Alzheimer’s disease (AD) is the leading cause of dementia among the elderly and sixth leading cause of death in the US. No effective therapeutic intervention is currently available to treat, cure or halt this devastating disease. The prevalence of AD is expected to rise rapidly in coming years and is predicted to overwhelm our socioeconomic reserves. Thus, unveiling critical pathogenic mechanisms of the disease and risk factors that modulate the disease progression and onset is an urgent matter to slow this upswell. While aging and genetics are unequivocal risk factors for AD, there is a growing evidence supporting the pivotal role of environmental or modifiable factors in contributing the onset of AD. Among them, air pollution has recently been highlighted as a prominent culprit accelerating cognitive decline and increasing a risk for AD. However, little work has been done to elucidate its underlying molecular mechanisms by which exposure to air pollution contributes to the pathological development of AD. The purpose of this dissertation is to investigate the neurotoxic effects of airborne particulates in a humanized amyloid mouse model of AD. Previous reports indicate that PM exposure in non-AD model mice increases inflammatory markers such as cytokines, glial activation, and oxidative stress in the CNS and may increase expression of native mouse amyloid or alter its processing. Thus, we hypothesize that exposure to ultra-fine particulates exacerbates memory impairment by increasing neuroinflammation and amyloid β burden in a humanized amyloid mouse model of AD. To test this, we exposure wild-type and AppNL-G-F/+-KI mice to concentrated ultra-fine particulate matter both in utero and at adult ages. We show exposure dependent decreases in memory behavior tasks, as well as increases in Aβ plaque burden in the AppNL-G-F/+-KI model. However, memory impairment occurs independently of plaque burden increase, and only in the in utero exposure group were neuroinflammatory markers increased. In the adult exposure, we instead saw a decrease in the astrocytic glutamate transporter GLT-1. Previous work has shown a link between glutamate transporter loss and tau pathology, indicating a potential amyloid independent pathway linking particulate exposure and AD pathology. This research provides evidence of Aβ plaque burden increase dependent only on particulate exposure, as well as evidence that exposure during development impacts AD neuropathology in adult mice.

Exposure to elevated levels of ambient air pollution particulate matter (PM), is associated with cognitive decline. In this study we examined the impacts of different sizes of PM (concentrated coarse (Dp ≤ 10 µm), concentrated fine (Dp ≤ 2.5 µm), and ultrafine PM (Dp ≤ 200 nm)) on memory in mice using two standard behavioral tests; the object location test (OLM) and the novel object recognition test (ORM). Secondary variables that were examined via behavioral testing included: sex (M vs. F), diet (high fat vs. normal chow diet), and mouse genotype. We used C57BL/6 mice from 2 studies, CEDARS (3xTg-AD mice) which were genetically modified to develop Alzheimer’s Disease, and RESTORE (ApoE KO mice and Ldlr KO mice) which can be hyperlipedemic and develop arterial plaques. These models were chosen to examine the potential impacts of ambient PM on neurodegenerative changes in exposed normal and compromised mice. Both studies were examined individually with respect to impact of PM exposure on behavioral test outcomes and were then compared with respect to the secondary independent variables, genetic modification and sex. There were no statistically significant changes in the Cedars study as a function of PM size. There were, however, statistically significant changes in OLM and ORM memory tasks based on exposure length, sex, genetic predisposition, and diet. The data suggests that these factors can interact with environmental PM exposure to impact memory performance. This analysis did have some limitations including; multiple people analyzed the behavior data, which could have increased variability in the results and there were losses of animals in some groups due to attrition, which reduced the power of the statistical analyses.

Hepatitis C virus (HCV) is a major health problem which is affecting more than 170 million people worldwide. About 80% of HCV-infected patients develop chronic infections because they cannot clear the virus. Chronic infections increase the risks of progressing to severe liver diseases such as cirrhosis and hepatocellular carcinoma. HCV has several strategies to evade host interferon (IFN) responses. In this study, we hypothesize that a novel HCV viral protein, frameshift/alternate reading frame protein (F/ARFP) from HCV -2/+1 frame, contributes to the suppression of retinoic-acid-inducible gene-I (RIG-I)-mediated IFN induction. We found that F/ARFP suppressed type I IFN induction stimulated by HCV RNA pathogen-associated molecular pattern (PAMP) and synthetic double stranded RNA poly(IC). This suppression occurred independently of other HCV viral factors such as nonstructural protein NS3/4A. Point mutations in the full-length HCV sequence for introducing premature termination codons in the -2/+1 frame enhanced type I IFN induction. Taken together, we suggest that HCV F/ARFP suppresses type I IFN responses mediated by RIG-I. Additionally, we investigated the role of NAD(P)H oxidases (Noxs) in type I IFN pathway. Nox family enzymes function as important sources of regulated production of reactive oxygen species in cell signaling and regulation of gene expression. However, much remains to be investigated how Nox enzymes contribute to the immune response against virus infection. In this study, among seven members of Nox family, we hypothesize that Nox1 has a role in type I IFN pathway. We observed that the mRNA levels of Nox enzymes including Nox1, Nox4, Nox5, Duox1, and Duox2 were elevated by poly(IC). We showed that poly(IC)-triggered IFNβ1 mRNA induction was suppressed by DPI, a flavoprotein inhibitor, in Huh7 hepatoma cells. Further study suggested that Nox1 siRNA-transfected Huh7 cells decreased poly(IC)-stimulated IFNβ1 mRNA induction in Huh7 cells. Moreover, HCV replication was increased by Nox1 and Nox4 knockdown by corresponding siRNAs. Taken together, these data suggest that Nox1 may modulate type I IFN pathway. Therefore, our study may provide new insights into how HCV evades the host innate immune response and how Nox enzymes function as antiviral agents and points to possible new drug target for therapy.

Chronic exposure to copper is a putative environmental risk factor for AD. Although copper is an essential metal, environmental exposure to its inorganic cupric form (Cu2+) may exert toxicity. However, exact neurotoxic mechanisms of action of copper, and its contribution to AD neuropathology remain largely unelucidated. Copper is a natural antimicrobial agent, and here, we investigate its impact of copper exposure on the gut microbiome and integrity as well as effects on cognition and AD pathology. Recent studies implicate gut dysbiosis in the onset and progression of AD in both humans and animal models. We believe that chronic exposure to environmentally relevant dose of copper through drinking water will reduce richness and diversity of the gut microbiota in both WT and hAPPNL-G-F knock-in (APP-KI) mice. Thus, we hypothesize that this copper-induced dysbiosis may perturb host metabolism and inflammatory homeostasis to contribute to AD neuropathology. In this dissertation, male and female APP-KI and wildtype C57BL/6J mice were exposed to 1.3 ppm CuCl2 in drinking water ad-libitum for 3(pilot study) and 9(chronic study) months. The present studies use a host of behavioral, immunohistochemical, biochemical and microbial analyses to investigate the effect of copper on microbiota, amyloidosis, neuroinflammation and cognition in AD mice.

In our pilot study, we discovered that while there were no significant changes in AD neuropathology or inflammatory status in WT or APP-KI mice, we observed that WT mice microbiota were resilient to the effects of copper administration. However, beginning 1 month after initiation of exposure, APP-KI mice developed a significantly different microbial composition according to the Bray Curtis -diversity. This was largely driven by a decrease in Firmicutes, and specifically in the genus Allobaculum and an increase in the phylum Bacteroidetes, specifically in the genus S24-7. These changes mirror some of the changes observed in studies in other AD mouse models and AD patients.

In our chronic study, spanning from after weaning and until the mice were 10 months, old, we discovered no significant changes in cognition and a mild change in the cytokine profile of AD mice, with a reduction of IL-4 in plasma, following copper administration. With this longer treatment paradigm, we did observe increased in amyloidosis, both dense core and diffuse plaques, with copper treatment. Further, along with evidence that plaque-associated microglia are of a diseased phenotype, the amount of these microglia was significantly increased with copper treatment indicating that copper perturbs neuroinflammation. In the gut microbiota, we observe unexpected results. Copper directs the developmental trajectory of WT mice but with time the microbiota of AD mice is not changed much. This may indicate differing windows of susceptibility to copper exposure between the genotypes of mice and between pilot study and the chronic study paradigms.

Cell to cell communication is important in an array of biological systems including inflammasome activation and cellular fusion. This process of communication leads to multinucleated giant cell (MGC) formation including: macrophage fusion, osteoclasts and foreign body cells (5, 35). Chronic inflammation contributes to multinucleated cells forming granulomas at sites of infection (5). The specific mechanism of membrane fusion has yet to be understood. Identified modulators of membrane fusion include the hemichannel pannexin-1 in association with purinergic receptor, P2X7 (20). Preliminary results on another P2X receptor, P2X4, have indicated its possible association in membrane fusion and Chlamydia (C. trachomatis) L2 vacuole fusion. Our objective is to characterize P2X4's role in the mechanism of cellular fusion and L2 vacuole fusion. We will also investigate the interaction of P2X4 with pannexin-1 in both membrane fusion processes. These studies will contribute to the understanding of the role of P2X4 in inflammatory and autoimmune diseases.