activation of phosphoinositide 3-kinase (PI3K). Recently, mitochondria have emerged as major determinants of the sites of axon branching. In this work we reveal a new role of neurotrophins in mitochondria fission. We report that NGF promote a rapid burst of mitochondria fission, followed by a new steady state of mitochondria length and density. Mek- Erk and PI3k pathways are required for NGF-induced fission. Mek-Erk controls fission through Drp1 activation, while we suggest that PI3K may contributes to the actin dependent aspect of fission. Drp1 mediated fission is required for NGF- induced branching in sensory neurons in vitro and the branching of sensory axons along the developing spinal cord. We reveal that fission is also required for the intra-axonal translation of the actin regulatory proteins Cortactin and Arp2 subunit from the Arp2/3 complex, an important aspect of NGF induced branching. Collectively, these observations reveal a novel role of neurotrophins in mitochondria fission and the formation of collateral branching / Neuroscience

No / Neuroimmune dysregulation characterizes atopic disease, but its nature and clinical impact remain ill-defined. Induced by stress, the neurotrophin nerve growth factor (NGF) may worsen cutaneous inflammation. We therefore studied the role of NGF in the cutaneous stress response in a mouse model for atopic dermatitis–like allergic dermatitis (AlD). Combining several methods, we found that stress increased cutaneous but not serum or hypothalamic NGF in telogen mice. Microarray analysis showed increased mRNAs of inflammatory and growth factors associated with NGF in the skin. In stress-worsened AlD, NGF-neutralizing antibodies markedly reduced epidermal thickening together with NGF, neurotrophin receptor (tyrosine kinase A and p75 neurotrophin receptor), and transforming growth factor-β expression by keratinocytes but did not alter transepidermal water loss. Moreover, NGF expression by mast cells was reduced; this corresponded to reduced cutaneous tumor necrosis factor-α (TNF-α) mRNA levels but not to changes in mast cell degranulation or in the T helper type 1 (Th1)/Th2 cytokine balance. Also, eosinophils expressed TNF receptor type 2, and we observed reduced eosinophil infiltration after treatment with NGF-neutralizing antibodies. We thus conclude that NGF acts as a local stress mediator in perceived stress and allergy and that increased NGF message contributes to worsening of cutaneous inflammation mainly by enhancing epidermal hyperplasia, pro-allergic cytokine induction, and allergy-characteristic cellular infiltration.

Thesis (Ph. D.)--West Virginia University, 2003. / Title from document title page. Document formatted into pages; contains xiii, 82, [148] p. : ill. (some col.). Includes abstract. Includes bibliographical references.

Thesis (M.Sc.)--Memorial University of Newfoundland, Memorial University of Newfoundland, 1998. / Typescript. Bibliography: leaves 118-132.