This research involves an examination of the olfactory ability of individuals with Frontotemporal Dementia, Alzheimer's disease, and geriatric individuals with cognitive complaints owing to Major Depressive Disorder or Generalized Anxiety Disorder. The purpose of this study was to determine if olfactory differences were useful in differentiating between demented and non-demented individuals. Due to the pathway of the olfactory tract, it can be expected that there would be equal olfactory deficits in those diagnosed with Frontotemporal Dementia and Alzheimer's disease, and that these deficits would be worse than those found in geriatric individuals with Major Depressive Disorder or Generalized Anxiety Disorder. Five hypotheses were investigated. The first utilized an ANCOVA and found that the olfaction of the demented group was worse than that of the non-demented, psychiatric group. The second utilized a series of Discriminant Function Analyses and F-tests and determined that olfaction best classified demented and non-demented individuals. The third implemented a Mixed Model ANOVA to assess for lateralized smell deficits within the demented group and between the demented and non-demented groups and found no main effects of lateralization or interaction effects. The fourth hypothesis investigated the relationship between smell and commonly used neuropsychological tests in a Frontotemporal Dementia sample, and found that there was no difference between the relationship of tests measuring the frontal and temporal lobes to those measuring other cerebral areas. The fifth hypothesis replicated Hypothesis 4 in an Alzheimer's disease sample and found that there was a significant difference between the relationship of tests measuring the frontal and temporal lobes to those measuring other cerebral areas. Primarily, this study demonstrated that dementia patients present with significantly more olfactory deficits than a psychiatric sample with subjective cognitive complaints. Additionally, olfaction correctly distinguished the Dementia Group from the Psychiatric Group with 81.6% accuracy, 90.2% sensitivity and 67.6% specificity. Alone, these classification statistics are quite impressive, appearing equivalent, or possibly superior to the classification statistics of commonly used neuropsychological tests of memory, executive functioning, and visuospatial ability. This study concluded that adding an olfactory measure to an assessment battery provides clinically relevant data, and enhances the diagnostic accuracy of the battery. However, though this study found the Alberta Smell Test was a valuable addition to a test battery, the absence of lateralized findings demonstrated the unirhinal format, by which the Alberta Smell Test is administered, does not provide diagnostically relevant information above and beyond the information a clinician will gain from birhinal assessment.

This study examined the relationship between five neurocognitive domains, including memory, visual-spatial skills, executive functioning, language, and attention, and three instrumental activities of daily living (IADLs), (financial management, medication management, and driving), as well as three basic activities of daily living (ADLs), including dressing, feeding, and grooming. Subjects were community dwelling older adults diagnosed with Alzheimer's Disease (AD) ranging in severity from mild to severe. Correlation and regression models were used to analyze the relationship between neurocognitive domains and self-care activities. All of the neurocognitive domains were significantly but mildly correlated with declines in IADLs, and all of the neurocognitive domains, except memory, were significantly but mildly correlated with ADL decline. None of the regression models, except driving, produced significant results. The hypotheses were based on the belief that overlap occurs between declines in cognitive domains and self-care skills. The results of this study support the assertion that global cognitive decline occurs followed by declines in IADLs and later declines in ADLs. Thus, instead of specific neurocognitive domains predicting decline in individual self-care skills, this research indicates that the degree of severity of cognitive decline is predictive of impairments in IADLs or ADLs with milder global impairment predicting IADLs and more pronounced decline predicting declines in ADLs. Degree of severity may have been more predictive because while individuals vary in cognitive symptom presentation, the course of AD always progresses from mild to more severe. The role of over-learning also potentially impacted the results of the study. Individuals learn and daily practice ADLs at a young age causing them to require less higher level cognitive skills (over-learned). Since over-learned IADLs require less higher level cognition, they were preserved longer in AD course, thus declining after global cognitive impairment. Clinicians need to thoroughly assess the degree to which an individual has over-learned a task in order to make the most accurate recommendations. If an individual has milder global decline, IADLs are likely to be impaired, and if an individual has moderate global decline with memory reaching a floor, ADLs are likely to be impaired.

The search for reliable early indicators of age-related cognitive decline represents an important avenue in aging research. Most research on late-life development charts cognitive change as a function of chronological age (CA), however, although CA is a commonly used developmental index, it offers little insight into the mechanisms underlying cognitive decline. In contrast, biological age (BioAge), reflecting the vitality of essential biological processes, represents a promising operationalization of developmental time. My overall programmatic doctoral research interests involve the identification of biological risk factors that predict age-related cognitive decline, impairment and dementia. In this dissertation document, I present: an overview of my empirical contributions to the BioAge and cognitive aging literature throughout my doctoral training; the dissertation project which uses preliminary data from the PREVENT study and provides evidence that elevated plasma pro-inflammatory proteins are associated with cognitive status (healthy controls (HC) vs Alzheimer’s disease dementia (AD)), cognitive performance and are related to poorer cognitive performance in amnestic mild cognitive impairment (a-MCI); and a discussion on the broad implications of the project results and future directions in BioAge research. / Graduate

Alzheimer's disease (AD) is the most common cause of a progressive decline of cognitive function in aged humans. It is thought to be a result of the formation of amyloid plaques in the brain that are largely composed of β-amyloid (Aβ), which is one of the cleavage products of the amyloid precursor protein (APP). The amyloidogenic processing of APP to produce the Aβ peptides requires sequential proteolytic cleavages by the β- and γ-secretases. APP is first cleaved by the β-secretase to produce APP-C99, and this product is a substrate for further processing by the γ-secretase that cleaves within its transmembrane domain to produce N-terminal Aβ peptides and the C-terminal APP intracellular domain (AICD). On the basis of similarities to the Notch processing pathway, it has been postulated that the AICD may play a role in gene regulation following its release in response to some form of extracellular signal. In order to better understand the production and fate of the AICD, I have investigated the potential for exploiting a cell-free system to study its generation and properties. Having generated a number of model APP-derived fragments and shown them to be efficiently membrane integrated in vitro, I went on to study AICD production. I discovered that AICD-like fragments are extremely labile when synthesised in a rabbit reticulocyte lysate system and are rapidly degraded via a metalloproteinase, most likely the insulin degrading enzyme (IDE). The in vitro stability of these model AICD-like fragments was dependent upon the precise chain length of the polypeptide and N-terminal processing may preface the activity of IDE in vitro. The rapid degradation of the AICD in vitro is in close agreement with previous in vivo studies, and taken together such data are consistent with a role for the AICD in a signalling pathway of some form. A variety of approaches were also taken to try to generate the AICD by the γ-secretase mediated cleavage of the APP-C99 fragment, a biologically relevant substrate. In no case was any evidence of such cleavage observed in vitro and hence I conclude that the endoplasmic reticulum does not possess an active form of the γ-secretase. Preliminary in vivo-based studies did provide evidence for the γ-secretase cleavage of APP-C99 fragments, consistent with current models implying that such processing takes place at the cell surface and/or in endosomes and not at the endoplasmic reticulum.

published_or_final_version / Physiology / Doctoral / Doctor of Philosophy

published_or_final_version / abstract / Anatomy / Doctoral / Doctor of Philosophy

published_or_final_version / abstract / Electrical and Electronic Engineering / Doctoral / Doctor of Philosophy

published_or_final_version / Anatomy / Doctoral / Doctor of Philosophy

The development of a highly sensitive ruthenium-based fluorescent staining solution isdescribed in this dissertation. The in-house synthesized ruthenium complex (RuMS)containing both sulfonated and non-sulfonated ligand has detection limit of 1 ng ofprotein that is better than colloidal coomassie, silver and ruthenium complex containingall sulfonated ligands (RuBPS). RuMS stain has 100-fold dynamic range and does notinterfere with subsequent mass spectral identification of proteins. The capability of inhousesynthesis of the staining solution makes it a viable cost-effective alternative to theexpensive commercially available fluorescent stain, Sypro Ruby. The low detection limit,broad linear dynamic range and compatibility with mass spectrometry, make thedevelopment of this stain a worthwhile pursuit. The staining solution was utilized insubsequent applications of two-dimensional gel electrophoresis (2-DE) technology.Proteomics methodology utilizing 2-DE and mass spectrometry was applied toinvestigate the effect of malathion on the proteome of human neuroblastoma cells.Results indicated that out of 122 proteins that were identified from the neuroblastomaproteome, sixteen proteins were down-regulated while five proteins were significantlyup-regulated after treatment with malathion. Significant down-regulation of calciummodulators like calmodulin and calgizarrin and other key chaperones makes themalathion-treated cells highly prone to oxidative stress. With increased awareness inpesticide related adverse effects, identification of altered proteins in malathion-treatedhuman neuroblastoma cells is a critical finding.Proteomics is a major area of research in the identification of biomarkers for diseases. Anovel immunoprecipitation method developed in this work allowed for successfulisolation and identification of albumin-interactome in cerebrospinal fluid (CSF) that isusually under-represented in standard CSF analysis using 2-DE. A key finding is thedifferential expression of various isoforms of proteins in CSF albumin-interactome fromAlzheimer's disease (AD) subjects. The data implicate the acidic isoform ofprostaglandin D2 synthase (PGDS2) as a potential biomarker for AD. An understandingof the differential expression of these protein isoforms in AD will provide insight into theetiology of the disease and this can have far-reaching implication on drug developmentleading to the cure or even preventation of the disease.

Alzheimer’s disease (AD) is a multifactorial, progressive, age-related neurodegenerative disease. Oxidative stress hypothesis is most prevalent and is gaining significant support. Inspite of the progress achieved on oxidative stress related damages in AD brain; the modification occurring on the various cellular antioxidant enzymes antioxidant has not been identified. Tyrosine nitration, a marker for peroxynitrite induced oxidative damage to protein is widespread in AD brain and Manganese superoxide dismutase (MnSOD), primary mitochondrial antioxidant enzyme is prone to peroxynitrite induced nitration and inactivation. Nitration of proteins involved in energy metabolism has been demonstrated in AD brain, which may explain the altered glucose metabolic status existing in AD brain. In the present study, we investigated the effect of tyrosine nitration of MnSOD on energy metabolism by the use of AD mouse model and cultured neuronal cells. The AD mouse model was generated from a double homozygous knock-in mouse, designated as APP/PS-1 mice, by incorporating the Swedish familial AD mutations in APP and P264L familial AD mutation in PS – 1. These animals develop age dependent increase in Aβ deposition beginning at 6 months along with an increase in insoluble Aβ1-40/Aβ1-42 levels. Genotype and age associated increase in nitration of MnSOD without any change in protein levels was also observed. MnSOD activity and mitochondrial respiration was decreased in APP/PS-1 mice. There was also concomitant increase in levels of lactate, an index of glycolytic activity in APP/PS-1 mice. To directly investigate the role of MnSOD inactivation in mitochondrial function and subsequent alteration in glycolytic activity, SH-SY5Y neuroblastoma cells line was used and treated with peroxynitrite. Enhanced nitration and reduction in the activity of MnSOD was observed upon peroxynitrite treatment. Peroxynitrite treatment also induced mitochondrial dysfunction, but MnSOD was inactivated at a concentration of peroxynitrite 10 times lower than that required to inhibit mitochondrial respiration. Mitochondrial dysfunction was alleviated by SOD mimetic and reproduced by MnSOD siRNA. The decline in mitochondrial function did not result in decreased ATP levels but was accompanied by an up-regulated glycolysis signified by high levels of lactate and lactate dehydrogenase activity but decreased activity of pyruvate dehydrogenase. These changes were prevented by SOD mimetic and were promoted by MnSOD siRNA. Specific reduction of MnSOD in MnSOD heterozygous knock-out mice resulted in decreased RCR and complex I activity with increased lactate levels. Taken together, these data demonstrate a critical role of MnSOD in influencing the mitochondrial function and thereby the switch in the energy metabolism switch that might occur under the pathological condition of MnSOD deficiency.