Biology subjects

Gordon, T.

Publications and source records attributed to Gordon, T..

13 recordsLinked to original sources

bioRxiv2026

A Nerve-Dependent NGF Receptor Switch Controls Corneal Epithelial Renewal In Neurotrophic Keratopathy

Corneal epithelial integrity depends on continuous epithelial renewal by limbal epithelial stem cells (LESCs), a process tightly linked to sensory innervation. Loss or impairment of innervation causes neurotrophic keratopathy (NK), a sight-threatening degenerative disease for which rhNGF, the only FDA-approved pharmacologic therapy, often has limited efficacy in advanced or refractory disease. The mechanistic basis for this limited response remains unclear. Using surgical, genetic, and pharmacologic approaches in a rodent model of NK with corneal Schwann cell ablation or structural and functional denervation, together with primary human LESCs, we examined how denervation alters NGF receptor signaling during epithelial repair. In innervated corneas, NGF promoted epithelial regeneration through TrkA. Denervation, however, increased expression of a second NGF receptor, anti-regenerative p75NTR, and activation of its effector JNK, and reduced the activity of the TrkA effector AKT in LESCs. In this altered receptor context, denervation-induced elevation of endogenous NGF amplified p75NTR signaling, thereby explaining the failure of topical rhNGF to rescue severely denervated NK phenotype corneas. Conversely, selective TrkA activation, either with the clinical-stage agonist tavilermide, or pharmacologic or genetic inhibition/ablation of p75NTR, restored AKT signaling and rescued epithelial healing in denervated corneas independent of reinnervation. These findings identify a nerve-dependent NGF receptor switch as a key regulator of corneal epithelial renewal and establish receptor-selective modulation as a mechanistically rational therapeutic strategy for treating NK. One Sentence SummaryCorneal denervation shifts limbal epithelial stem cell signaling from pro-regenerative TrkA-AKT toward anti-regenerative p75NTR-JNK, explaining the limited efficacy of recombinant human nerve growth factor (rhNGF; cenegermin), the only approved pharmacologic therapy for neurotrophic keratopathy, in severely denervated corneas and identifying receptor-selective modulation as a mechanistically distinct therapeutic strategy.

cell biology↗

bioRxiv2025

Regional Signaling Controls Stem Cell-Mediated Regeneration in an Invertebrate Chordate

Many tissues harbor quiescent stem cells that activate after injury, yet how local signals regulate this transition is not well understood. The solitary ascidian Ciona robusta provides a unique model, as bottom body fragments regenerate while upper fragments fail to do so. By comparing these regenerative and non-regenerative contexts, we reveal striking differences in transcriptional dynamics and signaling environments. Combining flow cytometry, scRNA-seq, transplantation, and fate mapping, we identified a candidate stem cell population with robust proliferative and differentiation potential following transplantation. However, regenerative capacity does not simply reflect stem cell abundance, but instead depends on region-specific signaling cues. Local expression of metabolic, immune and differentiation-related factors further underscores the importance of spatially distinct environments in shaping outcomes. Our findings show how a shared injury response can diverge into regeneration versus failure, highlighting principles that may be leveraged to enhance tissue repair in other systems.

developmental biology↗

bioRxiv2025

Lower Airway Dysbiosis in NTM+ Bronchiectasis is Associated with NET-Predominant Severe Phenotypes

RationaleThe discoveries of neutrophilic inflammation and Pseudomonas-dominant pulmonary dysbiosis have helped pave the way for host-directed therapy in bronchiectasis. Substantial knowledge gaps remain about the interplay between neutrophilic signatures and microbes in non-tuberculous mycobacterial lung disease (NTM-LD), a phenotypically diverse lung infection that is increasingly prevalent in the United States and other parts of the world. ObjectivesEvaluate the lower airway microbiota and neutrophilic traits in NTM- and NTM+ bronchiectasis. Methods16S rRNA gene sequencing, cell counts and neutrophil extracellular trap (NET) immunoassays were performed on bronchoscopic lower airway samples in 200 bronchiectasis subjects (108 NTM-, 92 NTM+). A preclinical model of oral commensal micro-aspiration and NTM infection was used to profile the murine lower airways with flow cytometry and a NET assay. Measurements and Main ResultsLower airways of NTM+ bronchiectasis patients were enriched with Mycobacterium and oral commensals (e.g., Veillonella, Prevotella). NET levels were higher in NTM+ BAL. Mycobacterium and oral commensals co-occurred with NET and neutrophils in network studies. Distinct oral commensal taxa associated with severe disease phenotypes such as cavitary disease and exacerbators. In a murine micro-aspiration model, the combination of oral commensals and Mycobacterium led to a sustained pro-inflammatory immune response marked by an increase in Th17, {gamma}8T cells, PD-1+ T lymphocytes as well as higher NET levels. ConclusionsOur analyses showed that distinct microbiome features beyond the primary pathogen can contribute to neutrophilic inflammation and severe disease phenotypes in bronchiectasis/ NTM-LD.

immunology↗

bioRxiv2025

Germline stem cell isolation, lineage tracing, and aging in a protochordate

Germline stem cells (GSCs), the source of gametes, are the only stem cells capable of passing genes to future generations and are therefore considered units of natural selection. Yet, the factors that influence GSC fitness, and thus govern GSC competition, which exist in both protochordates and mammals, remain poorly understood. We studied how aging affects GSC fitness in the protochordate Botryllus schlosseri, an evolutionary crosspoint between invertebrates and vertebrates. GSCs were isolated and distinguished from developing and mature gametes using flow cytometry and scRNA-Seq, facilitated by a new PacBio genome assembly. Moreover, their function was validated through a novel lineage tracing approach that combines membrane-labeled GSC transplantation with scRNA-Seq. Leveraging our method to isolate them, single-cell transcriptomics showed significant age-related changes between young and old GSCs. Spermatids and sperm, however, showed minimal changes, suggesting that reproductive aging is governed by GSCs rather than by gametes. Reduced expressions of markers like DDX4 and PIWIL1 in aged GSCs mirrored trends in mammalian datasets, pointing to a conserved GSC-driven aging mechanism across chordate evolution. This study provides new techniques that lay the foundation to investigate further drivers of GSC fitness and highlights fertility-related genes as promising targets for therapies to preserve reproductive health.

cell biology↗

bioRxiv2024

sc-SPLASH provides ultra-efficient reference-free discovery in barcoded single-cell sequencing

Typical high-throughput single-cell RNA-sequencing (scRNA-seq) analyses are primarily conducted by (pseudo)alignment, through the lens of annotated gene models, and aimed at detecting differential gene expression. This misses diversity generated by other mechanisms that diversify the transcriptome such as splicing and V(D)J recombination, and is blind to sequences missing from imperfect reference genomes. Here, we present sc-SPLASH, a highly efficient pipeline that extends our SPLASH framework for statistics-first, reference-free discovery to barcoded scRNA-seq (10x Chromium) and spatial transcriptomics (10x Visium); we also provide its optimized module for preprocessing and k-mer counting in barcoded data, BKC, as a standalone tool. sc-SPLASH rediscovers known biology including V(D)J recombination and cell-type-specific alternative splicing in human and trans-splicing in tunicate (Ciona) and when applied to spatial datasets, detects sequence variation including tumor-specific somatic mutation. In sponge (Spongilla) and tunicate (Ciona), we uncover secreted repeat proteins expressed in immune-type cells and regulated during development; the sponge genes were absent from the reference assembly. sc-SPLASH provides a powerful alternative tool for exploring transcriptomes that is applicable to the breadth of lifes diversity.

bioinformatics↗

bioRxiv2023

ERK1/2 inhibition disrupts alcohol memory reconsolidation and prevents relapse

Relapse to alcohol abuse after periods of abstinence, often caused by cue-induced alcohol craving, is a major challenge in the treatment of alcohol addiction. Therefore, disruption of the cue-alcohol associative memories can diminish the risk of relapse. Upon retrieval, memories become temporarily labile before they reconsolidate in a process that requires protein synthesis. Accumulating evidence suggests that the mammalian target of rapamycin complex 1 (mTORC1), which is responsible for the translation of a subset of dendritic proteins, is crucial for memory reconsolidation. Here, we explored the involvement of two regulatory pathways of mTORC1, namely phosphoinositide 3-kinase (PI3K)-AKT and extracellular regulated kinase1/2 (ERK1/2), in the reconsolidation process in a rat model of non-operant alcohol self-administration. We found that retrieval of alcohol memories using an odor-taste cue increased ERK1/2 activation in the amygdala, but did not affect the PI3K-AKT pathway. Importantly, inhibition of ERK1/2 shortly after alcohol memory retrieval impaired reconsolidation and led to long-lasting suppression of relapse to alcohol drinking. Additionally, we show that attenuation of alcohol memories and relapse was also induced by post-retrieval administration of lacosamide, an inhibitor of collapsin response mediator protein-2 (CRMP2) - a translational product of mTORC1 that is functionally regulated by PI3K-AKT signaling. Together, our findings provide evidence for the crucial role of ERK1/2 and CRMP2 in the reconsolidation of alcohol memories, and mark the FDA-approved drug, lacosamide, as a potential treatment for alcohol use disorder.

neuroscience↗

bioRxiv2023

Methods for cell isolation and analysis of the highly regenerative tunicate Polycarpa mytiligera

Polycarpa mytiligera is the only molecularly characterized solitary ascidian capable of regenerating all organs and tissue types. The cellular basis for regeneration in P. mytiligera is largely unknown, and methods for isolating live cells from this species for functional analyses are unavailable. Here, we developed a method for isolating live cells from P. mytiligera, overcoming major experimental challenges, including the dissociation of its thick body wall and native cellular autofluorescence. We demonstrated the applicability of our approach for tissue dissociation and cell analysis using three flow cytometry platforms, and by using broadly used non-species-specific cell labeling reagents. In addition to live cell isolation, proof-of-concept experiments showed that this approach was compatible with gene expression analysis of RNA extracted from the isolated cells, and with ex vivo analysis of phagocytosis. The ability to purify live cells will promote future studies of cell function in P. mytiligera regeneration.

developmental biology↗

bioRxiv2021

A Biodegradable, Tacrolimus-releasing Nerve Wrap Promotes Peripheral Nerve Regeneration

Axonal regeneration following nerve repair is slow and often incomplete, resulting in poor functional recovery and sometimes lifelong disability. Yet, there are no FDA-approved therapies available to promote nerve regeneration. Tacrolimus accelerates axonal regeneration, but systemic side-effects presently outweigh its potential benefits for peripheral nerve surgery. We have developed a biodegradable drug delivery system for the sustained local release of tacrolimus at the nerve repair site, with suitable properties for large-scale manufacturing and clinical application, aiming to promote axonal regeneration and functional recovery with minimal systemic drug exposure. Tacrolimus is encapsulated in polycarbonate-urethane nanofibers and electrospun to generate an implantable nerve wrap that releases therapeutic doses of bioactive tacrolimus over 31 days. Size and drug loading are adjustable for applications in small and large caliber nerves, and the wrap degrades within 120 days into biocompatible byproducts. Tacrolimus released from the nerve wrap promotes axon elongation in vitro and accelerates nerve regeneration and functional recovery in preclinical nerve repair models while systemic drug exposure is reduced by 80% compared to systemic delivery. Given its surgical suitability and preclinical efficacy and safety, this system may provide a readily translatable approach to support axonal regeneration and recovery in patients undergoing nerve surgery.

neuroscience↗

bioRxiv2021

Stemness activity underlying whole brain regeneration in a basal chordate

Central nervous system (CNS) regeneration extent is highly diverse across the metazoans, with adult mammals demonstrating limited ability1,2. Understanding how neurons regenerate following injury remains a central challenge in regenerative medicine. Although conserved pathways associated with neural regeneration have been identified3,4, a study describing the stepwise morphogenetic changes that take place throughout a complete CNS regeneration is lacking. Utilizing the highly regenerative tunicate model Polycarpa mytiligera5, we characterized the morphological, cell proliferation, and transcriptomic dynamics that lead to entire CNS regeneration. The regenerated CNS of adult P. mytiligera expressed key neurodevelopmental markers that are not otherwise present in the adult CNS. Removal of the entire CNS resulted in high cell proliferation in the regenerated area. Transcriptome analysis revealed enhanced stem-cell related gene activity, with high expression of P53 and piRNA pathways preceding the activation of Notch, Wnt, and Nanos pathways. The CNS regeneration atlas created here depicts the transcriptomic landscape of the entire CNS regeneration process, revealing the core pathways that regulate neuronal response to injury, and the regeneration stage at which they are most pronounced. The molecular and cellular mechanisms controlling regenerative capacity that this atlas reveals could be used to develop approaches to enhancing neurogenesis in closely-related chordate species, including humans.

developmental biology↗

bioRxiv2021

Optical Tissue Clearing Enables Rapid, Precise and Comprehensive Assessment of Three-Dimensional Morphology in Experimental Nerve Regeneration Research

Morphological analyses are key outcome assessments for nerve regeneration studies but are historically limited to tissue sections. Novel optical tissue clearing techniques enabling three-dimensional imaging of entire organs at a subcellular resolution have revolutionized morphological studies of the brain. To extend their applicability to experimental nerve repair studies we adapted these techniques to nerves and their motor and sensory targets in rats. The solvent-based protocols rendered harvested peripheral nerves and their target organs transparent within 24 h while preserving tissue architecture and fluorescence. Optical clearing is compatible with conventional laboratory techniques, including retrograde labelling studies, and computational image segmentation, providing fast and precise cell quantitation. Further, optically cleared organs enable three-dimensional morphometry at an unprecedented scale including dermatome-wide innervation studies, tracing of intramuscular nerve branches and mapping of neurovascular networks. Given their wide-ranging applicability, rapid processing times and low costs, tissue clearing techniques are likely to be a key technology for next-generation nerve repair studies.

neuroscience↗

bioRxiv2020

Global Age-Specific Patterns of Cyclic Gene Expression Revealed by Tunicate Transcriptome Atlas

Expression levels of circadian clock genes, which regulate 24-hour rhythms of behavior and physiology, have been shown to change with age. However, a study holistically linking aging and circadian gene expression is missing. Using the colonial chordate Botryllus schlosseri, we combined transcriptome sequencing and stem cell-mediated aging phenomena to test how circadian gene expression changes with age. This revealed that B. schlosseri clock and clock-controlled genes oscillate organism-wide, with daily, age-specific amplitudes and frequencies. These age-related, circadian patterns persist at the tissue level, where dramatic variations in cyclic gene expression of tissue profiles link to morphological and cellular aging phenotypes. Similar cyclical expression differences were found in hundreds of pathways associated with known hallmarks of aging, as well as pathways that were not previously linked to aging. The atlas we developed points to alterations in circadian gene expression as a key regulator of aging. One Sentence SummaryThe Ticking Clock: Systemic changes in circadian gene expression correlates with wide-ranging phenotypes of aging

developmental biology↗

bioRxiv2020

Failures of nerve regeneration caused by aging or chronic denervation are rescued by restoring Schwann cell c-Jun

After nerve injury, myelin and Remak Schwann cells reprogram to repair cells specialized for regeneration. Normally providing strong regenerative support, these cells fail in aging animals, and during the chronic denervation that results from the slow growth of axons. This impairs axonal regeneration and causes a significant clinical problem. In mice, we find that repair cells express reduced c-Jun protein as the regenerative support provided by these cells declines in aging animals and during chronic denervation. In both cases, genetically restoring Schwann cell c-Jun levels restores regeneration to that in controls. We identify potential gene candidates mediating this effect and implicate Shh in the control of Schwann cell c-Jun levels. This establishes that a common mechanism, reduced c-Jun in Schwann cells, regulates the success and failure of nerve repair both during aging and chronic denervation. This provides a molecular framework for addressing important clinical problems, and suggests molecular pathways that can be targeted to promote repair in the PNS.

neuroscience↗

bioRxiv2019

Molecular and Morphological Signatures of Chordate Development: Two Distinct Pathways, One Tunicate

All chordates, including urochordates such as tunicates, develop through embryogenesis. The chordate larvae of colonial tunicates metamorphose to lose all chordate structures such as notochord, neural tube, segmented musculature, and then develop by asexual reproduction [blastogenesis], whereby stem cells form tissues and organs. These two developmental pathways establish the same body axis, morphogenetic patterning and organ formation. It is unknown if this convergent morphology implies convergent cellular and molecular mechanisms, and whether the stem cells that mediate these processes differ. Using the colonial tunicate Botryllus schlosseri, we combined transcriptome sequencing and multiple microscopy techniques to study the molecular and morphological signatures of cells at each developmental stage of embryogenesis and blastogenesis. This revealed that the molecular programs are distinct, but the blastogenic tissue-specific stem cells and embryonic precursor populations share similar molecular profiles. By comparing embryogenesis in other chordates we found shared developmental principles, highlighting transcription factors as key evolutionary conserved elements. This study establishes a platform for advancing the science of stem cell biology and regulation of development and regeneration.

developmental biology↗