Biology subjects

Furnari, J. L.

Publications and source records attributed to Furnari, J. L..

5 recordsLinked to original sources

bioRxiv2026

THE MICROANATOMY OF INTRACRANIAL HUMAN THROMBI BY SINGLE NUCLEI TRANSCRIPTOMICS

Rationale: Thrombi are fragile, heterogeneous, and difficult to study with available methods, yet understanding their molecular composition and local microenvironment in stroke is critical to elucidating stroke pathophysiology and improving patient outcomes. Objective: We used a novel platform for harvesting and analyzing intracranial thrombi from endovascular thrombectomy for large vessel occlusion to define the cellular and transcriptomic landscape of thrombi. Methods and results: We performed single-nuclei RNA sequencing (snRNA-seq) on three human intracranial thrombi to define their molecular composition and microenvironment. Across 5,143 nuclei, we identified 8 distinct cellular populations, including monocytes and T cells. Monocytes displayed transcriptional changes on gene and regulon levels suggestive of transdifferentiation. We integrated our snRNA-seq data with publicly available single cell RNA-sequencing (scRNA-seq) data from asymptomatic and symptomatic atherosclerotic carotid plaques, and scRNA-seq data from healthy brain vasculature. We identified shared transcriptional programs between carotid and thrombi monocytes along dimensions including inflammation and endothelial activation, supporting the notion that thrombi monocytes undergo transdifferentiation in stroke. We also inferred potential ligand-receptor interactions that may occur at the site and time of occlusion. Conclusion: Our novel application of snRNA-seq to thrombectomy specimens has allowed us to uncover the full diversity of cell types present in thrombi, including low-abundance and novel cell populations, and to provide evidence of cellular plasticity and intercellular communication at the site of occlusion. This detailed definition of the cellular populations involved in stroke will facilitate mapping of novel interventional targets with direct functional relevance and facilitate greater personalization of post-reperfusion therapy.

molecular biology↗

bioRxiv2026

Electrophysiologically Targeted Biopsies Reveal the Transcriptional Landscape of Focal Epilepsy

Up to 30% of patients with epilepsy have intractable seizures, yet the mechanisms of focal ictogenesis remain unclear. Tissue involvement in ictal regions is heterogeneous, with different regions playing distinct roles in ictogenesis, seizure propagation, and resistance to spread. These roles are reflected in electrophysiologic differences between the seizure focus and the ictal penumbra, where evidence of synaptic spread is present but excitatory firing is constrained by largely intact inhibition. Investigation of the disruption of the normal interplay between excitatory and inhibitory activity, thought to underlie ictogenesis across a range of epilepsy etiologies, is limited by network complexity and cellular heterogeneity in human tissue samples. In this study, we relate cellular and molecular alterations to excitatory-inhibitory disruption in network dynamics defined by electrophysiologic features. This work may aid in the identification of clinically relevant tissue biomarkers and support novel preclinical therapeutic approaches for treatment-resistant focal epilepsy disorders. We developed a novel intracranial EEG guided, MRI-localized approach to sample paired biopsies from 11 patients with drug-resistant focal epilepsy with diverse etiologies, which were then studied using single-nucleus RNA sequencing (snRNAseq) and immunohistochemistry (IHC). EEG recorded from stereotactically implanted depth arrays (sEEG) was used to identify regions of epileptic involvement, based on findings from prior simultaneous clinical and microelectrode recordings. This approach addresses the intrinsic heterogeneity due to etiology and cortical architecture through paired, within-patient comparisons. We identified distinct cell-type specific transcriptional signatures that differentiate cellular populations in the seizure focus and ictal penumbra in intractable focal epilepsies. Our findings provide a link between tissue composition and gene expression that correlate with electrographic features in a heterogeneous seizure landscape. Our findings support common pathways of seizure generation and spread that are conserved across disease etiologies. Relative depletion of interneuron populations in the seizure focus supports the hypothesis of disrupted inhibition as a driver of epileptiform activity in the seizure focus. The enrichment of plasticity-associated gene signatures in the penumbra suggests a complex interaction of these regions with the seizure focus, as well as the role of the penumbra in enabling or limiting seizure expansion. This study provides a novel methodology for tissue sampling in epilepsy and uncovers biologically relevant tissue signatures that provide grounds for future work in targeting cellular and molecular alterations present in focal epilepsies.

neuroscience↗

bioRxiv2025

A proteogenomic tool uncovers protein markers for human microglial states

Human microglial heterogeneity has been largely described using transcriptomic data. Here, we introduce a microglial proteomic data resource and a Cellular Indexing of Transcriptomes and Epitopes by Sequencing panel enhanced with antibodies targeting 17 microglial cell surface proteins (mCITE-Seq). We evaluated mCITE-Seq on HMC3 microglia-like cells, induced-pluripotent stem cell-derived microglia (iMG), and freshly isolated primary human microglia. We identified novel protein microglial markers such as CD51 and relate expression of 101 cell surface proteins to transcriptional programs. This results in the identification and validation of three protein marker combinations with which to purify microglia enriched with each of 23 transcriptional programs; for example, CD49D, HLA-DR and CD32 enrich for GPNMBhigh ("disease associated") microglia. Further, we identify and validate proteins - SIRPA, PDPN and CD162 - that differentiate microglia from infiltrating macrophages. The mCITE-Seq panel enables the transition from RNA-based classification and facilitates the functional characterization and harmonization of model systems.

neuroscience↗

bioRxiv2024

A pharmacological toolkit for human microglia identifies Topoisomerase I inhibitors as immunomodulators for Alzheimer's disease

AbstractWhile efforts to identify microglial subtypes have recently accelerated, the relation of transcriptomically defined states to function has been largely limited to in silico annotations. Here, we characterize a set of pharmacological compounds that have been proposed to polarize human microglia towards two distinct states - one enriched for AD and MS genes and another characterized by increased expression of antigen presentation genes. Using different model systems including HMC3 cells, iPSC-derived microglia and cerebral organoids, we characterize the effect of these compounds in mimicking human microglial subtypes in vitro. We show that the Topoisomerase I inhibitor Camptothecin induces a CD74high/MHChigh microglial subtype which is specialized in amyloid beta phagocytosis. Camptothecin suppressed amyloid toxicity and restored microglia back to their homeostatic state in a zebrafish amyloid model. Our work provides avenues to recapitulate human microglial subtypes in vitro, enabling functional characterization and providing a foundation for modulating human microglia in vivo.

neuroscience↗

bioRxiv2023

A cell state specific metabolic vulnerability to GPX4-dependent ferroptosis in glioblastoma

Glioma cells hijack developmental transcriptional programs to control cell state. During neural development, lineage trajectories rely on specialized metabolic pathways. However, the link between tumor cell state and metabolic programs is poorly understood in glioma. Here we uncover a glioma cell state-specific metabolic liability that can be leveraged therapeutically. To model cell state diversity, we generated genetically engineered murine gliomas, induced by deletion of p53 alone (p53) or with constitutively active Notch signaling (N1IC), a pathway critical in controlling cellular fate. N1IC tumors harbored quiescent astrocyte-like transformed cell states while p53 tumors were predominantly comprised of proliferating progenitor-like cell states. N1IC cells exhibit distinct metabolic alterations, with mitochondrial uncoupling and increased ROS production rendering them more sensitive to inhibition of the lipid hydroperoxidase GPX4 and induction of ferroptosis. Importantly, treating patient-derived organotypic slices with a GPX4 inhibitor induced selective depletion of quiescent astrocyte-like glioma cell populations with similar metabolic profiles.

cancer biology↗