Biology subjects

Cologna, S. M.

Publications and source records attributed to Cologna, S. M..

13 recordsLinked to original sources

bioRxiv2026

Immunohistochemistry-compatible gel-assisted mass spectrometry imaging

While lipids are fundamental to cellular signaling and structure, probing native lipid compositions and distributions across complex tissue architectures at single-cell resolution remains challenging due to inherent trade-offs in spatial resolution, sensitivity, and molecular coverage in current spatial lipidomics methods. Here, we report a multimodal imaging approach that seamlessly integrates gold-standard immunohistochemical labeling with gel-assisted mass spectrometry imaging to enable cell-type-specific, single-cell spatial lipidomics with modern instrumentation. Using intact brain tissue as a testbed, we demonstrate in situ measurement of cerebellar Purkinje cells and spatially mapped lipids across timepoints and cerebellar subregions within the pathological landscape of a neurodegenerative, lysosomal storage disorder. With single-cell lipidomic profiling, we delineated spatiotemporally distinct accumulation of specific glycosphingolipids and phospholipids within Purkinje cells and non-Purkinje cells in the diseased brain. Furthermore, unsupervised single-cell lipidomic clustering elucidated disease-progression-and subregion-dependent molecular divergence between healthy and neurodegenerative states within the cerebellum.

neuroscience↗

bioRxiv2026

Unveiling Lipid Dysregulation: Lipidomics of Mouse Brain and Isolated Myelin in Niemann - Pick Disease Type C1

Niemann-Pick Disease Type C1 (NPC1) is a fatal, neurodegenerative disorder, characterized by lysosomal lipid accumulation and dysmyelination. Previous studies have documented some lipid abnormalities in the null mouse (Npc1-/-) focused on the whole brain and liver. However, the specific lipidomic alterations in severely affected brain regions, such as cerebellum and isolated myelin remain understudied. We present a comprehensive LC-MS-based lipidomic analysis of the cerebellum and cortex of Npc1-/- mice during disease progression stages, along with the first comprehensive characterization of the myelin lipidome in NPC1 disease. Our results reveal that the cerebellum accumulates lipid species, including sphingolipids and glycerophospholipids progressively, while the cortex shows an overall decline in lipid levels, indicating region-specific lipid dysregulation. Notably, bis(monoacylglycero)phosphates and their precursors--including lysophosphatidylglycerol and hemibismonoacylglycerophosphate exhibit significant accumulation, with a preference for docosahexaenoic acid (DHA)-containing species. Despite known cholesterol storage defects in NPC1, we observed reduced free cholesterol levels in both regions, which we attribute to myelin loss. Myelin-specific lipidomics demonstrated extensive dysregulation, particularly in cortical myelin, including severe losses in sulfatides, ether-lipids, and acylcarnitine, alongside striking accumulation of hydroxy-ceramides. These findings identify novel lipid alterations in brain subregions and myelin, offering critical insight into the lipid perturbations under the loss of NPC1, and highlight lipid targets that may be crucial for therapeutic intervention and biomarker development.

biochemistry↗

bioRxiv2025

Multiomic Analysis Reveals Molecular Pathways Associated with Intestinal Aggregation of α-Synuclein

Aggregates of the protein -synuclein may initially form in the gut before propagating to the brain in Parkinsons disease. Indeed, our prior work supports that enteroendocrine cells, specialized intestinal epithelial cells, could play a key role in the development of this disease. Enteroendocrine cells natively express -synuclein and synapse with enteric neurons as well as the vagus nerve. Severing the vagus nerve reduces the load of -synuclein aggregates in the brain, suggesting that this nerve is a conduit for gut-to-brain spread. Enteroendocrine cells line the gut lumen, as such, they are in constant contact with metabolites of the gut microbiota. We previously found that when enteroendocrine cells are exposed to nitrite--a potent oxidant produced by gut bacterial Enterobacteriaceae--a biochemical pathway is initiated that results in -synuclein aggregation. Here, we determined that dopamine production is critical to this mechanism of nitrite-induced -synuclein aggregation. Using enteroendocrine cells, we modulated dopamine biosynthesis and profiled the cellular proteome and lipidome. Proteomic signatures in dopamine-free cells were distinctly different than in enteroendocrine cells, highlighting pathways relevant to intestinal development of Parkinsons disease. Intriguingly, we observed that enteroendocrine cells maintain viability upon exposure to nitrite and in the presence of -synuclein aggregates. This cellular robustness suggests that dopamine-producing enteroendocrine cells may be a reservoir of toxic -synuclein aggregates, which can spread through a prion-like process. As a possible antidote, our findings show that benserazide--a chemical inhibitor of dopamine biosynthesis--limits formation of these aggregates in enteroendocrine cells. These studies lay a foundation for mechanistically informed therapeutic targets to prevent intestinal formation of -synuclein aggregates before they spread to the brain. For Table of Contents Use Only O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=104 SRC="FIGDIR/small/672706v1_ufig1.gif" ALT="Figure 1"> View larger version (24K): org.highwire.dtl.DTLVardef@1668b83org.highwire.dtl.DTLVardef@1515382org.highwire.dtl.DTLVardef@41b1d2org.highwire.dtl.DTLVardef@1018049_HPS_FORMAT_FIGEXP M_FIG C_FIG

molecular biology↗

bioRxiv2024

Small molecule modulator of neuronal lysosome positioning and function resolves Alzheimers Disease-linked pathologies in cultured human neurons

Abnormal increase in axonal lysosome abundance is associated with multiple neurodegenerative diseases including Alzheimers disease. However, the underlying mechanisms and disease relevance are not fully understood. We have recently identified RH1115 as a small molecule modulator of the autophagy-lysosomal pathway that regulates lysosome positioning in neurons. This allowed us to manipulate neuronal lysosome distribution including in axons and interrogate its contribution to both optimal neuronal functioning and to disease pathology. We demonstrate that the small molecule not only rescues aberrant buildup of both axonal autophagic and lysosomal intermediates but also reduces secreted A{beta}42 levels in human iPSC-derived neurons lacking the lysosomal adaptor, JIP3. We thus demonstrate that restoring efficient axonal lysosome transport has an anti-amyloidogenic effect in human neurons and is a promising therapeutic strategy for Alzheimers disease. Furthermore, we show that the small molecule enhances neuronal lysosome degradation, requires the lysosomal adaptor JIP4 to rescue the axonal lysosome pathology in the JIP3 KO neurons and increases levels of the JIP4-interacting lysosomal membrane protein, TMEM55B. Lastly, treatment with the small molecule led to a striking rescue of locomotor defects in JIP3 KO zebrafish larvae. Thus, we have identified a small molecule which can be impactful in neurodegenerative diseases that have a lysosomal pathology and have determined its molecular targets in modulating axonal lysosome abundance.

cell biology↗

bioRxiv2024

Optimization of systemic AAV9 gene therapy in Niemann-Pick disease type C1 mice

Niemann-Pick disease, type C1 (NPC1) is a rare, fatal neurodegenerative disorder caused by pathological variants in NPC1, which encodes a lysosomal cholesterol transporter. FDA-approved treatments are limited and do not target the underlying genetic defect. Both systemic and central nervous system (CNS) delivery of AAV9-hNPC1 have shown significant disease amelioration in NPC1 murine models. To assess the impact of dose in null Npc1m1N/m1N mice, we systemically administered three different doses of AAV9-hNPC1 at 4 weeks old. Then, to assess the impact of age, we administered the medium dose before phenotypic onset or at early- or late-stage of disease progression (4, 6 or 8 weeks old, respectively). Higher vector doses and earlier treatment were associated with significantly increased lifespan, slower disease progression, and enhanced CNS transduction. In Npc1I1061T/I1061T mice, a model recapitulating a common human hypomorphic variant, similar benefits ensued. Our findings help define dose ranges, treatment ages, and efficacy in hypomorphic models of NPC1 deficiency and suggest that higher doses of AAV9-hNPC1 in pre-symptomatic disease states are likely to yield better outcomes in NPC1 individuals. Summary BlurbSystemic AAV9-hNPC1 gene therapy in null Npc1m1N mice at higher doses or with earlier administration and treatment of hypomorphic Npc1I1061T mice delays disease progression and increases lifespan.

neuroscience↗

bioRxiv2024

In Vivo Simvastatin and Brain Radiation in a Model of HER2+ Inflammatory Breast Cancer Brain Metastasis

PurposeInhibiting HMG-CoA reductase with simvastatin prevents breast cancer metastases in preclinical models and radiosensitizes monolayer and stem-like IBC cell lines in vitro. Given the extensive use of simvastatin worldwide and its expected penetration into the brain, we examined whether regulating cholesterol with simvastatin affected IBC3 HER2+ brain metastases. Methods and MaterialsBreast cancer cell lines KPL4 and MDA-IBC3 were examined in vitro for DNA repair after radiation with or without statin treatment. Brain metastasis endpoints were examined in the MDA-IBC3 brain metastasis model after ex vivo exposure to lipoproteins and after tail vein injections with and without whole-brain radiotherapy (WBR) and oral statin exposure. ResultsEx vivo preculture of MDA-IBC3 cells with very low-density lipoprotein (vLDL) enhanced the growth of colonized lesions in the brain in vivo compared with control or high-density lipoprotein (HDL), and concurrent oral simvastatin/ WBR reduced the incidence of micrometastatic lesions evaluated 10 days after WBR. However, statin, with or without WBR, did not reduce the incidence, burden, or number of macrometastatic brain lesions evaluated 5 weeks after WBR. ConclusionsAlthough a role for cholesterol biosynthesis is demonstrated in DNA repair and response to whole brain radiation in this model, durable in vivo efficacy of concurrent whole brain irradiation and oral statin was not demonstrated.

cancer biology↗

bioRxiv2023

Caveolin-1 Autonomously Regulates Hippocampal Neurogenesis Via Mitochondrial Dynamics

Hippocampal neurogenesis plays instrumental roles in learning and memory. However, the mechanisms underlying neurogenesis are not fully understood. Here we show that the expression of Caveolin-1 (Cav-1), the principal component of caveolae, peaks in neural progenitor cells (NPCs) during neurogenesis. Using NestinCreERT2;Cav-1fl/fl male mice, and CRISPR-sham (Cav-1 Ctrl) and CRISPR/Cas9-edited (Cav-1 KO) human induced-pluripotent stem cells, we observed that Cav-1 deletion led to reduced stem cell proliferation and enhanced differentiation into neurons. This was manifested by increased neuronal dendritic tree surface area and enhanced mouse performance in contextual discrimination. Proteomic analysis revealed that Cav-1 plays a role in mitochondrial pathways in NPCs. Cav-1 localized to the mitochondria in NPCs and co-immunoprecipitated with mitofusion 2. Mitochondrial morphology was elongated in Cav-1 KO NPCs and the expression of mitofusion 2 was increased in mitochondrial fractions. Restoration of Cav-1 levels rescued elongated mitochondrial morphology and altered neuronal differentiation in Cav-1 KO NPCs. Together, this study identifies Cav-1 as a novel regulator of neurogenesis and -dependent learning and memory. Significance StatementThe hippocampal dentate gyrus (DG) orchestrates adult hippocampal neurogenesis (AHN). The precise mechanisms governing AHN remain elusive. Caveolin-1 regulates neurogenesis through mitochondrial fission-fusion process, suggesting Caveolin-1 as a novel regulator of AHN and underscoring the impact of AHN on cognition.

neuroscience↗

bioRxiv2023

Gel-assisted mass spectrometry imaging

Compatible with label-free detection and quantification, mass spectrometry imaging (MSI) is a powerful tool for spatial investigation of biomolecules in intact specimens. Yet, the spatial resolution of MSI is limited by the methods physical and instrumental constraints, which often preclude it from single-cell and subcellular applications. By taking advantage of the reversible interaction of analytes with superabsorbent hydrogels, we developed a sample preparation and imaging workflow named Gel-Assisted Mass Spectrometry Imaging (GAMSI) to overcome these limits. With GAMSI, the spatial resolution of lipid and protein MALDI-MSI can be enhanced severalfold without changing the existing mass spectrometry hardware and analysis pipeline. This approach will further enhance the accessibility to (sub)cellular-scale MALDI-MSI-based spatial omics.

biochemistry↗

bioRxiv2023

DBDA matrix increases ion abundance of fatty acids and sulfatides in MALDI-TOF and mass spectrometry imaging studies

MALDI-TOF MS is a powerful tool to analyze biomolecules owing to its soft ionization nature and generally results in simple spectra of singly charged ions. Moreover, implementation of the technology in imaging mode provides a means to spatially map analytes in situ. Recently, a new matrix, DBDA (N1,N4-dibenzylidenebenzene-1,4-diamine) was reported to facilitate the ionization of free fatty acids in the negative ion mode. Building on this finding, we sought to implement DBDA for MALDI mass spectrometry imaging studies in brain tissue and successfully map oleic acid, palmitic acid, stearic acid, docosahexaenoic acid and arachidonic acid using mouse brain sections. Moreover, we hypothesized that DBDA would provide superior ionization for sulfatides, a class of sulfolipids, with multiple biological functions. Herein we also demonstrate that DBDA is ideal for MALDI mass spectrometry imaging of fatty acids and sulfatides in brain tissue sections. Additionally, we show enhanced ionization of sulfatides using DBDA compared to three different traditionally used MALDI matrices. Together these results provide new opportunities for studies to measure sulfatides by MALDI-TOF MS including in imaging modes.

biochemistry↗

bioRxiv2022

Probing the force-from-lipid mechanism with synthetic polymers

A central feature of mechanotransduction is the ability of mechanosensitive channels to respond to mechanical stimuli from the surrounding lipid bilayer. Accordingly, the mechanical properties of membranes should play an important role in modulating force transmission to embedded channels, yet the nature of this relationship remains unclear for a wide class of mechanosensitive channels across prokaryotic and eukaryotic systems. Here, we use a synthetic amphiphile to modulate the membrane mechanical properties of cell-derived vesicles and probe channel activation. Using precise membrane mechanical characterization approaches that have rarely been used in conjunction with electrophysiology techniques, we directly characterize three membrane properties and the activation threshold of the E. coli mechanosensitive channel of large conductance (MscL). Our study reveals that decreases in the membrane area expansion modulus, KA, and bending rigidity, kc, correlate with increases in the pressure required to activate MscL and that this effect is reproducible with the mammalian channel, TREK-1. MD simulations demonstrate that polymer-mediated changes in interfacial tension is the best mechanism to describe these experimental results. Together, our results bolster the force-from-lipids mechanism by demonstrating the generality of the relationship between changes in specific membrane mechanical properties and the gating pressure of MscL and TREK-1. In addition, our results reveal the mechanical mechanism by which membrane amphiphiles alter the activity and sensitivity of mechanosensitive channels through changes in long-range force transmission.

biophysics↗

bioRxiv2022

An integrated approach to protein discovery and detection from complex biofluids

Ovarian cancer, a leading cause of cancer related deaths among women, has been notoriously difficult to routinely screen for and diagnose early. Researchers and clinicians continue to seek routinely usable, non-invasive, screening methods as early detection significantly improves survival. Biomarker screening is ideal; however, currently available ovarian cancer biomarkers lack desirable sensitivity and specificity. Furthermore, the most fatal forms, high grade serous cancers often originate in the fallopian tube; therefore, sampling from the vaginal environment provides more proximal sources for tumor detection. To address these shortcomings and leverage proximal sampling, we developed an untargeted mass spectrometry microprotein profiling method and identified a signature of cystatin A, validated this protein in an animal model, and sought to overcome the limits of detection inherent to mass spectrometry by demonstrating that cystatin A is present at 100 pM concentrations using a label-free microtoroid resonator. The findings highlight the potential utility for early-stage detection where cystatin A levels would be low. Significance StatementIt is now clear that high-grade serous ovarian cancer can originate in the fallopian tube epithelium. These tumors colonize the ovary and then metastasize throughout the peritoneum. This discovery has raised important, and yet unaddressed, questions how we might be able to detect and screen for this deadly disease for which there is no routine screening. We have leveraged vaginal lavages from a murine model of the disease as a complex biological fluid for untargeted discovery of microproteins using mass. We improved our limits of detection by conjugating a cystatin A antibody to the surface of a microtoroid resonator to allow us to specifically detect cystatin A from vaginal lavages at early time points across biological replicates.

cancer biology↗

bioRxiv2021

Loss of Depalmitoylation Exaggerates Synaptic Upscaling and Leads to Neuroinflammation in a Lysosomal Storage Disease

Palmitoylation and depalmitoylation are the dichotomic processes of lipid modification regulating protein trafficking, recycling, and degradation, thereby controlling proteostasis. Despite our understanding of palmitoylation, depalmitoylation is far less studied. Here, we study a lysosomal depalmitoylating enzyme, palmitoyl-protein thioesterase 1 (PPT1), associated with the devastating neurodegenerative condition CLN1 disease and show that dark-rearing Ppt1-/- mice, which induces synaptic upscaling in vivo, worsen the symptoms. In Ppt1-/- cortical neurons, upscaling induction triggers exaggerated responses of synaptic calcium-permeable AMPA receptors composed of palmitoylated GluA1 subunits. Consequently, Ppt1-/- visual cortex exhibits hypersynchrony in vivo. Remarkably, we also find an overload of palmitoylated A-kinase anchor protein 5 (Akap5) in Ppt1-/- mouse brains, leading to microglial activation through NFAT. These findings indicate Ppt1 acts as a gatekeeper of homeostatic plasticity by regulating the proteostasis of palmitoylated synaptic proteins. Moreover, our results suggest that perturbed depalmitoylation results in neuroinflammation, which is common to neurodegenerative diseases.

neuroscience↗

bioRxiv2020

Visualising Cholesterol in Brain by On-Tissue Derivatisation and Quantitative Mass Spectrometry Imaging

Despite being a critical molecule for neurobiology and brain health, mass spectrometry imaging (MSI) of cholesterol has been under reported compared to other lipids, due to the difficulty in ionising the sterol molecule. In the present work we have employed an on-tissue enzyme-assisted derivatisation strategy to improve detection of cholesterol in brain tissue sections. We report distribution and levels of cholesterol across specific brain structures of the mouse brain, in a model of Niemann-Pick type C1 (NPC1) disease, and during brain development. MSI revealed how cholesterol changes during development and that in the adult is highest in pons and medulla of the brain stem. Cholesterol was significantly reduced in the corpus callosum and other brain regions in the Npc1 null mouse, confirming hypomyelination at the molecular level. Our study demonstrates the potential of MSI to the study of sterols in neuroscience.

biochemistry↗