Biology subjects

Bruce, A.

Publications and source records attributed to Bruce, A..

8 recordsLinked to original sources

bioRxiv2026

Therapeutic signature mapping of paired direct and indirect LPS injury in an ex vivo human lung perfusion platform reveals injury-specific druggable programs

Acute Respiratory Distress Syndrome (ARDS) remains highly morbid and lacks approved disease-modifying pharmacotherapies. Direct (pulmonary) and indirect (extrapulmonary) insults may initiate biologically distinct early injury programs, but human tissue-level evidence from the first hours is scarce. Here we establish a paired, acellular ex vivo lung perfusion (EVLP) platform using human donor lungs unsuitable for transplantation to model direct (endobronchial) and indirect (perfusate) lipopolysaccharide (LPS) injury within the same donor. We profiled lung tissue proteomes at 4 h post-insult and performed therapeutic nomination by querying proteomics-derived injury signatures against the CLUE L1000 perturbational compendium with independent cross-platform validation. Both models developed histological injury and robust cytokine release. Direct injury preferentially enriched neutrophil degranulation, extracellular matrix remodelling and metabolic reprogramming modules, whereas indirect injury showed prominent complement/coagulation perturbation with greater endothelial activation markers in perfusate. Cross-platform prioritisation converged on tractable signalling and epigenetic axes, including JAK/STAT, PI3K/AKT/mTOR, SYK, CDK and HDAC inhibitor classes - yielding a tiered shortlist for EVLP intervention testing. This intact human lung perturbation platform enables injury-stratified mechanistic inference and therapeutic prioritisation in early lung injury relevant to ARDS.

immunology↗

bioRxiv2026

Cytoglobin regulates ventricular morphogenesis and diastolic function through NO-sGC-cGMP signaling during development.

AimsHypoplastic left heart syndrome (HLHS) is a severe congenital heart disease characterized by ventricular hypoplasia and impaired cardiac function. Clinically, inhaled nitric oxide (NO) therapy is used to reduce pulmonary vascular resistance and improve cardiopulmonary stability in HLHS patients. However, whether NO signaling contributes to HLHS pathogenesis remains unknown. Cytoglobin (Cygb) is a heme-protein traditionally thought to limit NO bioavailability and downstream activation of the soluble guanylate cyclase (sGC) cyclic guanosine monophosphate (cGMP) signaling pathway. Unexpectedly, our recent work shows that Cygb enhances NO signaling through activation of NO synthase, leading to downstream activation of sGC-cGMP signaling. In zebrafish embryos, Cygb-dependent NO signaling is required for normal cilia motility and the establishment of correct cardiac laterality. Here, our aim was to determine whether Cygb-dependent NO-sGC signaling linked to cilia function regulates cardiac morphogenesis and contributes to ventricular hypoplasia in HLHS. Methods and ResultsWe found that loss of Cygb (cygb2) in zebrafish disrupts NO-sGC signaling during cardiogenesis, altering cardiac progenitor organization and migration within the anterior lateral plate mesoderm. Disruption of these processes impairs heart tube morphogenesis, thereby producing a compact ventricle wall characterized by increased wall thickness (despite preserved cardiomyocyte number) reduced ventricle size and decreased stroke volume, recapitulating key features of HLHS. Genetic disruption of the sGC -subunit (gucy1a1) and pharmacological NO scavenging phenocopy the cygb2 mutant phenotype, resulting in reduced cGMP levels, compact ventricular architecture and decreased stroke volume. Consistently, restoration of NO-sGC signaling in cygb2 mutants rescues early cardiac progenitor patterning, ventricular morphology and stroke volume. ConclusionsThese findings identify Cygb-dependent NO-sGC signaling as a critical developmental pathway for ventricular development and performance, temporally linking cardiac progenitor dynamics to cilia-dependent signaling associated with left-right patterning. This study further suggests that pharmacological activation of sGC may provide a therapeutic strategy for hypoplastic ventricular disease. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=186 HEIGHT=200 SRC="FIGDIR/small/711730v4_ufig1.gif" ALT="Figure 1"> View larger version (39K): org.highwire.dtl.DTLVardef@13a1515org.highwire.dtl.DTLVardef@3e1a6eorg.highwire.dtl.DTLVardef@1394ec2org.highwire.dtl.DTLVardef@161c81c_HPS_FORMAT_FIGEXP M_FIG C_FIG

developmental biology↗

bioRxiv2025

Measurement of nitric oxide production in mouse tracheal epithelial cell cultures differentiated at the air liquid interface.

Airway ciliary dysfunction often leads to impaired mucociliary clearance, a defect frequently associated with dysregulation of canonical nitric oxide (NO) signaling. Indeed, many airway diseases exhibit aberrant levels of exhaled NO, a feature commonly used as a diagnostic indicator in patients. While this measurement is routinely performed clinically, there is currently no validated approach to assess NO release from airway ciliated cells in vitro, limiting mechanistic studies of NO regulation and function. An established method to study airway diseases in vitro is the use of cultured mouse tracheal epithelial cells (MTECs) differentiated at the air-liquid interface (ALI). Here, using this model, we develop a protocol to measure NO released from ciliated cells and accumulated in apical mucus, collected from the ALI-differentiated cultures. Using established instrumentation for NO analysis and a triiodide-based assay, we quantify nitrite, a stable NO metabolite and biomarker of NO production. For assay validation, cells were treated with the NO donor Diethylenetriamine NONOate (DETA-NONOate or DETA/NO) or the NO synthase inhibitor N({omega})-Nitro-L-arginine methyl ester (L-NAME). Together, these approaches demonstrate that this method reliably detects NO in samples from ALI-differentiated airway epithelia, providing an accurate in vitro platform to quantify NO release and model diseases associated with abnormal NO metabolism. SUMMARYThe goal of this current protocol is to describe a cell culture-based method to study airway diseases associated with altered nitric oxide (NO) signaling. This method describes the quantification of NO in airway epithelia differentiated at the air-liquid interface (ALI) using a triiodide-based chemiluminescence assay.

molecular biology↗

bioRxiv2025

Chamber-Specific Transcriptomic Insight into Cardiac Development using Guinea Pig and Human Heart Tissue

The heart undergoes significant molecular and functional adaptations throughout postnatal development. However, to date, our understanding of these dynamic changes in the human heart is limited. Moreover, advances in pediatric cardiac research can be hindered by a lack of preclinical models that accurately reflect human heart maturation. Guinea pigs may serve as a useful model for human cardiac research, as the guinea pig and human myocardium have similar ion channel expression and cardiovascular drug responsiveness. Despite these similarities, gene expression patterns during postnatal heart development have not been comprehensively investigated. In this study, we first characterized transcriptional changes in neonatal, juvenile, and adult guinea pig hearts - identifying gene ontologies and pathways associated with cardiac maturation. Second, we compared the transcriptional profile of right atria and left ventricular tissue to highlight unique and shared chamber-specific patterns in guinea pigs over time. Finally, we conducted a cross-species comparison of the right atrial transcriptome between humans and guinea pigs to identify conserved maturation markers and gene expression patterns. Our findings provide a molecular framework for understanding age- and chamber-specific cardiac development, supporting the guinea pig as a promising preclinical model for studying human heart maturation. By identifying conserved gene programs and developmental markers across species, this study lays the groundwork for age-specific pharmacological strategies and computational models that can help to refine treatment decisions and outcomes for pediatric cardiology patients. New and NoteworthyExisting knowledge on postnatal heart development and cardiomyocyte maturation is limited. We investigated age-dependent transcriptional changes in neonatal, juvenile, and adult guinea pig hearts - and then conducted a cross-species comparison to identify age-specific patterns that are conserved in the guinea pig and human atria. Expanding our knowledge of chamber- and age-specific gene expression patterns can inform and guide the selection of cardiovascular therapies in the pediatric population, where developmental differences are understudied.

genomics↗

bioRxiv2025

Gut bacteria gatekeep host-specific colonization of a commensal fungus that boosts type 2 immunity

Naturalized, wilded, wildling, and dirty/pet store mouse models represent a spectrum of approaches designed to make laboratory mice more immunologically and physiologically similar to wild or human contexts by increasing their exposure to naturally occurring microbes and pathogens. In this study, we screened the gut mycobiome of pet store mice, and identified Kazachstania pintolopesii as a dominant fungus in pet store mice across various geographical locations. K. pintolopesii strains isolated from mice in geographically distinct pet stores stably colonize the gastrointestinal tract of laboratory mice, independent of gut bacterial composition, maintaining high fungal burdens for extended periods. K. pintolopesii rapidly became the dominant fungus in the mouse gut in conventional, antibiotic, and germ-free settings, outcompeting other non-murine fungal strains. Pet store-derived K. pintolopesii exhibited unique immunological properties distinct from typical anti-fungal responses. Unlike C. albicans colonization, K. pintolopesii did not induce circulating neutrophil expansion or Th17 cell populations in the gut mucosa. When administered systemically, K. pintolopesii-infected mice showed 100% survival with minimal fungal burden in kidneys, contrasting sharply with lethal C. albicans infections. Adaptive immune deficiency (Rag1 knockout mice) did not affect K. pintolopesii colonization or host response, indicating that B and T cell-mediated immunity does not restrain this fungus. K. pintolopesii colonization provided no cross-protection against systemic candidiasis further establishing lack of immune activation. These findings demonstrate that K. pintolopesii establishes a benign host-fungal relationship through neutrophil-independent mechanisms, avoiding classical anti-fungal immune activation while maintaining stable gut colonization. Instead, it selectively induces strong type 2 mucosal immune responses, increasing tuft and goblet cell counts and stimulating Th2 and group 2 innate lymphoid cell (ILC2) populations. This immune profile confers notable protection against intestinal nematode infection, demonstrated by reduced Heligmosomoides polygyrus egg counts. Altogether, K. pintolopesii serves as an exemplary model for commensal mycobiota, revealing distinct mechanisms for host tolerance and immune modulation.

microbiology↗

bioRxiv2024

Connecting Transcriptomics with Computational Modeling to Reveal Developmental Adaptations in the Human Pediatric Myocardium

BackgroundNearly 1% or 1.3 million babies are born with congenital heart disease (CHD) globally each year - many of whom will require palliative or corrective heart surgery within the first few years of life. A detailed understanding of cardiac maturation can help to expand our knowledge on cardiac diseases that develop during gestation, identify age-appropriate cardiovascular drug therapies, and inform clinical care decisions related to surgical repair, myocardial preservation, or postoperative management. Yet, to date, our knowledge of the temporal changes that cardiomyocytes undergo during postnatal development is largely limited to animal models. MethodsRight atrial tissue samples were collected from n=117 neonatal, infant, and pediatric patients undergoing correct surgery due to (acyanotic) CHD. Patients were stratified into five age groups: neonate (0-30 days), infant (31-364 days), toddler to preschool (1-5 years), school age (6-11 years), and adolescent to young adults (12-32 years). We measured age-dependent adaptations in cardiac gene expression, and used computational modeling to simulate action potential and calcium transients. ResultsEnrichment of differentially expressed genes (DEG) was explored, revealing age-dependent changes in several key biological processes (cell cycle, cell division, mitosis), cardiac ion channels, and calcium handling genes. Gene-associated changes in ionic currents exhibited both linear trends and sudden shifts across developmental stages, with changes in calcium handling (INCX) and repolarization (IK1) most strongly associated with an age-dependent decrease in the action potential plateau potential and increase in triangulation, respectively. We also note a shift in repolarization reserve, with lower IKr expression in younger patients, a finding likely tied to the increased amplitude of IKs triggered by elevated sympathetic activation in pediatric patients. ConclusionThis study provides valuable insights into age-dependent changes in human cardiac gene expression and electrophysiology among patients with CHD, shedding light on molecular mechanisms underlying cardiac development and function across different developmental stages.

cell biology↗

bioRxiv2022

Genetic and pharmacological reduction of CDK14 mitigates synucleinopathy

Parkinsonas disease (PD) is a debilitating neurodegenerative disease characterized by the loss of midbrain dopaminergic neurons (DaNs) and the abnormal accumulation of -Synuclein (-Syn) protein. Currently, no treatment can slow nor halt the progression of PD. Multiplications and mutations of the -Syn gene (SNCA) cause PD-associated syndromes and animal models that overexpress -Syn replicate several features of PD. Decreasing total -Syn levels, therefore, is an attractive approach to slow down neurodegeneration in patients with synucleinopathy. We previously performed a genetic screen for modifiers of -Syn levels and identified CDK14, a kinase of largely unknown function as a regulator of -Syn. To test the potential therapeutic effects of CDK14 reduction in PD, we ablated Cdk14 in the -Syn preformed fibrils (PFF)-induced PD mouse model. We found that loss of Cdk14 mitigates the grip strength deficit of PFF-treated mice and ameliorates PFF-induced cortical -Syn pathology, indicated by reduced numbers of pS129 -Syn-containing cells. In primary neurons, we found that Cdk14 depletion protects against the propagation of toxic -Syn species. We further validated these findings on pS129 -Syn levels in PD patient neurons. Finally, we leveraged the recent discovery of a covalent inhibitor of CDK14 to determine whether this target is pharmacologically tractable in vitro and in vivo. We found that CDK14 inhibition decreases total and pathologically aggregated -Syn in human neurons, in PFF- challenged rat neurons and in the brains of -Syn-humanized mice. In summary, we suggest that CDK14 represents a novel therapeutic target for PD-associated synucleinopathy.

neuroscience↗

bioRxiv2017

Zebrafish yolk syncytial nuclei migrate along a dynamic microtubule network

In teleosts, the yolk syncytial layer is a multinucleate syncytium that functions as an extraembryonic signaling center to pattern the mesendoderm, coordinate morphogenesis and supply nutrients to the embryo. The zebrafish is an excellent system for studying this morphogenetically active tissue. The external yolk syncytial nuclei (e-YSN) undergo microtubule dependent epiboly movements that distribute the nuclei over the yolk. How e-YSN epiboly proceeds, and what role the yolk microtubule network plays is not understood but currently it is proposed that e-YSN are pulled vegetally as the microtubule network shortens from the vegetal pole. Data from our live imaging studies suggest that the yolk microtubule network is dismantled from the animal and vegetal regions and show that a region of stabilized microtubules forms before nuclear migration begins. e-YSN do not appear to be pulled vegetally but rather move along a dynamic microtubule network. We also show that overexpression of the KASH domain of Syne2a impairs e-YSN movement, implicating the LINC complex in e-YSN migration. This work provides new insights into the role of microtubules in morphogenesis of an extraembryonic tissue.\n\nSummary StatementAnalysis of yolk syncytial nuclear migration during zebrafish epiboly reveals that nuclei migrate along and largely beneath a dynamically yolk microtubule network.

developmental biology↗