Thesis (Ph. D.)--University of Michigan, 1995. / eContent provider-neutral record in process. Description based on print version record.

Thesis (D. Min.)--Bethel Seminary, St. Paul, MN, 2001. / Abstract. Includes bibliographical references (leaves 159-167).

The problem of this study was to better understand spouse caregivers' leisure involvement, experience, and barriers and their relationships with perceived burden. Thirty-six wife and 19 husband caregivers of persons with Alzheimer's disease and related disorders volunteered to participate in this study, either by mailed questionnaire or interview. Respondents were primarily female, white, with an average age of 72 years. The conclusions of the study were: (a) caregivers significantly reduce both their leisure involvement; (b) self-reported health, perceived social supports, income level, use of paid help, and leisure activity patterns are major factors associated with caregivers' leisure; and (c) leisure barriers are a significant contributor to caregivers' perceived burden. Recommendations were presented for caregivers, practitioners, and future study.

Despite the tremendous advances of the last century, the cause of Alzheimer disease (AD) remains unclear. Genetic analysis of families with Alzheimer disease has revealed a disease-associated variant of the APOE gene, which encodes apolipoprotein E, a transporter of lipids in the blood and central nervous system. The effect of the AD-associated isotype, termed ApoE-E4, on disease risk has been validated, though it is unclear by what mechanism apoE-E4 confers AD risk. Mitochondria have long been implicated in AD pathogenesis, as the canonical histopathological findings of amyloid plaques and tau tangles occur in the setting of mitochondrial dysfunction. The disrupted processes include calcium homeostasis, cholesterol metabolism, phospholipid synthesis, and mitochondrial dynamics, and are all regulated by a subcompartment of the ER that is in physical contact with mitochondria. This compartment, called the mitochondria-associated ER membrane, or MAM, has been found to be overactive in AD patient cell lines and cell models of AD. Given that MAM is dysfunctional in AD and that ApoE-ε4 is the most important risk factor for AD, this dissertation examines if ApoE4 contributes to the MAM dysfunction seen in AD. The MAM dysfunction seen in AD patients and in cell models of AD has been best characterized in the context of familial AD, and it is the purpose of this study to extend those findings to the more common, sporadic, form of the disease. Familial AD is the result of autosomal dominant mutations in one of three genes, amyloid precursor protein (APP), presenilin 1 (PSEN1), or presenilin 2 (PSEN2). APP is the protein from which amyloid-beta, the component of amyloid plaques, is cleaved. The presenilins constitute the enzymatic core of the γ-secretase complex, which cleaves amyloid-beta from a precursor APP molecule. Both PSEN1 (PS1) and PSEN2 (PS2) localize at the MAM, and their action is speculated to influence MAM activity. Fibroblasts from familial AD patients, which contained mutations in APP, PSEN1 or PSEN2, showed a marked increase in MAM activity when compared to that of age-matched controls. In mouse embryonic fibroblasts, one can recapitulate this increased MAM activity by knocking out presenilins 1 and 2. In these Psen1/2 double knockout (DKO) cells, the typical measures of MAM function, i.e. increased cholesteryl ester and phosphatidylethanolamine synthesis, calcium transport from ER to mitochondria, and co-localization of ER and mitochondria by confocal and electron microscopy, mimicked the same phenotype found in fibroblasts obtained from familial AD patients, which suggests that the presenilins are negative regulators of ER-mitochondrial apposition. The precise mechanism by which they regulate the ER-mitochondria interface, whether directly as part of a tethering complex, or indirectly though the metabolism of APP-derived substrates, is unclear. While the effect of familial AD mutations on MAM has been characterized, the mechanism of mitochondrial dysfunction seen in the more common sporadic form of the disease remains obscure. Sporadic AD patients harbor no mutations in APP, PSEN1, or PSEN2, but rather inherit mutations in other genes which do not guarantee the development of the disease, but are instead considered risk factors. The most important of these risk factors, in terms of both amount of AD risk conferred and prevalence in the population, is ApoE. Embedded in the phospholipid monolayer of lipoproteins, ApoE is involved in the transport of phospholipids, cholesterol, and cholesteryl esters in plasma and the central nervous system (CNS). In the CNS, it is the most abundant apolipoprotein, and is secreted primarily by astrocytes and taken up by neurons. Once endocytosed, ApoE can follow three different pathways: degradation by the lysosome, intracellular retention in early endosomes, or re-lipidation and re-secretion out of the cell. Our approach takes advantage of the physiological role of ApoE as part of a high densitylike lipoprotein particle (HDL). Using astrocytes from ApoE targeted gene replacement mice in which murine APOE has been replaced by either human APOE-E3 or human APOE-E4, cultured media containing ApoE3 and ApoE4-lipoproteins can be produced and applied to target cells that do not express ApoE, such as neurons or fibroblasts. These target cells can then be analyzed for MAM activity. To examine the contribution of ApoE towards MAM dysfunction, target cells, either neurons or fibroblasts, were grown in the presence of astrocyte conditioned media (ACM) from ApoE targeted gene replacement mice. Several measures of phospholipid and cholesteryl ester synthesis were performed to analyzed MAM function. To confirm that the alterations in phospholipid synthesis were the result of altered MAM activity, the same assay was performed in cells in which a protein tethers that bind mitochondria and ER were genetically ablated. Finally, to confirm that the effects seen were the result of the HDL particles and not the result of other components of the ACM, lipoproteins were extracted from ACM by density ultracentrifugation and applied to fibroblasts. In all of the assays performed, ApoE4 conditioned media or ApoE4 isolated lipoproteins were able to induce a significant increase in MAM activity, whereas ApoE4 from recombinant sources did not. These data suggest a contribution of ApoE4 towards MAM dysfunction seen in AD. The mechanism of these ApoE4 induced MAM alterations remains to be deduced. One may speculate that given the role of ApoE in cholesterol transport outside of the cell, its intracellular retention may impact the distribution of cholesterol within the cell. MAM is a cholesterol rich subdomain with lipid raft-like properties, and any change in the cholesterol content or lipid nature of this membrane may alter its activity. To test this hypothesis, MAM was biochemically extracted from ApoE3 and ApoE4 treated cells and analyzed for cholesterol and lipidomic content. The results described in this thesis demonstrate an AD-like effect in wildtype cells when treated with ApoE-E4, and that the mechanism for these alterations may be due to disturbances in cholesterol distribution in the MAM.

Epigenetic mechanisms play tremendous roles in the development and management of neural processing. The important mechanisms include inactivation of transcription via methylation, histone modification via acetylation/deacetylation, and miRNA regulation. These modifications allow for expression or silencing of genes, without manipulation of nucleotide sequence. An individual's internal and external environments provide input for quotidian epigenetic regulation. Aberrations in the form of regulation have been increasingly linked to neurological disorders, in addition to the established correlation to tumorigenesis. In recent years, deviations from normal epigenetic patterns have been observed in cases of Alzheimer's disease (AD). The brains of patients with AD have been shown to display significantly less methylation overall, as compared to age-matched controls. Of particular concern, the methylation, which normally keeps the promoter of the APP gene silenced, occur far less frequently in AD patient allowing for the progression of amyloid deposition and subsequent tau pathology. In addition to the hypomethylation present in AD, many AD cases present with a concurrent hypoacetylation on histones in the hippocampus. There is strong evidence suggesting that the reduced levels of acetylation are due to over-activation of histone deacetylases. Post-mortem examinations of the brains of AD patients have shown that the brain-derived neurotrophic gene, which is crucial for neural processing associated with maturation and memory, has low levels of acetylation halting its transcription. While low levels of methylation and acetylation seem to contribute to the pathogenesis of AD, regulatory miRNA levels can have adverse effects whether they are aberrantly reduced or increased. Patients with AD tend to show abnormally augmented expression of miRNA-125b, miRNA-128, and miRNA-9 in the hippocampus, while a reduced expression of miRNA-107. Deregulation of these miRNAs have been linked to the progression of AD and include amyloid deposition, tau pathology, and oxidative stress through inflammatory processes. The latter quandary of oxidative stress has been shown to be crucial for the early progression of AD. Reactive oxygen species disallow the methylation of genes due to steric hindrance at the CpG islands of DNA where DNA methyltransferases act. Research shows that increases in oxidative stress are correlated to decreases in methylation, which allows for APP expression. While these alterations to normal epigenetic patterns occur internally, there is a breadth of changes that the external environment imposes to exacerbated AD pathogenesis. Most heavily studied of these external environmental factors is lead exposure. There is a strong correlation between lead exposure in individuals who carry the ApoE4 gene and increased mRNA transcription of the APP gene. Lead is thought to demethylate the promoter of the APP gene and allow for amyloid processes to occur. Inadequate nutrition, specifically deficits in choline and folate, has been linked to hypomethylated states due to an inefficient "methylation/remethylation cycle" leading to an accumulation of homocysteine characteristic of AD. With the emphasis epigenetic deregulation has in the progression of AD, epigenetic treatments need to be seriously considered as therapeutic avenues. Current drugs treat the symptoms and acute conditions of AD, but through epigenetic modifications, the pathology of the diseases can be directly addressed. Potential therapeutic avenues include the use of methyl donors, highly specific histone deacetylase inhibitors, and miRNA biomarkers. Methyl donors can help alleviate the hypomethylated state and prevent further APP expression and amyloid deposition. Currently, the histone deacetylase inhibitors are being used as global inhibitors, but have adverse effects including non-specific and premature cell death. By further researching these inhibitors and finding a mechanism to attack specific histone deacetylases (such as HDAC6 in AD), the efficacy of this aspect of treatment will be greatly increased. The current use of miRNAs as epigenetic regulators to turn off unwanted genetic expression is ineffective due to a major problem of effective delivery to target zones. By using the gene sequences of miRNAs as biomarkers, an AD patient's genomic sequence can be mapped, marking which areas require regulation. This process is necessary because of the inter-individuality of miRNA regulation between each case of AD. Also, the problem of some anti-miRNA molecules not being able to cross the blood brain barrier needs to be addressed using a novel transport mechanism, as direct brain injections are not feasible. The simplest, and highly effective, therapeutic avenue is a healthy lifestyle. Daily exercise and proper nutrition hinder inflammatory process and oxidative stress and can prevent progression of AD through allowing higher brain perfusion for cognitive functioning.

Alzheimer's Disease (AD) is the most common cause of dementia in elderly people. Although the exact pathogenesis of AD remains unclear, accumulation of β- amyloid (Aβ) plaques seems to be among the causative events. In view of this, Aβ- PET imaging is considered to be a powerful non-invasive diagnostic tool that could also contribute to the development of therapies by monitoring responses. However, Aβ-PET ligands approved so far can only detect heavy plaque load and cannot replace post-mortem examination of brain tissue. The aim of this multidisciplinary study was to develop structurally novel PET tracers for AD. We focused on barbiturates for two main reasons: (i) barbiturates have an excellent ability to cross the blood-brain-barrier, (ii) they are chelators of cations involved in AD. A group of seven “cold” fluorinated barbiturates, along with the corresponding precursors for the “hot” radiosyntheses, was designed and synthesised. All the experimental logP values fell into the optimum range for brain uptake. Barbiturate 1a (Figure I) was selected for further investigations. Upon assessment of its affinity and specificity for Aβ, the radiosynthesis of [18F]1a was optimised. The imaging potential of this tracer was investigated in vivo in pre-clinical mouse models of AD. Brain PET/CT scans with [18F]1a showed reproducible brain uptake and clearance in three different mouse genotypes (WT, APP/PS1 and PLB2-Tau). The significantly higher uptake observed in APP/PS1 mice provided evidence for (i) the in vivo targeting of Aβ- plaques and (ii) the specificity of the tracer towards Aβ pathology. Finally, we designed a second-generation of barbiturates incorporating stilbene groups as dual metal/Ab targeting tracers and we developed a partial synthesis. With this study we paved the way for a larger scale research endeavour that may ultimately result in the rational design of an optimised lead tracer with the potential to ultimately translate into clinical use.

No description available.

published_or_final_version / Physiology / Doctoral / Doctor of Philosophy

published_or_final_version / Biochemistry / Doctoral / Doctor of Philosophy

This thesis focused on the physiological impact of uncertainty on caregivers of Alzheimer's disease victims. A convenience sample of 30 subjects was used. The uncertainty level was assessed using Parent's Perception of Uncertainty in Illness Scale. Physiological arousal was determined by assaying urinary cortisol and catecholamine levels. The results of the study showed that uncertainty and physiological stress were inversely related. This led to the conclusion that uncertainty was beneficial in that it offered a degree of hope. Knowledge of the disease process increased the stress perceived due to the devastation of Alzheimer's disease and its incurable state. Younger individuals had higher physiological stress than older individuals for comparable amounts of uncertainty. Multiple reasons for this finding are postulated. They include the thought that the younger caregivers may fear developing the disease. It also may be that younger individuals need certainty about the future.