Biology subjects

Greene, C.

Publications and source records attributed to Greene, C..

11 recordsLinked to original sources

bioRxiv2025

Endothelial tight junctions and cell-matrix adhesions reciprocally control blood-brain barrier integrity

Brain endothelial cells (ECs) rely on mechanical cues to provide a physical barrier that protects the brain. Yet how ECs integrate forces to establish and maintain the blood-brain barrier (BBB) remains poorly understood. Here, we show that the two main endothelial force-bearing systems, tight junctions and cell-matrix adhesions, reciprocally control BBB integrity. Using a combination of super-resolution imaging and biophysical techniques, we reveal increasing mechanical loads on cell-cell junctions vs. cell-matrix adhesions in human stem cell-derived ECs during BBB maturation. This force redistribution is enabled by cytoskeletal remodeling, a compacted pattern of the tight junction protein claudin-5, and the emergence of specialised perinuclear cell-matrix adhesions. Mechanistically, we find an inverse relationship between claudin-5 levels and the expression of key cell-matrix adhesion proteins zyxin and vinculin in vitro and in mice. Finally, we demonstrate that this mechanobiological signature associated with BBB maturation is reversed upon BBB dysfunction after seizures in mice and in human patients with temporal lobe epilepsy. Collectively, our findings establish a novel interplay between mechanoresponsive elements in brain ECs, with implications for BBB stabilisation therapy in epilepsy.

neuroscience↗

bioRxiv2024

Selective modification of ascending spinal outputs in acute and neuropathic pain states

Pain hypersensitivity arises from the plasticity of peripheral and spinal somatosensory neurons, which modifies nociceptive input to the brain and alters pain perception. We utilized chronic calcium imaging of spinal dorsal horn neurons to determine how the representation of somatosensory stimuli in the anterolateral tract, the principal pathway transmitting nociceptive signals to the brain, changes between distinct pain states. In healthy conditions, we identify stable, narrowly tuned outputs selective for cooling or warming, and a neuronal ensemble activated by intense/noxious thermal and mechanical stimuli. Induction of an acute peripheral sensitization with capsaicin selectively and transiently retunes nociceptive output neurons to encode low-intensity stimuli. In contrast, peripheral nerve injury-induced neuropathic pain results in a persistent suppression of innocuous spinal outputs coupled with activation of a normally silent population of high-threshold neurons. These results demonstrate the differential modulation of specific spinal outputs to the brain during nociceptive and neuropathic pain states.

neuroscience↗

bioRxiv2023

The Secondary Somatosensory Cortex Gates Mechanical and Thermal Sensitivity

The cerebral cortex is vital for the perception and processing of sensory stimuli. In the somatosensory axis, information is received by two distinct regions, the primary (S1) and secondary (S2) somatosensory cortices. Top-down circuits stemming from S1 can modulate mechanical and cooling but not heat stimuli such that circuit inhibition causes blunted mechanical and cooling perception. Using optogenetics and chemogenetics, we find that in contrast to S1, an inhibition of S2 output increases mechanical and heat, but not cooling sensitivity. Combining 2-photon anatomical reconstruction with chemogenetic inhibition of specific S2 circuits, we discover that S2 projections to the secondary motor cortex (M2) govern mechanical and thermal sensitivity without affecting motor or cognitive function. This suggests that while S2, like S1, encodes specific sensory information, that S2 operates through quite distinct neural substrates to modulate responsiveness to particular somatosensory stimuli and that somatosensory cortical encoding occurs in a largely parallel fashion.

neuroscience↗

bioRxiv2020

Modulating the Blood-Brain Barrier by Light Stimulation of Molecular-Targeted Nanoparticles

The blood-brain barrier (BBB) tightly regulates the entry of molecules into the brain by tight junctions that seals the paracellular space and receptor-mediated transcytosis. It remains elusive to selectively modulate these mechanisms and to overcome BBB without significant neurotoxicity. Here we report that light stimulation of tight junction-targeted plasmonic nanoparticles selectively opens up the paracellular route to allow diffusion through the compromised tight junction and into the brain parenchyma. The BBB modulation does not impair vascular dynamics and associated neurovascular coupling, or cause significant neural injury. It further allows antibody and adeno-associated virus delivery into local brain regions. This novel method offers the first evidence of selectively modulating BBB tight junctions and opens new avenues for therapeutic interventions in the central nervous system. One Sentence SummaryGentle stimulation of molecular-targeted nanoparticles selectively opens up the paracellular pathway and allows macromolecules and gene therapy vectors into the brain.

neuroscience↗

bioRxiv2020

Methamphetamine enhances caveolar transport of therapeutic drugs across the rodent blood-brain barrier

The blood-brain barrier (BBB) is a multifactorial and multicellular vascular interface separating the systemic environment from the central nervous system (CNS). It gates cerebral penetration of circulating molecules and cells and is the principal reason for low accumulation of many therapeutics in the brain. Low dose methamphetamine (METH) induces fluid phase transcytosis across the BBB in vitro and could therefore be used to enhance CNS drug delivery. Here we show, that low dose intravascular METH induced significant leakage exclusively via caveolar transport at the intact BBB in rodents ex vivo. Notably, METH-induced leakage was suppressed at 4{degrees}C and in Caveolin-1 (CAV1) knockout mice. Furthermore, METH strongly enhanced brain penetration of therapeutic molecules, namely doxorubicin (DOX), a small chemotherapeutic agent, and aflibercept (AFL), a ca. 100 kDA recombinant protein. Lastly, METH improved the therapeutic efficacy of DOX in a mouse model of human glioblastoma (GBM), as measured by a 25% increase in median survival time (p = 0.0024). Collectively, our data indicated that METH can facilitate preclinical assessment of novel experimental treatments and has the potential to enhance drug delivery to the diseased CNS.

neuroscience↗

bioRxiv2020

Structural Disconnection of the Tool Use Network After Left Hemisphere Stroke Predicts Limb Apraxia Severity.

Producing a tool use gesture is a complex process drawing upon the integration of stored knowledge of tools and their associated actions with sensory-motor mechanisms supporting the planning and control of hand and arm actions. Understanding how sensory-motor systems in parietal cortex interface with semantic representations of actions and objects in the temporal lobe remains a critical issue, and is hypothesized to be a key determinant of the severity of limb apraxia, a deficit in producing skilled action after left hemisphere stroke. We used voxel-based and connectome-based lesion symptom mapping with data from 57 left hemisphere stroke participants to assess the lesion sites and structural disconnection patterns associated with poor tool use gesturing. We found that structural disconnection between the left inferior parietal lobule, lateral temporal lobe (left middle temporal gyrus) and ventral temporal cortex (left medial fusiform gyrus) predicted the severity of tool use gesturing performance. Control analyses demonstrated that reductions in right-hand grip strength were associated with motor system disconnection, bypassing regions supporting tool use gesturing. Our findings provide causal evidence that limb apraxia may arise, in part, from disconnection of conceptual representations in the temporal lobe from mechanisms enabling skilled action production in the inferior parietal lobule.

neuroscience↗

bioRxiv2019

Potent and lasting seizure suppression by systemic delivery of antagomirs targeting miR-134 timed with blood-brain barrier disruption

RNA therapies such as oligonucleotides (OGNs) offer precision treatments for a variety of neurological diseases, including epilepsy but their deployment is hampered by the blood brain barrier (BBB). Here we used brain imaging and assays of serum proteins and tracer extravasation, to determine that BBB disruption occurring after status epilepticus in mice was sufficient to permit passage of systemically-injected antisense OGNs targeting microRNA-134 (Ant-134) into the brain parenchyma. A single intraperitoneal injection of Ant-134 two hours after status epilepticus in mice resulted in potent suppression of spontaneous recurrent seizures, reaching a 99.5% reduction during recordings at three months. The duration of spontaneous seizures, when they occurred, was also reduced in Ant-134-treated mice. These studies indicate that systemic delivery of Ant-134 reaches the brain and produces disease-modifying effects after systemic injection in mice when timed with BBB disruption and may be a clinically-viable approach for this and other disease-modifying microRNA therapies.

neuroscience↗

bioRxiv2018

Specific histone modifications associate with alternative exon selection during mammalian development

Alternative splicing (AS) is frequent during early mouse embryonic development. Specific histone post-translational modifications (hPTMs) have been shown to regulate exon splicing by either directly recruiting splice machinery or indirectly modulating transcriptional elongation. In this study, we hypothesized that hPTMs regulate expression of alternatively spliced genes for specific processes during differentiation. To address this notion, we applied an innovative machine learning approach to relate global hPTM enrichment to AS regulation during mammalian tissue development. We found that specific histone modifications, H3K36me3 and H3K4me1, play a dominant role in skipped exon selection among all the tissues and developmental time points examined. In addition, we used iterative random forest model to identify interactions of several hPTMs that associated with skipped exon selection during tissue development. Collectively, our data demonstrated a link between hPTMs and alternative splicing which will drive further experimental studies on the functional relevance of these modifications to alternative splicing.

bioinformatics↗

bioRxiv2017

A Multimodal Strategy Used By A Large c-di-GMP Network

The Pseudomonas fluorescens genome encodes for 50+ proteins involved in -di-GMP signaling. Here, we demonstrate that when tested across 188 nutrients, these enzymes and effectors appear capable of impacting biofilm formation. Transcriptional analysis of network members across [~]50 nutrient conditions indicates that altered gene expression can explain a subset, but not all, of biofilm-formation responses to the nutrients. Additional organization of the network is likely achieved through physical interaction, as determined via probing [~]2000 interactions by bacterial two-hybrid assays. Our analysis revealed a multimodal regulatory strategy, using combinations of ligand-mediated signals, protein-protein interaction and/or transcriptional regulation to further fine-tune c-di-GMP-mediated responses. These results create a profile of a large c-di-GMP network that is used to make important cellular decisions, opening the door to future model building and the ability to engineer this complex circuitry in other bacteria.\n\nAbstract ImportanceCyclic diguanylate (c-di-GMP) is a key signalling molecule regulating bacterial biofilm formation, and many microbes have up to dozens of proteins that make, break or bind this dinucleotide. Thus, a major open question in the field is how signalling specificity is conferred in this context with a soluble signalling molecule. Here, we take a systems approach, using mutational analysis, transcriptional studies and bacterial two-hybrid analysis to interrogate this network. We find that the network typically combines two or more modes of regulation (i.e., transcriptional control with protein-protein interaction) to generate an observed output.

microbiology↗

bioRxiv2017

Enter the matrix: Interpreting unsupervised feature learning with matrix decomposition to discover hidden knowledge in high-throughput omics data

Omics data contains signal from the molecular, physical, and kinetic inter- and intra-cellular interactions that control biological systems. Matrix factorization techniques can reveal low-dimensional structure from high-dimensional data that reflect these interactions. These techniques can uncover new biological knowledge from diverse high-throughput omics data in topics ranging from pathway discovery to time course analysis. We review exemplary applications of matrix factorization for systems-level analyses. We discuss appropriate application of these methods, their limitations, and focus on analysis of results to facilitate optimal biological interpretation. The inference of biologically relevant features with matrix factorization enables discovery from high-throughput data beyond the limits of current biological knowledge--answering questions from high-dimensional data that we have not yet thought to ask.

systems biology↗

bioRxiv2017

Machine learning analysis identifies Drosophila Grunge/Atrophin as an important learning and memory gene required for memory retention and social learning.

High throughput experiments are becoming increasingly common, and scientists must balance hypothesis driven experiments with genome wide data acquisition. We sought to predict novel genes involved in Drosophila learning and long-term memory from existing public high-throughput data. We performed an analysis using PILGRM, which analyzes public gene expression compendia using machine learning. We evaluated the top prediction alongside genes involved in learning and memory in IMP, an interface for functional relationship networks. We identified Grunge/Atrophin (Gug/Atro), a transcriptional repressor, histone deacetylase, as our top candidate. We find, through multiple, distinct assays, that Gug has an active role as a modulator of memory retention in the fly and its function is required in the adult mushroom body. Depletion of Gug specifically in neurons of the adult mushroom body, after cell division and neuronal development is complete, suggests that Gug function is important for memory retention through regulation of neuronal activity, and not by altering neurodevelopment. Our study provides a previously uncharacterized role for Gug as a possible regulator of neuronal plasticity at the interface of memory retention and memory extinction.

genetics↗