Biology subjects

Canoll, P. D.

Publications and source records attributed to Canoll, P. D..

9 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

Disrupted O-GalNAc glycosylation as a mechanism and biomarker of SLC35A2-associated epilepsy

Rare germline and somatic variants in SLC35A2 cause a spectrum of severe glycosylation disorders that commonly present with epilepsy. SLC35A2 encodes the Golgi transporter for UDP-galactose, but how its deficiency leads to severe neurodevelopmental disorders is unknown. Using a mouse model deficient for Slc35a2 in the forebrain, we identified a specific defect in O-GalNAc glycan synthesis, while other galactose-containing glycoconjugates remained intact. O-GalNAc glycans were absent from their normal location within neuronal tracts of the corpus callosum, and truncated precursors accumulated in the cortex on critical extracellular matrix molecules. Cultured primary neurons lacking Slc35a2 showed impaired development, hyperexcitability, and impaired O-GalNAc glycosylation. Finally, human brain tissue from cases of SLC35A2-associated intractable epilepsy displayed a strong correlation between variant burden and truncated O-GalNAc glycans. These findings provide a mechanistic link between genetic causes of SLC35A2-associated epilepsy and protein O-glycosylation that can be targeted for biomarker and therapeutic development.

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↗

bioRxiv2022

FLASH and Conventional Radiation Induce Differential Immune Responses in Diffuse Intrinsic Pontine Glioma, Highlighting Potential for Combination Immunotherapy

PurposeDiffuse Midline Glioma (DMG) is a fatal tumor traditionally treated with radiotherapy (RT) and previously characterized as having a non-inflammatory tumor immune microenvironment (TIME). FLASH is a novel RT technique using ultra-high dose rate, which is associated with decreased toxicity and effective tumor control. However, the effect of FLASH and conventional (CONV) RT on the DMG TIME have not yet been explored. MethodsHere, we perform single-cell RNA sequencing and flow cytometry on immune cells isolated from an orthotopic syngeneic murine model of brainstem DMG following the use of FLASH (90Gy/sec) or CONV (2Gy/min) dose-rate RT, and compare to unirradiated tumor (SHAM). ResultsAt day 4 post-RT, FLASH exerts similar effects as CONV in the predominant microglial (MG) population, including the presence of two activated subtypes. However, at day 10 post-RT, we observe a significant increase in type 1 interferon alpha receptor (IFNAR+) in MG in CONV and SHAM compared to FLASH. In the non-resident myeloid clusters of macrophages (MACs) and dendritic cells (DCs), we find increased type 1 interferon (IFN1) pathway enrichment for CONV compared to FLASH and SHAM by scRNA-seq. We observe this trend by flow cytometry at day 4 post-RT in IFNAR+ MACs and DCs, which equalizes by day 10 post-RT. DMG control and murine survival are equivalent between RT dose rates. ConclusionOur work is the first to map CONV and FLASH immune alterations of the DMG TIME with single-cell resolution. While DMG tumor control and survival are similar between CONV and FLASH, we find that changes in immune compartments differ over time. Importantly, while both RT modalities increase IFN1, we find that the timing of this response is cell-type and dose-rate dependent. These temporal differences, particularly in the context of tumor control, warrant further study.

immunology↗

bioRxiv2022

SARS-CoV-2 infection results in lasting and systemic perturbations post recovery

SARS-CoV-2 has been found capable of inducing prolonged pathologies collectively referred to as Long-COVID. To better understand this biology, we compared the short- and long-term systemic responses in the golden hamster following either SARS-CoV-2 or influenza A virus (IAV) infection. While SARS-CoV-2 exceeded IAV in its capacity to cause injury to the lung and kidney, the most significant changes were observed in the olfactory bulb (OB) and olfactory epithelium (OE) where inflammation was visible beyond one month post SARS-CoV-2 infection. Despite a lack of detectable virus, OB/OE demonstrated microglial and T cell activation, proinflammatory cytokine production, and interferon responses that correlated with behavioral changes. These findings could be corroborated through sequencing of individuals who recovered from COVID-19, as sustained inflammation in OB/OE tissue remained evident months beyond disease resolution. These data highlight a molecular mechanism for persistent COVID-19 symptomology and characterize a small animal model to develop future therapeutics.

microbiology↗

bioRxiv2021

Hyperoxia Inhibits the Growth of Mouse Forebrain Oligodendrocyte Progenitors

NG2 chondroitin sulfate proteoglycan positive oligodendrocyte progenitor cells (OPCs) reside throughout the brain. They divide asymmetrically and differentiate into myelinating oligodendrocytes throughout adulthood. OPCs have been successfully isolated from rodents using several techniques including magnetic beads, immunopanning and exploiting differential centripetal adhesion. Whereas rat OPCs are relatively simple to propagate in vitro, it has been difficult to expand mouse OPCs. Therefore, we evaluated the effects of oxygen levels, growth factors and extracellular matrix components to produce a simple and reproducible method to prepare large numbers of nearly homogenous cultures of primary mouse OPCs from postnatal day 0-2 mouse telencephala. Using the McCarthy and de Vellis mechanical separation method OPCs were separated from mixed culture of glial cells. When the OPCs were plated onto fibronectin coated tissue culture plates in a biochemically defined medium that contained fibroblast growth factor-2 (FGF-2) and platelet derived growth factor AA (PDGFAA), and they were maintained in a standard tissue culture incubator, they proliferated very slowly. By contrast, mouse OPCs doubled approximately every 7 days when maintained in a 2% oxygen, nitrogen buffered environment. After 3 passages, greater than 99% of these OPCs were NG2+/PDGFR+. In medium containing only FGF-2, mouse OPCs progressed to late stage OPCs whereupon A2B5 expression decreased and O4 expression increased. When these cells were differentiated between passages 1 and 3, the majority of the OPCs differentiated into MBP+ mature oligodendrocytes However, cells that were repeatedly passaged beyond 4 passages progressed to a late O4+ OPC (even with mitogens present) and when differentiated by mitogen removal a minority of the OPCs differentiated into MBP+ cells. These studies reveal significant differences between mouse and rat OPCs and an inhibitory role for oxygen in mouse OPC proliferation.

neuroscience↗

bioRxiv2019

Sex Differences in Predictors of Seizure in Contrast-Enhancing Gliomas at Clinical Presentation: A Network Approach

BackgroundBrain tumor related epilepsy (BTE) is a major co-morbidity related to the management of patients with brain cancer. Despite published practice guidelines recommending against anti-epileptic drug (AED) utilization in patients with gliomas, there is heterogeneity in prescription practices of AEDs in these patients. In an attempt to impact BTE management, we statistically analyzed clinically relevant attributes (sex, age, tumor size, tumor growth kinetics, and tumor location) pertaining to seizure at presentation and used them to build a computational machine learning model to predict the probability of a seizure (at presentation).\n\nMethodsFrom our clinical data repository, we identified 223 patients (females, n=86; males, n=137) with pathologically-determined glioma and known seizure status at clinical presentation. Non-parametric and Fishers Exact tests were used to identify statistical differences in clinical characteristics. We utilized a random forest machine learning method for generating our predictive models by entire cohort and separated by male and female.\n\nFindingsPatients were divided into those that presented with seizure (SP, n=96, 43%; F, n= 28; M, n= 68) and those that presented without seizure (nSP, n=127, 57%, F n=58, M n=69). Females presented with seizures significantly less often than males (x2=6{middle dot}28, p=0{middle dot}01). SP patients had significantly smaller T1Gd radius compared to nSP (SP 11{middle dot}30mm, nSP 18.66mm, p<0{middle dot}0001). Tumor size and patient age were significant negative predictors for SP; patients with larger tumors, older age and less tumor diffusivity (p/D) were at lower risk for SP.\n\nInterpretationDespite heterogeneity across our patient cohort, there is strong evidence of a role for patient sex, tumor size, tumor invasion, and patient age in predicting the incidence of seizures at diagnosis. Future studies, with prospectively detailed data collection, may provide clearer insights into the incidence of seizures through a patients treatment course.

cancer biology↗

bioRxiv2019

CD8+ T-cell-mediated immunoediting influences genomic evolution and immune evasion in murine gliomas

Cancer immunoediting shapes tumor progression by the selection of tumor cell variants that can evade immune recognition. Given the immune evasion and intra-tumor heterogeneity intrinsic to gliomas, we hypothesized that CD8+ T-cells mediate immunoediting in these tumors. We evaluated glioma progression in the absence of CD8+ T-cells by depleting this immune cell population in transgenic murine gliomas. Upon transplantation, gliomas that developed in the absence of CD8+ T-cells engrafted poorly in recipients with intact immunity but engrafted well in those with CD8+ T-cell depletion. Gliomas developed in absence of CD8+ T-cells exhibited increased chromosomal instability, MAPK signaling, gene fusions, and macrophage/microglial infiltration. MAPK activation correlated with macrophage/microglial recruitment in this model and in the human disease. Our results indicate that CD8+ T-cells mediate immunoediting during gliomagenesis, influencing the genomic stability of glioma and its microenvironment, leading to immune evasion.\n\nSignificanceImmune evasion renders cancer resistant to anti-tumoral immunity. Therapeutic intervention often fails for gliomas because of the plasticity of tumor cell variants that resist immune surveillance. Our results demonstrate a mechanism of immune evasion in gliomas that derives from CD8+ T-cells during the development and progression of this disease.

cancer biology↗

bioRxiv2018

Quantitative Assessment of Protein Activity in Orphan Tissues and Single Cells Using the metaVIPER Algorithm

We and others have shown that transition and maintenance of biological states is controlled by master regulator protein, which can be inferred by interrogating tissue-specific regulatory models (interactomes) with transcriptional signatures, using the VIPER algorithm. Yet, some tissues may lack molecular profiles necessary for interactome inference (orphan tissues), or, as for single cells isolated from heterogeneous samples, their tissue context may be undetermined. To address this problem, we introduce metaVIPER, a novel algorithm designed to assess protein activity in tissue-independent by integrative analysis of multiple, non-tissue-matched interactomes. This assumes that transcriptional targets of each protein will be recapitulated by one or more available interactome. We confirmed the algorithms value in assessing protein dysregulation induced by somatic mutations, as well as in assessing protein activity in orphan tissues and, most critically, in single cells, thus allowing transformation of noisy and potentially biased RNA-Seq signatures into reproducible protein-activity signatures.

systems biology↗