Tal Nuriel, Ph.D. - US grants

[unreadable] DESCRIPTION (provided by applicant): Alzheimer's disease is a serious neurodegenerative illness with a cruel manifestation and few treatment options. Although much has been uncovered about the pathogenesis of familial Alzheimer's disease, little is known about the mechanisms that bring about the sporadic form of the disease that affects about 95% of Alzheimer's patients. Specifically, how is it that environmental and secondary genetic cues are able to trigger the same pathophysiologic events that occur when the direct proteins involved in these events become mutated? In this proposal, I hypothesize that oxidative stress, and specifically nitric oxide, may be capable of exacerbating and perhaps even triggering the same overproduction of amyloid-beta that is seen in familial Alzheimer's disease. Preliminary data shows that gamma-secretase, one of the proteases responsible for amyloid-beta production, is susceptible to nitric oxide-induced S-nitrosylation, and that nitric oxide is capable of increasing gamma-secretase activity. These results suggests a simple but powerful mechanism for how the oxidative stress seen in many conditions that promote sporadic Alzheimer's disease may directly increase amyloid-beta production. In order to further investigate this possibility, I have proposed a series of experiments that will help to elucidate as well as characterize the role of gammasecretase S-nitrosylation on the overall production of amyloid-beta. Using multiple gamma-secretase activity assays, these experiments will test the ability of S-nitrosylation to stimulate gamma-secretase activity. In addition, the specific mechanism of gamma-secretase S-nitrosylation will be determined using both exogenous and endogenous sources of nitric oxide. If the results of these experiments show that gamma-secretase S-nitrosylation plays a significant role in the production of amyloid-beta, this would be a major discovery in the Alzheimer's field. Furthermore, it would have significant implications for possible new therapeutic methods for treating and perhaps even preventing the disease. Alzheimer's disease is a neurodegenerative illness that affects about 24 million people across the globe. This proposal presents a novel idea about what may be triggering Alzheimer's disease, suggesting that Alzheimer's may be caused by increased oxidative stress, which may be capable of modifying an important player in the disease. The results of this research could lead to new ways of treating Alzheimer's disease, including new, more specific forms of antioxidant therapy. [unreadable] [unreadable] [unreadable]

DESCRIPTION (provided by applicant): Carriers of the apolipoprotein E (APOE) ?4 gene are at significantly increased risk for developing Alzheimer's disease (AD). Although numerous theories have been proposed, the cause of this association remains unclear. The most widely accepted view is that the accelerated AD pathology observed in APOE ?4 carriers is due to a decreased ability of the apoE4 protein to clear A? from the brain. However, possession of the APOE ?4 gene also results in a number of other neurological deficits unrelated to A? clearance, including alterations in neuronal structure, thinner entorhinal cortex (EC) layers and poorer outcomes after stroke, suggesting that there may be other mechanisms involved in this process. In order to gain a more comprehensive understanding of the how the expression of different apoE isoforms affects the brain and how this may impact the risk of developing AD and other age-related neurological illnesses, we propose to use mass spectrometry to identify lipids and small-molecules whose levels are affected by changes in apoE isoform expression. To accomplish this, we will first extract lipids, small-molecules and proteins from pathology-free EC and primary visual cortex (PVC) tissues obtained from 14-month old mice and postmortem 19-55 year old individuals expressing differing apoE isoforms. The lipid and small-molecule fractions from these extracts will then be used to perform targeted lipidomics and untargeted metabolomics, followed by bioinformatic analysis and a variety of validation experiments, in order to determine the specific lipids, small-molecules and metabolic pathways that are affected by alternative apoE isoform expression in the brain. Thus far, preliminary studies have uncovered significant changes in energy metabolism pathways and several important lipid subclasses, demonstrating the power of these techniques for discovering previously unknown isoform-specific effects of apoE in the brain. We expect that the full study proposed herein will uncover further apoE isoform-specific changes in lipid and small-molecule levels and will lead to a greater understanding of how apoE4 influences AD pathology, potentially leading to new therapeutic strategies for AD and other neurological illnesses influenced by apoE4 expression.

Project Summary Possession of the ?4 allele of apolipoprotein E (APOE) is the major genetic risk factor for late onset Alzheimer?s disease (AD), although the direct cause remains a source of debate. While research suggests that APOE4 expression can increase the phosphorylation, aggregation and neurotoxicity of tau in the brain, it has yet to be investigated whether APOE4 can accelerate the propagation of tau through the brain, a process that is highly correlated with the cognitive decline of AD patients. Preliminary work that I have recently published shows that APOE4 expression in mice leads to an increase in brain activity in the entorhinal cortex (EC), one of the first brain regions affected by tau pathology during AD. This finding, along with additional data showing that neuronal hyperactivity can increase tau release and propagation, suggests that this APOE4-associated hyperactivity that I have observed may accelerate the propagation of tauopathy out of the EC and into neighboring brain regions. To test this hypothesis, I will utilize a novel mouse line in which human APOE3 and APOE4 mice are crossed with EC-Tau mice, which predominantly expresses tau in the EC. Preliminary data on these APOE/EC-Tau mice also suggests that this mouse model may be a useful tool for studying the ability of APOE4 to accelerate tauopathy-induced neurodegeneration in an AD-relevant brain region. Through a series of studies on these APOE/EC-Tau mice, I will elucidate the ability of APOE4 to both accelerate tauopathy propagation through the brain, as well as to increase tauopathy-induced neurodegeneration in the EC. As both APOE4 and tau significantly impact AD, elucidating the link between them would represent a major breakthrough in the field and has the potential to lead to novel therapeutic strategies for preventing or treating AD, especially among APOE4 carriers.

Project Summary Possession of the ?4 allele of apolipoprotein E (APOE) is a major risk factor for late onset Alzheimer's disease (AD), although the direct cause remains a source of debate. Recent data has shown that the APOE gene is upregulated in a novel microglial activation state found in AD and other neurodegenerative diseases termed disease associated microglia (DAM). Furthermore, APOE4 expression has been shown to induce a neurotoxic activation of microglia in the presence of various stimuli, including tauopathy-afflicted neurons, although the mechanism of this activation has yet to be elucidated. In order to discover the role and mechanism of APOE4- associated microglial activation and its resulting neurotoxicity of tauopathy-afflicted neurons, I have devised an innovative set of experiments that utilize both cutting-edge technology and novel resources. In Aim 1, single- cell RNA-sequencing will be performed on microglia purified from a novel mouse line in which human APOE3 and APOE4 mice are crossed with EC-Tau mice, which express a pro-aggregating mutant tau protein primarily in the entorhinal cortex (EC). In Aim 2, APOE4's effects on microglial activation will be investigated in the setting of human microglia, using both a newly created isogenic APOE human stem cell line and powerful bioinformatics analyses. And in Aim 3, a robust in vitro co-culture assay will be utilized in order to both uncover the mechanism responsible for APOE4-associated microglial activation and the resulting neurotoxicity and to identify novel therapeutic strategies for inhibiting these events. As both APOE4 and microglial activation significantly impact AD, discovery of the mechanistic link between these two important players and therapeutic strategies for modulating APOE4-associated microglial activation would represent a major breakthrough in the AD field and would greatly advance our goal of preventing or slowing the progression of AD, especially among APOE4 carriers.

Project Summary Possession of the ?4 allele of apolipoprotein E (APOE) is the major genetic risk factor for late onset Alzheimer?s disease (AD), although the direct cause remains a source of debate. While much research has focused on the association between APOE4 and A? pathology, there is significant evidence that APOE4 expression effects a wide-array of important pathways in the brain that are independent of A?, including my own work showing the effects of APOE4 expression on neuronal hyperactivity, endosomal-lysosomal dysregulation and bioenergetics deficits in AD-vulnerable brain regions. In order to expand on this work and to gain a more comprehensive picture of APOE4?s effects in individual cell populations in both mice and humans, I propose to perform single-nucleus RNA-sequencing on entorhinal cortex (EC) tissues from middle-aged mice and humans expressing APOE4 vs. APOE3. Data generated from this sequencing strategy will be robustly analyzed using modern bioinformatics techniques, and gene and pathway hits will be further validating using molecular biology approaches. As APOE4 is the most important genetic determinant of late-onset AD, discovery of cell-type specific effects of APOE4 on previously identified or novel pathways will greatly increase our knowledge of how late-onset AD develops and has the potential to uncover new therapeutic strategies for preventing or treating AD, especially among APOE4 carriers.

Project Summary Carriers of the apolipoprotein E (APOE) ?4 gene are at a significantly increased risk for developing Alzheimer?s disease (AD). Although numerous theories have been proposed, the cause of this association remains unclear. My own research has uncovered novel effects of APOE4 expression on important processes in the brain, including neuronal activity, the endosomal-lysosomal system and bioenergetic regulation. However, substantial questions remain about when, where and how these systems are effected by differential APOE isoform expression. In order to answer these questions and gain a more comprehensive understanding of how differential APOE isoform expression affects vital brain processes and pathways, I propose a series of cutting-edge experiments, performed on a newly created APOE mouse model. By conducting behavioral experiments, histological examinations, imaging and an array of spatial multi-omics experiments, this project aims to define the temporal, spatial and cellular progression of differential APOE isoform effects in the brain. Each of these experiments will be performed on young (4-6 month-old), aged (14-16 month-old), and old (24-26 month-old) APOE2, APOE3 and APOE4-KI mice. In Aim 1, we will conduct a series of behavioral tests, including Barnes maze, novel object recognition, and fear conditioning, as well as a histological analysis for endosomal-lysosomal disruptions, bioenergetic deficits, and changes in AD pathology markers. We will also conduct a detailed imaging analysis using fMRI to observe activity and structural changes in these mice. In Aim 2, we will conduct an in- depth spatial multi-omics analysis on these mice, including spatial transcriptomics and spatial metabolomics/lipidomics. And in Aim 3, we will explore the cellular contributions to differential APOE isoform expression, including a novel bioinformatics approach and conditionally knockout of APOE from astrocytes and microglia in the APOE-KI mice. We anticipate that the full study proposed herein will uncover important APOE isoform effects on multiple brain processes and pathways in a systems-biology manner, which will dramatically increase our understanding of how APOE isoform differences affect an individual?s susceptibility to AD, potentially leading to new therapeutic strategies for AD, especially among APOE4 carriers.