Faculty Bibliography

INTRODUCTION/BACKGROUND:MicroRNA (miRNA) activity is increasingly appreciated as a key regulator of pathophysiologic pathways in Alzheimer's disease (AD). However, the role of miRNAs during the progression of AD, including resilience and prodromal syndromes such as mild cognitive impairment (MCI), remains underexplored. METHODS:We performed miRNA-sequencing on samples of posterior cingulate cortex (PCC) obtained post mortem from Rush Religious Orders Study participants diagnosed ante mortem with no cognitive impairment (NCI), MCI, or AD. NCI subjects were subdivided as low pathology (Braak stage I/II) or high pathology (Braak stage III/IV), suggestive of resilience. Bioinformatics approaches included differential expression, messenger RNA (mRNA) target prediction, interactome modeling, functional enrichment, and AD risk modeling. RESULTS:We identified specific miRNA groups, mRNA targets, and signaling pathways distinguishing AD, MCI, resilience, ante mortem neuropsychological test performance, post mortem neuropathological burden, and AD risk. DISCUSSION/CONCLUSIONS:These findings highlight the potential of harnessing miRNA activity to manipulate disease-modifying pathways in AD, with implications for precision medicine. HIGHLIGHTS/CONCLUSIONS:MicroRNA (MiRNA) dysregulation is a well-established feature of Alzheimer's disease (AD). Novel miRNAs also distinguish subjects with mild cognitive impairment and putative resilience. MiRNAs correlate with cognitive performance and neuropathological burden. Select miRNAs are associated with AD risk with age as a significant covariate. MiRNA pathways include insulin, prolactin, kinases, and neurite plasticity.

Selective vulnerability of neuronal populations occurs in both Down syndrome (DS) and Alzheimer's disease (AD), resulting in disproportional degeneration of pyramidal neurons (PNs) affecting memory and executive function. Elucidating the cellular mechanisms underlying the selective vulnerability of these populations will provide pivotal insights for disease progression in DS and AD. Single population RNA-sequencing analysis was performed on neurons critical for executive function, prefrontal cortex Brodmann area 9 (BA9) layer III (L3) and layer V (L5) excitatory PNs in postmortem human DS and age- and sex-matched control (CTR) brains. Data mining was performed on differentially expressed genes (DEGs) from PNs in each lamina with DEGs divergent between lamina identified and interrogated. Bioinformatic inquiry of L3 PNs revealed more unique/differentially expressed DEGs (uDEGs) than in L5 PNs in DS compared to CTR subjects, indicating gene dysregulation shows both spatial and cortical laminar projection neuron dependent dysregulation. DS triplicated human chromosome 21 (HSA21) comprised a subset of DEGs only dysregulated in L3 or L5 neurons, demonstrating partial cellular specificity in HSA21 expression. These HSA21 uDEGs had a disproportionally high number of noncoding RNAs, suggesting lamina specific dysfunctional gene regulation. L3 uDEGs revealed overall more dysregulation of cellular pathways and processes, many relevant to early AD pathogenesis, while L5 revealed processes suggestive of frank AD pathology. These findings indicate that trisomy differentially affects a subpopulation of uDEGs in L3 and L5 BA9 projection neurons in aged individuals with DS, which may inform circuit specific pathogenesis underlying DS and AD.

Parkinson's disease (PD) is a prevalent neurodegenerative disorder, and recent evidence suggests that pathogenesis may be in part mediated by inflammatory processes, the molecular and cellular architectures of which are largely unknown. To identify and characterize selectively vulnerable brain cell populations in PD, we performed single-nucleus transcriptomics and unbiased proteomics to profile the prefrontal cortex from postmortem human brains of six individuals with late-stage PD and six age-matched controls. Analysis of nearly 80,000 nuclei led to the identification of eight major brain cell types, including elevated brain-resident T cells in PD, each with distinct transcriptional changes in agreement with the known genetics of PD. By analyzing Lewy body pathology in the same postmortem brain tissues, we found that α-synuclein pathology was inversely correlated with chaperone expression in excitatory neurons. Examining cell-cell interactions, we found a selective abatement of neuron-astrocyte interactions and enhanced neuroinflammation. Proteomic analyses of the same brains identified synaptic proteins in the prefrontal cortex that were preferentially down-regulated in PD. By comparing this single-cell PD dataset with a published analysis of similar brain regions in Alzheimer's disease (AD), we found no common differentially expressed genes in neurons but identified many shared differentially expressed genes in glial cells, suggesting that the disease etiologies, especially in the context of neuronal vulnerability, in PD and AD are likely distinct.

The posterior cingulate cortex (PCC) is a key hub of the default mode network underlying autobiographical memory retrieval, which falters early in the progression of Alzheimer's disease (AD). We recently performed RNA sequencing of post-mortem PCC tissue samples from 26 elderly Rush Religious Orders Study participants who came to autopsy with an ante-mortem diagnosis of no cognitive impairment but who collectively displayed a range of Braak I-IV neurofibrillary tangle stages. Notably, cognitively unimpaired subjects displaying high Braak stages may represent cognitive resilience to AD pathology. Transcriptomic data revealed elevated synaptic and ATP-related gene expression in Braak Stages III/IV compared with Stages I/II, suggesting these pathways may be related to PCC resilience. We also mined expression profiles for small non-coding micro-RNAs (miRNAs), which regulate mRNA stability and may represent an underexplored potential mechanism of resilience through the fine-tuning of gene expression within complex cellular networks. Twelve miRNAs were identified as differentially expressed between Braak Stages I/II and III/IV. However, the extent to which the levels of all identified miRNAs were associated with subject demographics, neuropsychological test performance and/or neuropathological diagnostic criteria within this cohort was not explored. Here, we report that a total of 667 miRNAs are significantly associated (rho > 0.38, P < 0.05) with subject variables. There were significant positive correlations between miRNA expression levels and age, perceptual orientation and perceptual speed. By contrast, higher miRNA levels correlated negatively with semantic and episodic memory. Higher expression of 15 miRNAs associated with lower Braak Stages I-II and 47 miRNAs were associated with higher Braak Stages III-IV, suggesting additional mechanistic influences of PCC miRNA expression with resilience. Pathway analysis showed enrichment for miRNAs operating in pathways related to lysine degradation and fatty acid synthesis and metabolism. Finally, we demonstrated that the 12 resilience-related miRNAs differentially expressed in Braak Stages I/II versus Braak Stages III/IV were predicted to regulate mRNAs related to amyloid processing, tau and inflammation. In summary, we demonstrate a dynamic state wherein differential PCC miRNA levels are associated with cognitive performance and post-mortem neuropathological AD diagnostic criteria in cognitively intact elders. We posit these relationships may inform miRNA transcriptional alterations within the PCC relevant to potential early protective (resilience) or pathogenic (pre-clinical or prodromal) responses to disease pathogenesis and thus may be therapeutic targets.

Neurodegenerative disorders, including Alzheimer's disease (AD) and Parkinson's disease (PD), represent debilitating conditions with complex, poorly understood pathologies. Epichaperomes, pathologic protein assemblies nucleated on key chaperones, have emerged as critical players in the molecular dysfunction underlying these disorders. In this study, we introduce the synthesis and characterization of clickable epichaperome probes, PU-TCO, positive control, and PU-NTCO, negative control. Through comprehensive in vitro assays and cell-based investigations, we establish the specificity of the PU-TCO probe for epichaperomes. Furthermore, we demonstrate the efficacy of PU-TCO in detecting epichaperomes in brain tissue with a cellular resolution, underscoring its potential as a valuable tool for dissecting single-cell responses in neurodegenerative diseases. This clickable probe is therefore poised to address a critical need in the field, offering unprecedented precision and versatility in studying epichaperomes and opening avenues for novel insights into their role in disease pathology.

The intricate network of protein-chaperone interactions is crucial for maintaining cellular function. Recent discoveries have unveiled the existence of specialized chaperone assemblies, known as epichaperomes, which serve as scaffolding platforms that orchestrate the reconfiguration of protein-protein interaction networks, thereby enhancing cellular adaptability and proliferation. This study explores the structural and regulatory aspects of epichaperomes, with a particular focus on the role of post-translational modifications (PTMs) in their formation and function. A key finding is the identification of specific PTMs on HSP90, particularly at residues Ser226 and Ser255 within an intrinsically disordered region, as critical determinants of epichaperome assembly. Our data demonstrate that phosphorylation of these serine residues enhances HSP90's interactions with other chaperones and co-chaperones, creating a microenvironment conducive to epichaperome formation. Moreover, we establish a direct link between epichaperome function and cellular physiology, particularly in contexts where robust proliferation and adaptive behavior are essential, such as in cancer and pluripotent stem cell maintenance. These findings not only provide mechanistic insights but also hold promise for the development of novel therapeutic strategies targeting chaperone assemblies in diseases characterized by epichaperome dysregulation, thereby bridging the gap between fundamental research and precision medicine.

Brain-derived neurotrophic factor (BDNF) is important in the development and maintenance of neurons and their plasticity. Hippocampal BDNF has been implicated in Alzheimer's disease (AD) because hippocampal levels in AD patients and AD animal models are often downregulated, suggesting that reduced BDNF contributes to AD. However, the location where hippocampal BDNF protein is most highly expressed, the mossy fiber (MF) axons of dentate gyrus granule cells (GCs), has been understudied, and not in controlled conditions. Therefore, we evaluated MF BDNF protein in the Tg2576 mouse model of AD. Tg2576 and wild-type (WT) mice of both sexes were examined at 2-3 months of age, when amyloid-β (Aβ) is present in neurons but plaques are absent, and 11-20 months of age, after plaque accumulation. As shown previously, WT mice exhibited high levels of MF BDNF protein. Interestingly, there was no significant decline with age in either the genotype or sex. Notably, MF BDNF protein was correlated with GC ΔFosB, a transcription factor that increases after 1-2 weeks of elevated neuronal activity. We also report the novel finding that Aβ in GCs or the GC layer was minimal even at old ages. The results indicate that MF BDNF is stable in the Tg2576 mouse, and MF BDNF may remain unchanged due to increased GC neuronal activity, since BDNF expression is well known to be activity dependent. The resistance of GCs to long-term Aβ accumulation provides an opportunity to understand how to protect vulnerable neurons from increased Aβ levels and therefore has translational implications.

We elucidated the molecular fingerprint of vulnerable excitatory neurons within select cortical lamina of individuals with Down syndrome (DS) for mechanistic understanding and therapeutic potential that also informs Alzheimer's disease (AD) pathophysiology. Frontal cortex (BA9) layer III (L3) and layer V (L5) pyramidal neurons were microisolated from postmortem human DS and age- and sex-matched controls (CTR) to interrogate differentially expressed genes (DEGs) and key biological pathways relevant to neurodegenerative programs. We identified > 2300 DEGs exhibiting convergent dysregulation of gene expression in both L3 and L5 pyramidal neurons in individuals with DS versus CTR subjects. DEGs included over 100 triplicated human chromosome 21 genes in L3 and L5 neurons, demonstrating a trisomic neuronal karyotype in both laminae. In addition, thousands of other DEGs were identified, indicating gene dysregulation is not limited to trisomic genes in the aged DS brain, which we postulate is relevant to AD pathobiology. Convergent L3 and L5 DEGs highlighted pertinent biological pathways and identified key pathway-associated targets likely underlying corticocortical neurodegeneration and related cognitive decline in individuals with DS. Select key DEGs were interrogated as potential hub genes driving dysregulation, namely the triplicated DEGs amyloid precursor protein (APP) and superoxide dismutase 1 (SOD1), along with key signaling DEGs including mitogen activated protein kinase 1 and 3 (MAPK1, MAPK3) and calcium calmodulin dependent protein kinase II alpha (CAMK2A), among others. Hub DEGs determined from multiple pathway analyses identified potential therapeutic candidates for amelioration of cortical neuron dysfunction and cognitive decline in DS with translational relevance to AD.

Protein-protein interactions (PPIs) play a crucial role in cellular function and disease manifestation, with dysfunctions in PPI networks providing a direct link between stressors and phenotype. The dysfunctional Protein-Protein Interactome (dfPPI) platform, formerly known as epichaperomics, is a newly developed chemoproteomic method aimed at detecting dynamic changes at the systems level in PPI networks under stressor-induced cellular perturbations within disease states. This review provides an overview of dfPPIs, emphasizing the novel methodology, data analytics, and applications in disease research. dfPPI has applications in cancer research, where it identifies dysfunctions integral to maintaining malignant phenotypes and discovers strategies to enhance the efficacy of current therapies. In neurodegenerative disorders, dfPPI uncovers critical dysfunctions in cellular processes and stressor-specific vulnerabilities. Challenges, including data complexity and the potential for integration with other omics datasets are discussed. The dfPPI platform is a potent tool for dissecting disease systems biology by directly informing on dysfunctions in PPI networks and holds promise for advancing disease identification and therapeutics.

Alzheimer's disease (AD) is a neurodegenerative disorder with limited therapeutic options. Accordingly, new approaches for prevention and treatment are needed. One focus is the human microbiome, the consortium of microorganisms that live in and on us, which contributes to human immune, metabolic, and cognitive development and that may have mechanistic roles in neurodegeneration. AD and Alzheimer's disease-related dementias (ADRD) are recognized as spectrum disorders with complex pathobiology. AD/ADRD onset begins before overt clinical signs, but initiation triggers remain undefined. We posit that disruption of the normal gut microbiome in early life leads to a pathological cascade within septohippocampal and cortical brain circuits. We propose investigation to understand how early-life microbiota changes may lead to hallmark AD pathology in established AD/ADRD models. Specifically, we hypothesize that antibiotic exposure in early life leads to exacerbated AD-like disease endophenotypes that may be amenable to specific microbiological interventions. We propose suitable models for testing these hypotheses.