In Noori et al. [1], we hypothesized that there is a shared gene expression signature underlying neurodegenerative proteinopathies including Alzheimer's disease (AD), Lewy body diseases (LBD), and the amyotrophic lateral sclerosis and frontotemporal dementia (ALS-FTD) spectrum. To test this hypothesis, we performed a systematic review and meta-analysis of 60 human central nervous system transcriptomic datasets in the public Gene Expression Omnibus and ArrayExpress repositories, comprising a total of 2,600 AD, LBD, and ALS-FTD patients and age-matched controls which passed our stringent quality control pipeline. Here, we provide the results of differential expression analyses with data quality reports for each of these 60 datasets. This atlas of differential expression across AD, LBD, and ALS-FTD may guide future work to elucidate the pathophysiological drivers of these individual diseases as well as the common substrate of neurodegeneration.

Alzheimer's disease (AD) is defined by the presence of abundant amyloid-β (Aβ) and tau neuropathology. While this neuropathology is necessary for AD diagnosis, it is not sufficient for causing cognitive impairment. Up to one third of community dwelling older adults harbor intermediate to high levels of AD neuropathology at death yet demonstrate no significant cognitive impairment. Conversely, there are individuals who exhibit dementia with no gross explanatory neuropathology. In prior studies, synapse loss correlated with cognitive impairment. To understand how synaptic composition changes in relation to neuropathology and cognition, multiplexed liquid chromatography mass-spectrometry was used to quantify enriched synaptic proteins from the parietal association cortex of 100 subjects with contrasting levels of AD pathology and cognitive performance. 123 unique proteins were significantly associated with diagnostic category. Functional analysis showed enrichment of serotonin release and oxidative phosphorylation categories in normal (cognitively unimpaired, low neuropathology) and "resilient" (unimpaired despite AD pathology) individuals. In contrast, frail individuals, (low pathology, impaired cognition) showed a metabolic shift towards glycolysis and increased presence of proteasome subunits.

Metformin, a diabetes drug with anti-aging cellular responses, has complex actions that may alter dementia onset. Mixed results are emerging from prior observational studies. To address this complexity, we deploy a causal inference approach accounting for the competing risk of death in emulated clinical trials using two distinct electronic health record systems. In intention-to-treat analyses, metformin use associates with lower hazard of all-cause mortality and lower cause-specific hazard of dementia onset, after accounting for prolonged survival, relative to sulfonylureas. In parallel systems pharmacology studies, the expression of two AD-related proteins, APOE and SPP1, was suppressed by pharmacologic concentrations of metformin in differentiated human neural cells, relative to a sulfonylurea. Together, our findings suggest that metformin might reduce the risk of dementia in diabetes patients through mechanisms beyond glycemic control, and that SPP1 is a candidate biomarker for metformin's action in the brain.

Omics studies have revealed that various brain cell types undergo profound molecular changes in Alzheimer's disease (AD) but the spatial relationships with plaques and tangles and APOE-linked differences remain unclear.

Insoluble fibrillar tau, the primary constituent of neurofibrillary tangles, has traditionally been thought to be the biologically active, toxic form of tau mediating neurodegeneration in Alzheimer's disease. More recent studies have implicated soluble oligomeric tau species, referred to as high molecular weight (HMW), due to their properties on size-exclusion chromatography, in tau propagation across neural systems. These two forms of tau have never been directly compared. We prepared sarkosyl-insoluble and HMW tau from the frontal cortex of Alzheimer patients and compared their properties using a variety of biophysical and bioactivity assays. Sarkosyl-insoluble fibrillar tau comprises abundant paired-helical filaments (PHF) as quantified by electron microscopy (EM) and is more resistant to proteinase K, compared to HMW tau, which is mostly in an oligomeric form. Sarkosyl-insoluble and HMW tau are nearly equivalent in potency in HEK cell bioactivity assay for seeding aggregates, and their injection reveals similar local uptake into hippocampal neurons in PS19 Tau transgenic mice. However, the HMW preparation appears to be far more potent in inducing a glial response including Clec7a-positive rod microglia in the absence of neurodegeneration or synapse loss and promotes more rapid propagation of misfolded tau to distal, anatomically connected regions, such as entorhinal and perirhinal cortices. These data suggest that soluble HMW tau has similar properties to fibrillar sarkosyl-insoluble tau with regard to tau seeding potential, but may be equal or even more bioactive with respect to propagation across neural systems and activation of glial responses, both relevant to tau-related Alzheimer phenotypes.

The prion-like propagation of tau in neurodegenerative disorders implies that misfolded pathological tau can recruit the normal protein and template its aggregation. Here, we report the methods for the development of sensitive biosensor cell lines for the detection of tau seeding activity.

Inflammation has been implicated in the pathogenesis of Alzheimer's disease (AD) and other neurodegenerative diseases. We have examined the potential role of some chemokine/chemokine receptors in this process. It is known that CXCR2 is a strongly expressed chemokine receptor on neurons and is strongly upregulated in AD in a subpopulation of neuritic plaques. Here, we show that one of the CXCR2 ligand GROalpha/KC can be a potent trigger for the ERK1/2 and PI-3 kinase pathways, as well as tau hyperphosphorylation in the mouse primary cortical neurons. GROalpha immunoreactivity can be detected in a subpopulation of neurons in normal and AD. Therefore, the CXCR2-ligand pair may have a potent pathophysiological role in neurodegenerative diseases.

Progressive microglial accumulation at amyloid-β (Aβ) plaques is a well-established signature of the pathology of Alzheimer's disease, but how and why microglia accumulate in the vicinity of Aβ plaques is unknown. To understand the distinct roles of Aβ on microglial accumulation, we quantified microglial responses to week-long lasting gradients of soluble Aβ and patterns of surface-bound Aβ in microfluidic chemotaxis platforms. We found that human microglia chemotaxis in gradients of soluble Aβ42 was most effective at two distinct concentrations of 23 pg.mL(-1) and 23 ng.mL(-1) Aβ42 in monomers and oligomers. We uncovered that while the chemotaxis at higher Aβ concentrations was exclusively due to Aβ gradients, chemotaxis at lower concentrations was enhanced by Aβ-induced microglial production of MCP-1. Microglial migration was inhibited by surface-bound Aβ42 in oligomers and fibrils above 45 pg.mm(-2). Better understanding of microglial migration can provide insights into the pathophysiology of senile plaques in AD.

Presenilin-mediated endoproteolysis of transmembrane proteins plays a key role in physiological signaling and in the pathogenesis of Alzheimer disease and some cancers. Numerous inhibitors have been found via library screens, but their structural mechanisms remain unknown. We used several biophysical techniques to investigate the structure of human presenilin complexes and the effects of peptidomimetic γ-secretase inhibitors. The complexes are bilobed. The head contains nicastrin ectodomain. The membrane-embedded base has a central channel and a lateral cleft, which may represent the initial substrate docking site. Inhibitor binding induces widespread structural changes, including rotation of the head and closure of the lateral cleft. These changes block substrate access to the catalytic pocket and inhibit the enzyme. Intriguingly, peptide substrate docking has reciprocal effects on the inhibitor binding site. Similar reciprocal shifts may underlie the mechanisms of other inhibitors and of the "lateral gate" through which substrates access to the catalytic site.

Parkinson's disease is a neurodegenerative disorder characterized by Lewy bodies, a pathological hallmark comprised mostly of aggregated alpha synuclein. Accumulating evidence demonstrates the association of smaller oligomeric aggregates to disease etiology and many therapeutic approaches are aimed at inhibiting and reducing the aggregation process. Molecular chaperones and co-chaperones play a key role in protein homeostasis and have potential as therapeutics to inhibit alpha synuclein associated toxicity. Here we use a gene therapy approach to evaluate the applicability of the Hsp70 co-chaperone CHIP (C-terminal Hsp70 interacting protein) as a therapeutic candidate and examine its direct effect on alpha synuclein aggregates in vivo. Utilizing a novel viral vector mediated rat model to directly detect alpha synuclein aggregates, we show that CHIP can mediate the degradation of alpha synuclein aggregates in vivo. However, our studies also reveal that CHIP may potentially degrade tyrosine hydroxylase which would compromise the applicability of CHIP as a therapeutic approach for Parkinson's disease.

Optical imaging using multiphoton microscopy and whole body near infrared imaging has become a routine part of biomedical research. However, optical imaging methods rely on the availability of either small molecule reporters or genetically encoded fluorescent proteins, which are challenging and time consuming to develop. While directly labeled antibodies can also be used as imaging agents, antibodies are species specific, can typically not be tagged with multiple fluorescent reporters without interfering with target binding, and are bioactive, almost always eliciting a biological response and thereby influencing the process that is being studied. We examined the possibility of developing highly specific and sensitive optical imaging agents using aptamer technology. We developed a fluorescently tagged anti-Aβ RNA aptamer, β55, which binds amyloid plaques in both ex vivo human Alzheimer's disease brain tissue and in vivo APP/PS1 transgenic mice. Diffuse β55 positive halos, attributed to oligomeric Aβ, were observed surrounding the methoxy-XO4 positive plaque cores. Dot blots of synthetic Aβ aggregates provide further evidence that β55 binds both fibrillar and non-fibrillar Aβ. The high binding affinity, the ease of probe development, and the ability to incorporate multiple and multimodal imaging reporters suggest that RNA aptamers may have complementary and perhaps advantageous properties compared to conventional optical imaging probes and reporters.

In Parkinson's disease and other Lewy body disorders, the propagation of pathology has been accredited to the spreading of extracellular α-synuclein (α-syn). Although the pathogenic mechanisms are not fully understood, cell-to-cell transfer of α-syn via exosomes and other extracellular vesicles (EVs) has been reported. Here, we investigated whether altered molecular properties of α-syn can influence the distribution and secretion of α-syn in human neuroblastoma cells. Different α-syn variants, including α-syn:hemi-Venus and disease-causing mutants, were overexpressed and EVs were isolated from the conditioned medium. Of the secreted α-syn, 0.1-2% was associated with vesicles. The major part of EV α-syn was attached to the outer membrane of vesicles, whereas a smaller fraction was found in their lumen. For α-syn expressed with N-terminal hemi-Venus, the relative levels associated with EVs were higher than for WT α-syn. Moreover, such EV-associated α-syn:hemi-Venus species were internalized in recipient cells to a higher degree than the corresponding free-floating forms. Among the disease-causing mutants, A53T α-syn displayed an increased association with EVs. Taken together, our data suggest that α-syn species with presumably lost physiological functions or altered aggregation properties may shift the cellular processing towards vesicular secretion. Our findings thus lend further support to the tenet that EVs can mediate spreading of harmful α-syn species and thereby contribute to the pathology in α-synucleinopathies.

To retrospectively determine the frequency of N-Methyl-D-Aspartate (NMDA) receptor (NMDAR) autoantibodies in patients with different forms of dementia.

Mucolipidosis IV (MLIV) is caused by mutations in the gene MCOLN1. Patients with MLIV have severe neurologic deficits and very little is known about the brain pathology in this lysosomal disease. Using an accurate mouse model of mucolipidosis IV, we observed early behavioral deficits which were accompanied by activation of microglia and astrocytes. The glial activation that persisted during the course of disease was not accompanied by neuronal loss even at the late stage. In vivo [Ca(2+)]-imaging revealed no changes in resting [Ca(2+)] levels in Mcoln1(-/-) cortical neurons, implying their physiological health. Despite the absence of neuron loss, we observed alterations in synaptic plasticity, as indicated by elevated paired-pulse facilitation and enhanced long-term potentiation. Myelination deficits and severely dysmorphic corpus callosum were present early and resembled white matter pathology in mucolipidosis IV patients. These results indicate the early involvement of glia, and challenge the traditional view of mucolipidosis IV as an overtly neurodegenerative condition.

The transition between soluble intrinsically disordered tau protein and aggregated tau in neurofibrillary tangles in Alzheimer's disease is unknown. Here, we propose that soluble tau species can undergo liquid-liquid phase separation (LLPS) under cellular conditions and that phase-separated tau droplets can serve as an intermediate toward tau aggregate formation. We demonstrate that phosphorylated or mutant aggregation prone recombinant tau undergoes LLPS, as does high molecular weight soluble phospho-tau isolated from human Alzheimer brain. Droplet-like tau can also be observed in neurons and other cells. We found that tau droplets become gel-like in minutes, and over days start to spontaneously form thioflavin-S-positive tau aggregates that are competent of seeding cellular tau aggregation. Since analogous LLPS observations have been made for FUS, hnRNPA1, and TDP43, which aggregate in the context of amyotrophic lateral sclerosis, we suggest that LLPS represents a biophysical process with a role in multiple different neurodegenerative diseases.

Alzheimer's disease (AD) is defined by the presence of intraneuronal neurofibrillary tangles (NFTs) composed of hyperphosphorylated tau aggregates as well as extracellular amyloid-beta plaques. The presence and spread of tau pathology through the brain is classified by Braak stages and thought to correlate with the progression of AD. Several in vitro and in vivo studies have examined the ability of tau pathology to move from one neuron to the next, suggesting a "prion-like" spread of tau aggregates may be an underlying cause of Braak tau staging in AD. Using the HEK293 TauRD-P301S-CFP/YFP expressing biosensor cells as a highly sensitive and specific tool to identify the presence of seed competent aggregated tau in brain lysate-i.e., tau aggregates that are capable of recruiting and misfolding monomeric tau-, we detected substantial tau seeding levels in the entorhinal cortex from human cases with only very rare NFTs, suggesting that soluble tau aggregates can exist prior to the development of overt tau pathology. We next looked at tau seeding levels in human brains of varying Braak stages along six regions of the Braak Tau Pathway. Tau seeding levels were detected not only in the brain regions impacted by pathology, but also in the subsequent non-pathology containing region along the Braak pathway. These data imply that pathogenic tau aggregates precede overt tau pathology in a manner that is consistent with transneuronal spread of tau aggregates. We then detected tau seeding in frontal white matter tracts and the optic nerve, two brain regions comprised of axons that contain little to no neuronal cell bodies, implying that tau aggregates can indeed traverse along axons. Finally, we isolated cytosolic and synaptosome fractions along the Braak Tau Pathway from brains of varying Braak stages. Phosphorylated and seed competent tau was significantly enriched in the synaptic fraction of brain regions that did not have extensive cellular tau pathology, further suggesting that aggregated tau seeds move through the human brain along synaptically connected neurons. Together, these data provide further evidence that the spread of tau aggregates through the human brain along synaptically connected networks results in the pathogenesis of human Alzheimer's disease.

Adeno-associated viral vectors (AAVs) have demonstrated potential in applications for neurologic disorders, and the discovery that some AAVs can cross the blood-brain barrier (BBB) after intravenous injection has further expanded these opportunities for non-invasive brain delivery. Anc80L65, a novel AAV capsid designed from in silico reconstruction of the viral evolutionary lineage, has previously demonstrated robust transduction capabilities after local delivery in various tissues such as liver, retina, or cochlea, compared with conventional AAVs. Here, we compared the transduction efficacy of Anc80L65 with conventional AAV9 in the CNS after intravenous, intracerebroventricular (i.c.v.), or intraparenchymal injections. Anc80L65 was more potent at targeting the brain and spinal cord after intravenous injection than AAV9, and mostly transduced astrocytes and a wide range of neuronal subpopulations. Although the efficacy of Anc80L65 and AAV9 is similar after direct intraparenchymal injection in the striatum, Anc80L65's diffusion throughout the CNS was more extensive than AAV9 after i.c.v. infusion, leading to widespread EGFP expression in the cerebellum. These findings demonstrate that Anc80L65 is a highly efficient gene transfer vector for the murine CNS. Systemic injection of Anc80L65 leads to notable expression in the CNS that does not rely on a self-complementary genome. These data warrant further testing in larger animal models.

Aggregation of amyloid beta peptide into senile plaques and hyperphosphorylated tau protein into neurofibrillary tangles in the brain are the pathological hallmarks of Alzheimer's disease. Despite over a century of research into these lesions, the exact relationship between pathology and neurotoxicity has yet to be fully elucidated. In order to study the formation of plaques and tangles and their effects on the brain, we have applied multiphoton in vivo imaging of transgenic mouse models of Alzheimer's disease. This technique allows longitudinal imaging of pathological aggregation of proteins and the subsequent changes in surrounding neuropil neurodegeneration and recovery after therapeutic interventions.

Alzheimer's disease (AD) is a neurodegenerative disorder that impairs memory and cognition. One of the major neuropathological hallmarks is the accumulation of the extracellular senile plaques that are mainly composed of amyloid beta (Abeta) protein. Plaques are associated with synapse loss, dystrophic neurites and altered neurite trajectories. A reversal of such morphological changes has been observed days after single dose anti-Abeta immunotherapy. In this study we investigated the extended effects of a single dose of passive anti-Abeta immunotherapy on morphological changes associated with senile plaques. We found that although plaque burden was not reduced 30 days after immunotherapy, there were fewer dystrophic neurites around each plaque, a recovery of synapse density, and normalization of neurite curvature near plaques. Taken together these results suggest that a single dose of immunotherapy is sufficient to cause lasting benefits to the morphology of cortical neurons, implying substantial plasticity of neural circuits despite the continued presence of plaques.

We determined the crystal structure of 1TM-alphaVbeta3, which represents the complete unconstrained ectodomain plus short C-terminal transmembrane stretches of the alphaV and beta3 subunits. 1TM-alphaVbeta3 is more compact and less active in solution when compared with DeltaTM-alphaVbeta3, which lacks the short C-terminal stretches. The structure reveals a bent conformation and defines the alpha-beta interface between IE2 (EGF-like 2) and the thigh domains. Modifying this interface by site-directed mutagenesis leads to robust integrin activation. Fluorescent lifetime imaging microscopy of inactive full-length alphaVbeta3 on live cells yields a donor-membrane acceptor distance, which is consistent with the bent conformation and does not change in the activated integrin. These data are the first direct demonstration of conformational coupling of the integrin leg and head domains, identify the IE2-thigh interface as a critical steric barrier in integrin activation, and suggest that inside-out activation in intact cells may involve conformational changes other than the postulated switch to a genu-linear state.