Biology subjects

Mancuso, R.

Publications and source records attributed to Mancuso, R..

14 recordsLinked to original sources

bioRxiv2026

Phage Display-Derived Cyclic Peptides as Ligand-Specific Modulators for β2-Integrin Receptors

The leukocyte-specific {beta}2-integrin receptor family exerts a wide range of functions: {beta}2-integrins are involved in leukocyte trafficking, where they mediate cell adhesion during inflammatory responses via binding to ICAM-1, ICAM-2, or JAM-C. Furthermore, they are essential for the recognition and phagocytosis of pathogens opsonized by complement. Accordingly, the {beta}2-integrin family is known to be involved in autoimmune and inflammatory diseases, such as systemic lupus erythematosus. Owing to their complex biology, involving multiple conformational transitions, different signaling pathways, and a broad spectrum of ligands, the development of {beta}2-integrin-targeted probes and therapeutics has remained challenging. We aimed to develop macrocyclic peptides, derived from phage display screening, which can be used to unravel ligand binding profiles of {beta}2-integrins with an emphasis on the I domain. The selection of suitable lead peptides, and the characterization of their interaction profiles with different I domains, was enabled by an established in-vitro assay platform. Various peptide sequences were enriched during several rounds of phage display against the I-domain of CR3, of which two peptides with particularly low micromolar binding affinity were further characterized. Both peptides showed direct binding to {beta}2-integrin I-domains and, in a competitive assay, dose-dependent inhibition of the I-domains interactions with their main ligands iC3b and ICAM-1, respectively. These ligand-interfering properties were confirmed in bead- and cell-based adhesion assays. The modulators developed here are expected to provide valuable insight into the (patho-)physiology of CR3 and the other members of the {beta}2-integrin family, as the two peptides were able to compete with different ligands. In the future, this may help to identify potential therapeutic approaches for autoimmune, inflammatory, and age-related diseases.

biochemistry↗

bioRxiv2026

Cross-protocol comparison of iPSC-microglia reveals hypofunction contributes to neuronal vulnerability and synaptic alterations in the MAPT-S305N model of frontotemporal dementia.

Progressive and chronic neuroinflammation is associated with numerous neurodegenerative diseases, including primary tauopathies such as frontotemporal dementia and progressive supranuclear palsy. Unlike Alzheimers disease, there is no clear genetic association implicating microglial dysfunction as a primary driver of tauopathy. As such, the contributions of microglia to tauopathy pathogenesis have been less well defined. Here, we explore the cell autonomous effects of the pathogenic MAPT-S305N variant on microglial function, across two distinct iPSC-microglia protocols, followed by examination of the non-cell autonomous effects of microglial MAPT genotype on neuronal health and function. We find that different protocols produce cells of equivalent microglial identity, but result in microglia in different functional states, thereby influencing reactivity and detectable phenotypes. Regardless, across both protocols we find that MAPT-S305N induces microglial hypoactivity, evidenced by impaired phagocytosis, reduced cytokine release and diminished regulation of synaptic function. We conclude that microglial hypoactivity may be an early event in disease pathogenesis, where MAPT mutation microglia fail to adequately respond to pathogenic stimuli, thereby contributing to subsequent neuronal vulnerability and susceptibility. Further studies are required to understand how and when this initial hypoactive state may switch to a toxic pro-inflammatory state, and whether early detection and correction may be of therapeutic value.

neuroscience↗

bioRxiv2025

Canonical nuclear envelope protein emerin regulates structure and integrity of the erythrocyte plasma membrane

Red blood cells (RBC) are essential for the survival of aerobic organisms. Mutations in RBC plasma membrane proteins can give rise to a spectrum of diseases characterized by hemolysis, inefficient gas exchange, and anemia. We tested whether emerin (EMD), a nuclear envelope (NE) protein, is also expressed in anuclear cell types to regulate membrane structure and function. We find that emerin is localized to the RBC plasma membrane and regulates the localization of other membrane proteins involved in actin branching and cytoskeletal dynamics. Additionally, we find that loss of Emd in mice results in decreased RBC corpuscular volume and hemoglobin content in both sexes, as well as a male-specific increase in RBC number and hematocrit, consistent with other male-dominated EMD phenotypes. Along with decreased RBC size, we observed a reduction in the rate of osmotic lysis in Emd KO animals, particularly males. We found that human EMD mutations also have a statistically significant association with RBC phenotypes, namely MCV. Individuals with emerin-related Emery-Dreifuss muscular dystrophy (EDMD) also demonstrate a mild yet consistent RBC phenotype according to patient CBC data. Together, these data suggest that emerin can have cell-specific localization and functions independent of its canonical repertoire in nucleated cells.

cell biology↗

bioRxiv2024

The Alzheimers Disease Risk Genes MS4A4A And MS4A6A Cooperate to Negatively Regulate Trem2 and Microglia states

Genetic variations in MS4A4A and MS4A6A are linked to the regulation of cerebrospinal fluid soluble TREM2 (sTREM2) levels and are associated with Alzheimers disease (AD) risk and progression. Using CRISPR knockout and MS4A4A-degrading antibodies in primary human microglia, non-human primates (NHP), and a xenotransplantation model of amyloid pathology, we provide evidence that MS4A4A and MS4A6A are negative regulators of both the transmembrane and soluble TREM2 proteins. They also negatively regulate microglia proliferation, survival, metabolism, lysosomal function, energetics, phagocytosis, and disease-fighting states. Mechanistically, we find that MS4A4A exerts negative regulation by interacting with MS4A6A and protecting it from degradation. MS4A6A in turn forms a complex with and blocks the co-receptor DAP12, which is required for the stability, cell surface localization, and signaling of TREM2 and other receptors. Taken together, the data indicate that MS4A4A and MS4A6A are cooperating, post-transcriptional negative regulators of TREM2 and microglial function, and potential drug targets for AD.

neuroscience↗

bioRxiv2024

The APOE isoforms differentially shape the transcriptomic and epigenomic landscapes of human microglia in a xenotransplantation model of Alzheimer's disease

Microglia play a key role in the response to amyloid beta in Alzheimers disease (AD). In this context, a major transcriptional response of microglia is the upregulation of APOE, the strongest late-onset AD risk gene. Of its three isoforms, APOE2 is thought to be protective, while APOE4 increases AD risk. We hypothesised that the isoforms functionally alter microglia by shaping their transcriptomic and chromatin landscapes. We used RNA- and ATAC-sequencing to profile gene expression and chromatin accessibility of human microglia isolated from a xenotransplantation model of AD. We identified widespread transcriptomic and epigenomic differences which are dependent on APOE genotype, and are corroborated across the profiling assays. Our results indicate that impaired microglial proliferation, migration and immune responses may contribute to the increased risk for late-onset AD in APOE4 carriers, while increased DNA-binding of the vitamin D receptor in APOE2 microglia may contribute to the isoforms protective role.

genomics↗

bioRxiv2023

S100A8-enriched microglia populate the brain of tau-seeded and accelerated aging mice

Long considered to fluctuate between pro- and anti-inflammatory states, it has now become evident that microglia occupy a variegated phenotypic landscape with relevance to aging and neurodegeneration. However, whether specific microglial subsets converge in or contribute to both processes that eventually affect brain function is less clear. To investigate this, we analyzed microglial heterogeneity in a tauopathy mouse model (K18-seeded P301L) and an accelerated aging model (senescence accelerated mouse prone 8, SAMP8) using cellular indexing of transcriptomes and epitopes by sequencing. We found that widespread tau pathology in K18-seeded P301L mice caused a significant change in the number and morphology of microglia, but only a mild overrepresentation of disease-associated microglia. At the cell population-level, we observed a marked upregulation of the calprotectin-encoding genes S100a8 and S100a9. In 9-months-old SAMP8 mice, we identified a unique microglial subpopulation that showed partial similarity with the disease-associated microglia phenotype and was additionally characterized by a high expression of the same calprotectin gene set. Immunostaining for S100A8 revealed that this population was enriched in the hippocampus, correlating with the cognitive impairment observed in this model. However, incomplete colocalization between their residence and markers of neuronal loss suggests regional specificity. Importantly, S100A8-positive microglia were also retrieved in brain biopsies of human AD and tauopathy patients as well as in a biopsy of an aged individual without reported pathology. Thus, the emergence of S100A8-positive microglia portrays a conspicuous commonality between accelerated aging and tauopathy progression, which may have relevance for ensuing brain dysfunction. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=162 SRC="FIGDIR/small/566543v1_ufig1.gif" ALT="Figure 1"> View larger version (28K): org.highwire.dtl.DTLVardef@d1e4e1org.highwire.dtl.DTLVardef@98adeorg.highwire.dtl.DTLVardef@7c15aaorg.highwire.dtl.DTLVardef@7265ca_HPS_FORMAT_FIGEXP M_FIG C_FIG

neuroscience↗

bioRxiv2023

The role of microRNAs to understand sex-based differences in Alzheimer's disease

BackgroundThe incidence of Alzheimers disease (AD) - the most frequent cause of dementia - is expected to increase as life expectancies rise across the globe. While sex-based differences in AD have previously been described, there remain uncertainties regarding any association between sex and disease-associated molecular mechanisms. Studying sex-specific expression profiles of regulatory factors such as microRNAs (miRNAs) could contribute to more accurate disease diagnosis and treatment. MethodsA systematic review identified five studies of microRNA expression in AD patients that incorporated information regarding the biological sex of samples in the Gene Expression Omnibus repository. A differential microRNA expression analysis was performed, considering disease status and patient sex. Subsequently, results were integrated within a meta-analysis methodology, with a functional enrichment of meta-analysis results establishing an association between altered miRNA expression and relevant Gene Ontology terms. ResultsMeta-analyses of miRNA expression profiles in blood samples revealed the alteration of sixteen miRNAs in female and twenty-two miRNAs in male AD patients. We discovered nine miRNAs commonly overexpressed in both sexes, suggesting a shared miRNA dysregulation profile. Functional enrichment results based on miRNA profiles revealed sex-based differences in biological processes; most affected processes related to ubiquitination, regulation of different kinase activities, and apoptotic processes in males, but RNA splicing and translation in females. Meta-analyses of miRNA expression profiles in brain samples revealed the alteration of six miRNAs in female and four miRNAs in male AD patients. We observed a single underexpressed miRNA in female and male AD patients (hsa-miR-767-5p); however, the functional enrichment analysis for brain samples did not reveal any specifically affected biological process. ConclusionsSex-specific meta-analyses supported the detection of differentially expressed miRNAs in female and male AD patients, highlighting the relevance of sex-based information in biomedical data. Further studies on miRNA regulation in AD patients should meet the criteria for comparability and standardization of information. HighlightsO_LIDeregulation of miRNA expression profiles occurs in a tissue- and sex-specific manner in AD patients C_LIO_LIMeta-analysis of blood samples revealed a partial overlapping pattern of altered miRNA expression in female and male AD patients C_LIO_LIFunctional enrichment based on AD-associated miRNA expression profiles in blood samples reveals sex-based differences: RNA splicing and translation in female AD patients and ubiquitination, regulation of different kinase activities, and apoptotic process in male AD patients C_LIO_LILinks between AD development and miRNA expression in brain tissue also demonstrate the influence of sex C_LI Plain English SummaryAlzheimers disease (AD) - a neurodegenerative disease mainly affecting older patients - is characterized by cognitive deterioration, memory loss, and progressive incapacitation in daily activities. While AD affects almost twice as many females as males, and cognitive deterioration and brain atrophy develop more rapidly in females, the biological causes of these differences remain poorly understood. MicroRNAs (miRNAs) regulate gene expression and impact a wide variety of biological processes; therefore, studying the differential expression of miRNAs in female and male AD patients could contribute to a better understanding of the disease. We reviewed studies of miRNA expression in female and male AD patients and integrated results using a meta-analysis methodology and then identified those genes regulated by the altered miRNAs to establish an association with biological processes. We found sixteen (females) and twenty-two (males) miRNAs altered in the blood of AD patients. Functional enrichment revealed sex-based differences in the affected altered biological processes - protein modification and degradation and cell death in male AD patients and RNA processing in female AD patients. A similar analysis in the brains of AD patients revealed six (females) and four (males) miRNAs with altered expression; however, our analysis failed to highlight any specifically altered biological processes. Overall, we highlight the sex-based differential expression of miRNAs (and biological processes affected) in the blood and brain of AD patients.

neuroscience↗

bioRxiv2022

Hexanucleotide repeat expansions in C9orf72 alter microglial responses and prevent a coordinated glial reaction in ALS

Neuroinflammation is an important hallmark in amyotrophic lateral sclerosis (ALS). Experimental evidence has highlighted a role of microglia in the modulation of motor neuron degeneration. However, the exact contribution of microglia to both sporadic and genetic forms of ALS is still unclear. We generated single nuclei profiles of spinal cord and motor cortex from sporadic and C9orf72 ALS patients, as well as controls. We particularly focused on the transcriptomic responses of both microglia and astrocytes. We confirmed that C9orf72 is highly expressed in microglia and shows a diminished expression in carriers of the hexanucleotide repeat expansion (HRE). This resulted in an impaired response to disease, with specific deficits in phagocytic and lysosomal transcriptional pathways. Astrocytes also displayed a dysregulated response in C9orf72 ALS patients, remaining in a homeostatic state. This suggests that C9orf72 HRE alters a coordinated glial response, which ultimately would increase the risk for developing ALS. Our results indicate that C9orf72 HRE results in a selective microglial loss-of-function, likely impairing microglial-astrocyte communication and preventing a global glial response. This is relevant as it indicates that sporadic and familial forms of ALS may present a different cellular substrate, which is of great importance for patient stratification and treatment.

neuroscience↗

bioRxiv2022

Deep proteomic analysis of human microglia and model systems reveal fundamental biological differences of in vitro and ex vivo cells

Using high resolution quantitative mass spectrometry, we have generated the most comprehensive human and mouse microglia proteomic datasets to date, consisting of over 11,000 proteins across all six microglia groups. Microglia from different sources share a core protein signature of over 5600 proteins, yet fundamental differences are observed between species and culture conditions, indicating limitations for human disease modelling in mouse or in in vitro cultures of microglia. Mouse ex vivo microglia show important differences at the proteome level such as differential expression of inflammation and Alzheimers Disease associated proteins. We identify a tenfold difference in the protein content of ex vivo and in vitro cells and significant proteome differences associated with protein synthesis, metabolism, microglia marker expression and environmental sensors. Culturing microglia induces rapidly increased growth, protein content and inflammatory protein expression. These changes can be restored by engrafting in vitro cells into the brain, with xenografted hESC-derived microglia closely resembling microglia from human brain. This data provides an important resource for the field and highlights important considerations needed when using model systems to study human physiology and pathology of microglia.

neuroscience↗

bioRxiv2022

A multi-pronged human microglia response to Alzheimer's disease Aβ pathology

Microglial activation and neuroinflammation are initial steps in the pathogenesis of Alzheimers disease (AD). However, studies in mouse models and human postmortem samples have yielded divergent results regarding microglia cell states relevant to AD. Here, we investigate 127,000 single cell expression profiles of human microglia isolated freshly from a xenotransplantation model for early AD. While human microglia adopt a disease-associated (DAM) profile, they display a much more pronounced HLA-cell state related to antigen presentation in response to amyloid plaques. In parallel, a distinctive pro-inflammatory cytokine and chemokine CRM response is mounted against oligomeric amyloid-{beta}. TREM2 and, to a lesser extent, APOE polymorphisms, modulate the response of microglia to amyloid-{beta} plaques, in contrast with the response to oligomeric A{beta}. Specific polygenic risk genes are enriched in each branch of these multi-pronged response of human microglia to amyloid pathology (ARM). ARM responses can be captured in post-mortem studies when reanalyzed in light of this novel, comprehensive data set. In conclusion, therapeutic strategies targeting microglia in AD need to carefully assess how they affect the different cell states, as the overall balance between distinct microglial profiles might determine a protective or damaging outcome.

neuroscience↗

bioRxiv2020

Single nucleus sequencing fails to detect microglial activation

Single nucleus RNA-Seq (snRNA-Seq) methods are used as an alternative to single cell RNA-Seq methods, as they allow transcriptomic profiling of frozen tissue. However, it is unclear whether snRNA-Seq is able to detect cellular state in human tissue. Indeed, snRNA-Seq analyses of human brain samples have failed to detect a consistent microglial activation signature in Alzheimers Disease. A comparison of microglia from single cells and single nuclei of four human subjects reveals that ~1% of genes is depleted in nuclei compared to whole cells. This small population contains 18% of genes previously implicated in microglial activation, including APOE, CST3, FTL, SPP1, and CD74. We confirm our findings across multiple previous single nucleus and single cell studies. Given the low sensitivity of snRNA-Seq to this population of activation genes, we conclude that snRNA-Seq is not suited to detecting cellular activation in microglia in human disease.

neuroscience↗

bioRxiv2019

Spatial and temporal transcriptomics reveal microglia-astroglia crosstalk in the amyloid-β plaque cell niche of Alzheimer’s disease

The linear cause-consequence relationship linking amyloid-{beta} peptide (A{beta}) accumulation to neuronal dysfunction in Alzheimer disease (AD) is gradually replaced by the concept that A{beta} initiates complex inflammatory-like cellular alterations that progressively become A{beta} independent and lead to brain dyshomeostasis. Little is known about the pathophysiology of this cellular phase of AD. We use here two orthogonal technologies, Spatial Transcriptomics and in situ sequencing, to analyse the transcriptome changes in cells in the amyloid-{beta} plaque niche in a knock-in mouse model for AD. We identify a multicellular co-expressed gene network of 57 Plaque-Induced Genes (PIGs) that define a series of co-ordinated and spatially restricted microglia, astroglia and oligodendrocyte responses to progressing amyloid plaques encompassing complement, oxidative stress and inflammation. A separate oligodendrocyte network suggests abnormal myelination. Spatial Transcriptomics provides an unprecedented approach to untangle the dysregulated cellular network in the vicinity of pathogenic hallmarks of AD and other brain diseases.

neuroscience↗

bioRxiv2019

Stem cell derived human microglia transplanted in mouse brain to study genetic risk of Alzheimer's Disease

Summary paragraphGenetics highlight the central role of microglia in Alzheimers disease but at least 36% of AD-risk genes lack good mouse orthologues. Here, we show that embryonic stem cell (ESC)-derived human microglia successfully engraft the mouse brain and recapitulate transcriptionally primary human microglia derived from human surgical samples. Upon exposure to oligomeric A{beta} a wide range of AD-risk genes are expressed that are not readily studied in current mouse models for AD. This work provides a unique humanized animal model that will allow elucidating the role of genetic risk in the pathogenesis of AD.

neuroscience↗

bioRxiv2018

Novel Alzheimer risk genes determine the microglia response to amyloid-β but not to TAU pathology

BackgroundThousands of SNPs associated with risk of Alzheimers disease (AD) in genome-wide association studies (GWAS) do not reach genome-wide significance. When combined, they contribute however to a highly predictive polygenic risk score. The relevance of these subthreshold risk genes to disease, and how their combined predictive power translates into functionally relevant disease pathways, is unknown. We investigate here at the genome-wide level and in an unbiased way to what extent AD risk genes show altered gene expression in the context of increasing A{beta} or Tau pathology in mouse models of AD.\n\nMethodsWe used an existing GWAS data set to generate lists of candidate AD genes at different levels of significance. We performed transcriptomic analysis on wild-type and transgenic APP/PS1 (APPtg) and Thy-TAU22 (TAUtg) mouse models at early and late stage of disease. We used unbiased weighted gene co-expression network analysis (WGCNA) to identify clusters of co-regulated genes responsive to A{beta} or TAU pathology. Gene set enrichment was used to identify clusters that were enriched for AD risk genes.\n\nFindingsConsistent and significant enrichment of AD risk genes was found in only one out of 63 coexpression modules. This module is highly responsive to A{beta} but not to TAU pathology. We identify in this module 18 AD risk genes (p-value=6{middle dot}5e-11) including 11 new ones, GPC2, TREML2, SYK, GRN, SLC2A5, SAMSN1, PYDC1, HEXB, RRBP1, LYN and BLNK. All are expressed in microglia, have a binding site for the transcription factor SPI1 (PU.1), and become significantly upregulated when exposed to A{beta}. A subset regulates FC-gamma receptor mediated phagocytosis.\n\nInterpretationGenetic risk of AD is functionally translated into a microglia pathway responsive to A{beta} pathology. This insight integrates aspects of the amyloid hypothesis with genetic risk associated to sporadic AD.

neuroscience↗