Andrea Crotti - Academia.edu (original) (raw)

Papers by Andrea Crotti

Research Square (Research Square), May 2, 2023

Single-cell technologies have revealed important cell heterogeneity across the human brain. In th... more Single-cell technologies have revealed important cell heterogeneity across the human brain. In the context of neurological diseases, the relationship between different cell subpopulations and pathological features can be crucial for their understanding. However, the combination of spatial, single-cell, and high-plex protein information together with analytical tools for the accurate segmentation of ramified cells remains a challenge to address in neuroscience. Here, we present CODEX-CNS, a modification of CO-Detection by indEXing (CODEX) technology for its use in human brain tissues. This technology consists of multiplexed fluorescent imaging that allows the detection of up to 100 proteins using DNA-barcoded antibodies. In this paper we bring technical advancements to the CODEX protocol and an improved data analysis pipeline for accurate segmentation of complex cell morphologies. As proof-of-principle, we were able to detect the different parenchymal brain cells and their cytoarchitecture, as well as bloodbrain barrier and meningeal components with a 32-plex antibody panel. More specifically, we used CODEX-CNS in human brain samples of healthy and Alzheimer's disease donors to study microglial phenotypes in relationship to their spatial context. Applying our customized cell segmentation algorithm and clustering analysis, we identified diverse microglial subpopulations differentially distributed between brain areas and according to their distance to amyloid-b plaques. These data provide a new approach for the neuroscience community that allows the characterization of microglial subpopulations at the protein level with both single-cell and spatial resolution.

We propose Mobile Probabilistic ADdressing (MPAD), a virtual-coordinates-based routing protocol f... more We propose Mobile Probabilistic ADdressing (MPAD), a virtual-coordinates-based routing protocol for Wireless Mesh Networks (WMNs). In contrast to other virtual coordinates based protocols such as Beacon Vector Routing (BVR), our approach collects statistical distributions of hop distances, thereby avoiding explicit link estimation. MPAD therefore captures topology changes arising from link variability and mobility more gracefully. We implemented our prototype for wireless mesh nodes and the OMNeT++ simulator, and evaluated it on a real testbed and in simulation. In the simulation we evaluated three different mobility scenarios. From the results that we obtained, we can conclude that MPAD is a feasible routing protocol for WMNs.

Nature Neuroscience, 2018

The Open Virology Journal, 2007

HIV-1 can be subdivided into distinct subtypes; the consequences of such a genomic variability re... more HIV-1 can be subdivided into distinct subtypes; the consequences of such a genomic variability remain largely speculative. The long terminal repeats (LTR) control HIV transcription and reflect the major differences of distinct viral subtypes. Three regions in the HIV-1 subtype B LTR are close matches to the Signal Transducer and Activator of Transcription (STAT) consensus sequence. Here, we show heterogeneity in these putative STAT binding sites among HIV-1 LTR subtypes A through G. Transfection of constitutively activated STAT5 lead to transcriptional activation of HIV-1 expression in 293T cells transfected with a reporter assay driven by HIV-1 LTR subtype B. Constitutively activated STAT5 transactivated the LTR of various subtypes in U937 cells with different potency. These findings support and expand the potential relevance of STAT5 activation in HIV infection and may bear relevance for a differential regulation of latency and expression of different subtypes of HIV-1.

PLOS ONE, 2019

BIN1 is the most important risk locus for Late Onset Alzheimer's Disease (LOAD), after ApoE. BIN1... more BIN1 is the most important risk locus for Late Onset Alzheimer's Disease (LOAD), after ApoE. BIN1 AD-associated SNPs correlate with Tau deposition as well as with brain atrophy. Furthermore, the level of neuronal-specific BIN1 isoform 1 protein is decreased in sporadic AD cases in parallel with neuronal loss, despite an overall increase in BIN1 total mRNA. To address the relationship between reduction of BIN1 and neuronal cell loss in the context of Tau pathology, we knocked-down endogenous murine Bin1 via stereotaxic injection of AAV-Bin1 shRNA in the hippocampus of mice expressing Tau P301S (PS19). We observed a statistically significant reduction in the number of neurons in the hippocampus of mice injected with AAV-Bin1 shRNA in comparison with mice injected with AAV control. To investigate whether neuronal loss is due to deletion of Bin1 selectively in neurons in presence Tau P301S, we bred Bin1 flox/flox with Thy1-Cre and subsequently with PS19 mice. Mice lacking neuronal Bin1 and expressing Tau P301S showed increased mortality, without increased neuropathology, when compared to neuronal Bin1 and Tau P301S-expressing mice. The loss of Bin1 isoform 1 resulted in reduced excitability in primary neurons in vitro, reduced neuronal c-fos expression as well as in altered microglia transcriptome in vivo. Taken together, our data suggest that the contribution of genetic variation in BIN1 locus to AD risk could result from a cell-autonomous reduction of neuronal excitability due to Bin1 decrease, exacerbated by the presence of aggregated Tau, coupled with a non-cell autonomous microglia activation.

Scientific Reports, 2019

Despite Bridging INtegrator 1 (BIN1) being the second most statistically-significant locus associ... more Despite Bridging INtegrator 1 (BIN1) being the second most statistically-significant locus associated to Late Onset Alzheimer’s Disease, its role in disease pathogenesis remains to be clarified. As reports suggest a link between BIN1, Tau and extracellular vesicles, we investigated whether BIN1 could affect Tau spreading via exosomes secretion. We observed that BIN1-associated Tau-containing extracellular vesicles purified from cerebrospinal fluid of AD-affected individuals are seeding-competent. We showed that BIN1 over-expression promotes the release of Tau via extracellular vesicles in vitro as well as exacerbation of Tau pathology in vivo in PS19 mice. Genetic deletion of Bin1 from microglia resulted in reduction of Tau secretion via extracellular vesicles in vitro, and in decrease of Tau spreading in vivo in male, but not female, mice, in the context of PS19 background. Interestingly, ablation of Bin1 in microglia of male mice resulted in significant reduction in the expression...

Identified as an Alzheimer’s disease (AD) susceptibility gene by genome wide-association studies,... more Identified as an Alzheimer’s disease (AD) susceptibility gene by genome wide-association studies, BIN1 has 10 isoforms that are expressed in the Central Nervous System (CNS). The distribution of these isoforms in different cell types, as well as their role in AD pathology still remains unclear. Utilizing antibodies targeting specific BIN1 epitopes in human postmortem tissue and analyzing RNA expression data from purified microglia, we identified three isoforms expressed specifically in neurons (isoforms 1, 2 and 3) and four isoforms expressed in microglia (isoforms 6, 9, 10 and 12). The abundance of selected peptides, which correspond to groups of BIN1 protein isoforms, was measured in dorsolateral prefrontal cortex, and their relation to neuropathological features of AD was assessed. Peptides contained in exon 7 of BIN1’s N-BAR domain were found to be significantly associated with AD-related traits and, particularly, tau pathology. Since only isoforms 1, 2 and 3 contain exon 7, it ...

Immunity, 2016

Microglia originate from erythromyeloid progenitors (EMPs) in the yolk sac and develop in the for... more Microglia originate from erythromyeloid progenitors (EMPs) in the yolk sac and develop in the forming CNS. Microglia are fundamental for the development and function of a healthy brain. By contrast, their role in immune host defense of the CNS remains speculative, given the immune privilege of this organ. Alterations in microglia functionality are involved in brain aging, as well as in neurodegenerative disease severity and progression. The combination of their ontogeny with the influence of the complex environment of the CNS makes microglia a unique cell population. Recent observations about microglia ontogeny combined with extensive gene expression profiling allow us to better capture the variety of nuances that microglia can manifest. Here, we provide a contemporary appraisal of microglial uniqueness based on their origin, functions, and expression profiles. Furthermore, we give an overview of the impact of aging and neurodegenerative diseases on microglia transcriptomes.

Alzheimer's & Dementia, 2019

Initiative (ADNI) participants, we derived 6 PRSs: 3 biomarkerPRSs (PRSA, PRST, PRSN), their comb... more Initiative (ADNI) participants, we derived 6 PRSs: 3 biomarkerPRSs (PRSA, PRST, PRSN), their combination (PRSA/T/N), a “Traditional” PRSTraditional, and an IGAP PRSIGAP-A/T/N (Figure 1, Table1). Specifically, we retained SNPs with GWAS p-value 5e-05. We calculated the correlation between SNPs, and to avoid collinearity we retained one SNP from each LD-block. Using the relevant endophenotype as the response variable, forward selection was applied for SNP selection and weight estimation. The APOE e4 SNP was forced into every PRS model by design. SNPs in PRSTraditional were selected by forward selection on Cox model. IGAP odds ratios used as PRSIGAP-A/T/N weights. Timedependent area under the curve (AUC) was used to assess the predictive performance of each PRSA, PRST, PRSN, PRSA/T/N, PRSIGAP-A/T/N PRSTraditional, as well as the simultaneous presence of PRSA, PRST, PRSN in a model. Results: Accounting simultaneously for PRSA, PRST, and PRSN in AD AAO prediction, achieves significantly better performance (Wilcoxon p-value < 2.2e-16) than the PRSTraditional, PRSIGAP-A/T/N or univariate PRSA, PRST, PRSN models (Figure 2). When assessed separately, PRSN has the best performance, followed by PRST, and PRSA. Conclusions: Individualized biomarker-specific PRS based on the patient’s biomarker profile may be more effective and informative compared to current PRS approaches. Further validation is warranted in larger samples. Optimized PRS profiling, after validation, may be a key supporting approach for the development of a precision medicine approach to AD.

Abstract: HIV-1 can be subdivided into distinct subtypes; the consequences of such a genomic vari... more Abstract: HIV-1 can be subdivided into distinct subtypes; the consequences of such a genomic variability remain largely speculative. The long terminal repeats (LTR) control HIV transcription and reflect the major differences of distinct viral subtypes. Three regions in the HIV-1 subtype B LTR are close matches to the Signal Transducer and Activator of Tran-scription (STAT) consensus sequence. Here, we show heterogeneity in these putative STAT binding sites among HIV-1 LTR subtypes A through G. Transfection of constitutively activated STAT5 lead to transcriptional activation of HIV-1 expression in 293T cells transfected with a reporter assay driven by HIV-1 LTR subtype B. Constitutively activated STAT5 transactivated the LTR of various subtypes in U937 cells with different potency. These findings support and ex-pand the potential relevance of STAT5 activation in HIV infection and may bear relevance for a differential regulation of latency and expression of different subtypes of HIV-1.

Post-entry events of efficient R5 vs. inefficient X4 HIV-1 replication

Trends in Immunology, 2015

Currently, the concept of &amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;am... more Currently, the concept of &amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;#39;neuroinflammation&amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;#39; includes inflammation associated with neurodegenerative diseases, in which there is little or no infiltration of blood-derived immune cells into the brain. The roles of brain-resident and peripheral immune cells in these inflammatory settings are poorly understood, and it is unclear whether neuroinflammation results from immune reaction to neuronal dysfunction/degeneration, and/or represents cell-autonomous phenotypes of dysfunctional immune cells. Here, we review recent studies examining these questions in the context of Huntington&amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;#39;s disease (HD), where mutant Huntingtin (HTT) is expressed in both neurons and glia. Insights into the cellular and molecular mechanisms underlying neuroinflammation in HD may provide a better understanding of inflammation in more complex neurodegenerative disorders, and of the contribution of the neuroinflammatory component to neurodegenerative disease pathogenesis.

Advances in Immunology, 2010

Chronic inflammation is associated with many neurodegenerative diseases, including multiple scler... more Chronic inflammation is associated with many neurodegenerative diseases, including multiple sclerosis, Parkinson&amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;#39;s disease, and Alzheimer&amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;#39;s disease. Increasing evidence that neuroinflammation contributes to disease severity has generated considerable interest in determining whether inhibition of inflammation pathways might be of therapeutic benefit. One potential avenue of intervention is provided by members of the nuclear receptor superfamily of ligand-dependent transcription factors that exert anti-inflammatory effects in many cell types. Here, we review recent studies providing insights into the distinct mechanisms that enable nuclear receptors to modulate immune responses, describe inflammatory components of neurodegenerative diseases, and discuss recent literature relevant to roles of nuclear receptors in influencing these processes.

Glia, 2013

Microglia cells function as sentinels for innate immunity in the central nervous system (CNS). To... more Microglia cells function as sentinels for innate immunity in the central nervous system (CNS). To perform this function, microglia express a diverse set of pattern recognition receptors (PRRs) for pathogen-associated molecular patterns (PAMPs) that include Toll-like receptors (TLRs) and inflammasomes. Several members of the TLR and inflammasome family also recognize endogenously derived molecules that are generated as a consequence of tissue injury or other pathological processes. Recognition of PAMPs or endogenous ligands by PRRs in microglia induces the robust activation of innate immune responses leading to the production of proinflammatory mediators and the activation of adaptive immunity. Activation of microglia is essential for clearance of infection and repair of tissue injury. However, uncontrolled inflammatory responses of microglia are also thought to contribute to the severity of many neurodegenerative diseases. Thus, activation of microglia must be properly and tightly regulated to maintain normal tissue homeostasis. Several mechanisms have been identified that appear to function in the active maintenance of quiescence under normal conditions and/or re-establish this state following resolution of infection or injury. These mechanisms involve communication with neurons and other glia through secreted molecules or surface expressing receptors as well as actions of members of the nuclear receptor (NR) superfamily of transcription factors. Here, we review recent advances in our understanding of the regulation of microglia activation and deactivation with a focus on counter-regulation of microglia activation by nuclear receptors. V