Biology subjects

Nguyen, N.

Publications and source records attributed to Nguyen, N..

At least 19 recordsLinked to original sources

bioRxiv2026

LEUTX mediates amplification of DUX4 signaling and aberrant myotube differentiation in facioscapulohumeral muscular dystrophy

Aberrant reactivation of an embryonic DUX4 transcription factor gene encoded in the D4Z4 macrosatellite repeats on chromosome 4q is considered the root cause of facioscapulohumeral muscular dystrophy (FSHD). While DUX4 target gene expression correlates with the clinical severity, DUX4 expression itself is very rare. How this limited DUX4 expression leads to definitive target gene activation and how these downstream genes contribute to FSHD pathology remain elusive. We previously found the evidence for LEUTX and ZSCAN4 representing two distinct DUX4 target subpathways. Here we present functional analyses in control and FSHD skeletal myocytes identifying LEUTX, but not ZSCAN4, as a major amplifier of DUX4 target gene expression. Furthermore, LEUTX uniquely disrupts myotube differentiation by impairing muscle gene induction and promoting persistent mitotic gene expression. Forced LEUTX expression causes aberrant DNA replication in myotube nuclei. A subset of misregulated genes is bound by LEUTX, strongly suggesting that they are direct targets of LEUTX. Notably, LEUTX requires the presence of DUX4 to gain access to DUX4 target gene promoters for their activation. Taken together, our results establish the critical role of LEUTX in DUX4 signal amplification and defective myotube differentiation, revealing distinct contributions of DUX4 target gene pathways in FSHD pathogenesis.

cell biology↗

bioRxiv2026

A binuclear copper enzyme platform for enantioconvergent radical (pseudo)halogenation

Despite their intriguing native metalloenzymology, naturally occurring copper enzymes remain largely underexploited for new-to-nature biocatalytic reactions. Herein, we report the systematic investigation and reprogramming of natural copper enzymes to catalyze unnatural free radical (pseudo)halogenation reaction in a highly enantioselective fashion. Evaluating Cu enzymes in the decarboxylative azidation of redox-active esters revealed activity across multiple Cu enzyme families, with type III binuclear Cu enzymes, particularly the Bacillus megaterium tyrosinase (BmTyr), exhibiting superior activity and enantioselectivity across both stabilized and unstabilized alkyl radicals upon further engineering. The strong halide binding affinity of the binuclear Cu system also enabled challenging enantioconvergent bromination, chlorination and isothiocyanation reactions, which remained inaccessible to repurposed nonheme Fe enzymes. Further EPR and UV-visible spectroscopic analyses confirmed the coupled binuclear nature of wild-type and engineered bacterial tyrosinases and provided insights into the origin of their enhanced activity. Collectively, this study establishes binuclear copper enzymes as a powerful platform for new-to-nature stereoselective radical reactions, expanding the scope of metalloenzyme catalysis beyond mononuclear systems.

biochemistry↗

bioRxiv2026

Microfluidic Devices for Imaging and Biochemistry Analysis of Microbes Under Mechanical Pressure

Growth-induced pressure arises when proliferating cell populations are confined within rigid microenvironments and is increasingly recognized as a determinant of microbial physiology in soils, biofilms, and host tissues. We present two complementary microfluidic devices that confine microbes within polydimethylsiloxane (PDMS) chambers and apply defined, optically read-out growth-induced pressure of up to 1.5 MPa. The first, the self-closing (SC) chip, confines cells in multiple small chambers accommodating hundreds of cells; it provides excellent nutrient supply and rapid medium exchange, supports single-cell imaging, and allows multiplexing of cellular or chemical conditions. The second, the pressure-recovery (PR) chip, confines cells in a single 5 cm-long chamber accommodating hundreds of thousands of cells; a scalpel-cut step recovers live cells from the channel within minutes for bulk biochemical assays. The two devices share a single two-layer soft-lithography fabrication process and a common brightfield wall-displacement pressure readout. Together, they enable single-cell imaging and bulk biochemical analysis under matched, defined pressure conditions. We illustrate the protocol with two representative validations: rapid {beta}-estradiol-induced transcription in the SC chip, in which nascent transcription foci appear within 5 min independently of the applied pressure, and the PR chip, which produces uniform growth-induced pressure along the entire 5 cm channel and recovers between 0.1x10^6 and 1x10^6 cells per device in a pressure-tunable manner.

biophysics↗

bioRxiv2026

ARCHER: Amortized cross-specimen pose estimation for cryo-electron microscopy

Single-particle cryo-electron microscopy (cryo-EM) pose estimation is traditionally solved anew for each dataset, where iterative refinement is done from scratch while the estimator learns to store the molecule in its weights. In this work, we show that pose inference is a generalizable, specimen-agnostic operation when conditioned explicitly on a reference volume. We introduce ARCHER, an amortized contrastive classifier that models the pose posterior over a discrete rotation grid. Trained across a variety of protein structures, it operates zero-shot without retraining per structure. This transferability is grounded in Fourier-space information mechanics, where all specimen dependence is captured by the reference structure's power spectrum and spatial extent. ARCHER achieves a median angular error of 5.0{degrees} on 100 held-out test structures and 2.5{degrees} on experimental particles, matching dedicated estimators within 0.16[A] in 3D reconstruction. Crucially, downstream conformational signal is preserved. The leading conformational coordinate correlates at 0.97 with deposited benchmarks, faithfully reconstructing free-energy basins and mobile domains. These results overall demonstrate that cryo-EM pose estimation can be generalized across different structures.

biophysics↗

bioRxiv2026

Remodeled cephalic arch geometry promotes disturbed flow at pre-maturation flow rates in hemodialysis patients

In the United States, roughly 550,000 people receive routine hemodialysis for end-stage renal disease. This treatment requires an arteriovenous access, most commonly a brachiocephalic fistula. However, these accesses often fail due to stenosis in the cephalic arch (CA), a common complication whose underlying causes remain unclear (Bennet et al., 2015). To characterize the hemodynamic environment in the CA, we employed patient-specific millifluidic models that were perfused with blood-mimicking fluid containing fluorescently labelled beads to visualize flow behaviors. We perfused our models across physiologic (28-45 mL/min) and elevated (60-423 mL/min) flow rates and quantified wall shear stress (WSS) and streamline angle as a measure of flow disturbance. Our findings show that the bulk curvature and remodeled wall topography each create regions of persistently low WSS, consistent with prior clinical observations (Hammes et al., 2016). Moreover, remodeled wall topography promotes disturbed flow at elevated flow rates, a hemodynamic profile associated with various vascular pathologies (Chiu & Chien, 2011). Independently performed computational fluid dynamics (CFD) modeling complements these results, showing that remodeled wall topography promotes vortex formation at elevated flow rates, as assessed by Q-criterion. Collectively, our experimental and computational results provide strong evidence for geometry-driven disturbed flow in the CA at elevated flow rates. Notably, we observed disturbed flow at flow rates as low as 81 mL/min, far below the 600 mL/min required for hemodialysis. Disturbed flow thus offers a plausible mechanism that relates access flow rates to the vascular pathologies that precede access failure. Significance StatementHemodialysis requires an arteriovenous fistula (AVF) that must remodel and mature to withstand chronically elevated blood flow rates. However, how remodeled vessel geometry interplays with elevated flow to shape local hemodynamics remains poorly understood. Here, we used millifluidic models of the cephalic arch (CA) to show that vessel geometry and elevated flow rates promote regions of low wall shear stress and disturbed flow. Notably, geometry-driven disturbed flow is observed at flow rates above physiologic levels but well below 600 mL/min, the flow rate necessary for adequate dialysis. Because disturbed flow is injurious to the endothelium, our findings are the first to show that vascular damage begins before fistula maturation.

biophysics↗

bioRxiv2026

Hyaluronan and CD44 targeting reverses early matrix changes, proinflammatory signals and fibrosis in primary sclerosing cholangitis

Primary sclerosing cholangitis (PSC) is a rare, progressive liver disease characterized by biliary inflammation and bile duct strictures and no approved medical therapy. Despite its clinical severity, the pathological mechanisms underlying PSC remain poorly understood, largely due to early diagnostic challenges. Here we provide complementary evidence in human PSC samples, transcriptomic data, mouse models, and 3D cholangiocyte cultures that underscore the importance of hyaluronic acid (HA) and its cognate receptor CD44 in PSC pathogenesis. HA is a glycosaminoglycan abundant in the extracellular matrix in inflammatory disorders, yet its role in PSC has not been well characterized. We demonstrate that in early-stage PSC, cholangiocytes aberrantly produce high molecular weight HA that accumulates in the peribiliary matrix, increasing local tissue stiffness. This mechanical signal is transduced by a CD44/Integrin {beta}1 receptor complex in cholangiocytes, driving cell proliferation, YAP mechanosignaling, pro-inflammatory cytokine production with a transition to a ductular reactive phenotype. CD44 knockdown in cholangiocyte cell lines and mouse models significantly lowered stiffness, and attenuated inflammation. Together, these findings reveal a mechano-inflammatory axis in which HA-driven matrix stiffening perpetuates biliary inflammation and disease progression, identifying HA targeting and CD44 as promising therapeutic strategies. One Sentence SummaryHyaluronan and CD44 mediate matrix changes and progressive fibrosis in primary sclerosing cholangitis.

pathology↗

bioRxiv2026

Persistent microbial material contributes to Alzheimer disease and is targetable by vaccination

Chronic neuroinflammation is increasingly recognized as a contributor to Alzheimer disease, yet the upstream stimuli that sustain it remain poorly defined. We investigated whether persistent microbial material contributes to Alzheimer disease using the conserved microbial polysaccharide poly-N-acetylglucosamine (PNAG). PNAG-containing microbial material colocalized with amyloid plaques in human Alzheimer disease brain tissue. In fully human neuronal and three-dimensional brain models, purified PNAG and PNAG-containing microbial vesicles activated Toll-like receptor 2-dependent inflammasome signaling and promoted amyloid-{beta} and phosphorylated tau accumulation. Vaccination targeting PNAG improved cognition, reduced glial activation and amyloid pathology, remodeled amyloid processing, and preserved gut microbial community structure in APP/PS1 mice. These findings identify persistent microbial material as an upstream contributor to Alzheimer disease-associated neuroinflammation and a potential therapeutic target.

neuroscience↗

bioRxiv2026

Adenosine A2B Receptor Activation: A Novel Therapeutic Strategy for Accelerating Liver Recovery After Acetaminophen Overdose

An acetaminophen (APAP) overdose is the leading cause of drug-induced hepatotoxicity and acute liver failure (ALF) in the United States. While N-acetylcysteine (NAC), is highly effective when administered early after an overdose, its efficacy decreases with delayed administration. Since most patients present late to the clinic, there is an urgent need for novel late-acting therapeutic options to prevent progression to ALF. We previously demonstrated the benefit of delayed activation of the Adenosine A2B Receptor (A2BAR) in attenuating APAP-induced hepatotoxicity and this study focuses on its effects on liver recovery after injury. Fasted male C57BL/6J mice were treated with 300 mg/kg APAP, followed by activation of A2BAR 6 or 9 h later and sacrifice 24, 48 or 72 h post-APAP with evaluation of liver injury, the innate immune response and liver regeneration. Delayed activation of A2BAR significantly enhanced liver recovery, with accelerated repopulation of the liver by Kupffer cells, increased macrophage migration to the necrotic areas and their faster resolution. A2BAR activation also upregulated lipid metabolism-related genes in non-parenchymal cells and cell proliferation and metabolism genes in hepatocytes. Remarkably, genes such as Cidec and Plin2, crucial for lipid droplet formation, were upregulated, indicating that A2ABR activation enhances lipid metabolism which plays a key role in providing energy for liver regeneration. Overall, these findings highlight the potential of A2BAR activation not only in protecting against liver injury, but also in promoting and accelerating liver regeneration by modulating the innate immune responses and metabolic pathways.

pharmacology and toxicology↗

bioRxiv2026

A replication hijacking mechanism for Tn3-family replicative transposition

Transposition of all classes of transposable elements generates DNA intermediates that must be processed by the host to be effective. However, the mechanisms whereby transposons communicate with cellular DNA-processing machineries remain poorly investigated. Here, we provide convergent genetic and biochemical evidence that replicative transposition of the Tn3-family transposon Tn4430 is strictly coupled to replication of the target. Blocking target replication abolishes transposition, while blocking replication of the transposon donor molecule has no effect. Furthermore, the insertion preference of Tn4430 was found to be altered by the direction of replication and potential replication impediments within the target, suggesting a functional link between the integration mechanism and replication fork progression. In vitro, the transposase TnpA was found to specifically bind to fork-like DNA structures that mimic replication intermediates. Compared to linear DNA fragments, these structures are efficient substrates for TnpA-catalysed end joining. Strand transfer occurred immediately downstream of the fork, poising the transposon for replication. Together, the data suggest a mechanism in which the transposon targets DNA replication intermediates to directly recruit the host replication machinery at the time of transposition. This "replication hijacking" mechanism contrasts with classical "replication hiring" mechanisms during which replication is recruited after strand transfer. SIGNIFICANCE STATEMENTBacterial transposons of the Tn3 family constitute a threat for human health, being continuously involved in the emergence and spread of new antimicrobial resistances amongst pathogens. The success of these elements is based on their replicative mode of transposition allowing them to produce a new copy of themselves whenever they move. Here, we provide evidence that, rather than recruiting the host replication machinery after integration as is proposed in textbook models for replicative transposition, Tn3-family transposons directly transpose into ongoing replication intermediates. We propose that this "replication hijacking" mechanism provides a means of synchronizing transposition with cell replication activity, thus optimizing the movement of transposons with the expansion of bacterial populations. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=57 SRC="FIGDIR/small/728999v1_ufig1.gif" ALT="Figure 1"> View larger version (10K): org.highwire.dtl.DTLVardef@c04da0org.highwire.dtl.DTLVardef@1285b5aorg.highwire.dtl.DTLVardef@859c5corg.highwire.dtl.DTLVardef@1dc9ce8_HPS_FORMAT_FIGEXP M_FIG C_FIG

molecular biology↗

bioRxiv2026

Selective targeting of the oligodendroglial GPR17 receptor improves myelin integrity and motor function in female SOD1G93A mice

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease with no definitive disease-modifying therapies available, underscoring the urgent need to identify novel druggable targets. The G protein-coupled receptor GPR17 is a critical regulator of oligodendrocyte maturation and has emerged as a candidate target in ALS, yet its relevance to human disease and its therapeutic potential remain unclear. Here, we demonstrate that pathological GPR17 upregulation defines a conserved, pathologically immature oligodendroglial state in human ALS that can be pharmacologically leveraged to restore myelin integrity and improve functional outcome in vivo. Publicly available transcriptomics datasets and histological analyses revealed an increased abundance of GPR17-expressing immature oligodendrocytes in post-mortem human spinal cord tissue from ALS cases. Moreover, sustained activation of GPR17 with a selective agonist induced GPR17 internalization in heterologous expression systems and promoted the differentiation of primary oligodendrocyte precursors derived from SOD1G93A mice. Translating these findings in vivo, chronic treatment with a brain-penetrant GPR17 agonist derived from the same pharmacological class significantly extended survival, delayed body weight loss, and improved motor performance in female SOD1G93A mice, whereas male mice showed no therapeutic benefit. These effects were associated with restored oligodendrocyte maturation, preserved myelin integrity, motor neuron survival, and attenuated reactive gliosis in the spinal cord of female SOD1G93A mice, while milder effects were observed in males. Together, these findings establish oligodendroglial GPR17 as a conserved and pharmacologically actionable target in ALS and show that sustained in vivo GPR17 agonism can reprogram altered oligodendroglial states and slow disease progression in a sex-dependent manner.

pharmacology and toxicology↗

bioRxiv2026

EVEE: Interpretable variant effect prediction from genomic foundation model embeddings

Scientific foundation models learn high-dimensional representations from diverse data modalities, yet what they encode and how to extract that knowledge remain open questions. Here we show that probing the internal representations of Evo 2, a 7-billion-parameter genomic foundation model, enables accurate and interpretable genetic variant effect prediction. We introduce a covariance-based probe that captures second-order structure from Evo 2 sequence embeddings to predict variant pathogenicity across variant types and functional consequences, matching or exceeding specialized predictors within their domains. To ground these predictions in known biological mechanisms, we train a complementary panel of probes on existing annotations to detect which genomic properties are disrupted by a variant. This categorized evidence is then integrated with each variants genomic context through a language model to generate variant-specific mechanistic hypotheses. Our pathogenicity predictions correlate with experimental measures of variant function, clinical penetrance, and biobank disease associations while the mechanistic hypotheses are consistent with expert reviews, known mechanism classes, and downstream molecular readouts. We release pathogenicity scores, disruption profiles, and contextualized interpretations for 4.2 million variants from the ClinVar database as an open resource through the Evo Variant Effect Explorer (EVEE). More broadly, this structured probing approach offers a general framework for interrogating foundation models across scientific disciplines and grounding their outputs in existing domain concepts.

genomics↗

bioRxiv2026

Engineered probiotics that sequester arsensite in a mouse gastrointestinal system

Chronic exposure to inorganic arsenic remains a major global health concern, as arsenite is frequently present in contaminated food and drinking water and readily absorbed through the gastrointestinal (GI) tract. Once internalized, arsenite accumulates in tissues and contributes to long-term health effects, including cancer, organ dysfunction, and neurological disorders. Despite extensive efforts to reduce environmental contamination, there are currently no practical strategies to prevent dietary arsenite from entering the human body during digestion. Here, we report a synthetic biology-based approach that uses engineered probiotics to detect and sequester arsenite directly within the GI tract before systemic absorption occurs. We engineered Escherichia coli Nissle 1917 (EcN), a probiotic strain, to function as a living arsenite-interception system. Central to this design is an arsenite-responsive genetic toggle switch that activates chelator expression upon exposure and sustains production under biostatic conditions, while automatically shutting off during active cell division to limit metabolic burden and enhance biosafety. In parallel, we engineered an arsenite-binding protein derived from the transcriptional regulator ArsR to eliminate DNA-binding activity while retaining high-affinity metal binding, yielding a non-toxic chelator suitable for intracellular sequestration. The resulting engineered strain efficiently removed arsenite from its surrounding environment in vitro while maintaining robust cell viability and growth. To translate these findings to an in vivo context, we developed a mass-transfer model describing arsenite distribution among the stomach lumen, engineered bacteria, and epithelial cells. This model guided the selection of a bacterial dose predicted to substantially deplete lumenal arsenite prior to epithelial uptake. Using this strategy, we demonstrated in a mouse GI model that oral administration of engineered EcN markedly reduced arsenite entry into the bloodstream compared with wild-type EcN or no-bacteria controls. Together, these results establish a programmable probiotic platform for intercepting dietary arsenite and highlight a potential strategy for preventing absorption of environmental toxicants using living microbial therapeutics.

synthetic biology↗

bioRxiv2025

Decreased endothelial cell retinoic acid signaling accelerates progression of single ventricle pulmonary arteriovenous malformations

BackgroundPulmonary arteriovenous malformations (PAVMs) are vascular complications that universally develop in patients with single ventricle congenital heart disease after Glenn surgery. However, the pathophysiological mechanisms underlying single ventricle PAVMs are poorly understood. To comprehensively evaluate molecular changes post-Glenn, we performed single-cell RNA sequencing (scRNAseq) on rat lung samples after Glenn surgery. MethodsUsing adult Sprague Dawley rats, we performed scRNAseq on unfiltered lung samples 3 weeks after left-sided Glenn or sham surgery. We compared endothelial cell (EC) differentially expressed genes (DEGs) in our model to two mouse models of hereditary hemorrhagic telangiectasia (HHT), a hereditary condition characterized by visceral AVMs. Finally, we modified the vitamin A (Vit A) content of Glenn and sham rat diets and re-assessed PAVM shunting and EC transcriptional differences. ResultsUsing scRNAseq (n=4 Glenn, n=4 sham), we identified 13 transcriptionally distinct lung cell clusters, including 3 EC clusters (general, capillary, lymphatic), with pronounced differences between Glenn and sham in the general EC cluster ([~]17% of genes). Comparison to HHT mouse models confirmed overlap of [~]18% of DEGs, including identification of significantly downregulated genes involved in and regulated by all-trans retinoic acid (ATRA) signaling in all 3 models. Dietary deficiency of Vit A intake, a precursor of ATRA, caused increased PAVM shunting (p<0.01) that was mitigated with excess dietary Vit A intake. Lastly, EC-specific RNAseq identified Vit A diet-induced gene expression differences, including regulation of PI3K signaling. ConclusionsIn this study, we report the novel application of scRNAseq to study mechanisms underlying single ventricle PAVMs in a surgical rat model. We identified multiple dysregulated biological processes in rat lung ECs post-Glenn, including decreased ATRA signaling and conserved gene expression patterns with HHT. Dietary modification of Vit A intake altered post-Glenn shunting and represents a novel potential therapeutic strategy for single ventricle PAVMs and HHT AVMs.

physiology↗

bioRxiv2025

Expanding the scope of Mycobacterium abscessus reference strains to improve pulmonary disease modeling

Mycobacterium abscessus (Mab) pulmonary disease is an emerging clinical challenge, particularly among individuals with immunosuppression or underlying structural lung conditions. There are currently no FDA-approved therapies for Mab disease. Existing treatment strategies using repurposed drugs are prolonged, complex, and yield low cure rates (30-50%), underscoring the urgent need for more effective therapeutics. Developing new treatments requires preclinical disease models that faithfully replicate human disease, and the choice of Mab strain is a key determinant of model relevance. The commonly used reference strain, ATCC 19977, was isolated from a non-pulmonary source but became the default due to its early availability. To evaluate its relevance for pulmonary disease modeling, we compared ATCC 19977 with 15 clinical Mab isolates derived from lung infections across diverse regions of the United States. In both in vitro assays and a validated mouse lung infection model, ATCC 19977 behavior differed from the clinical isolates for key traits including rapid systemic dissemination, failure to develop robust lung granulomas, and early mortality. In contrast, clinical isolates demonstrated greater pulmonary tropism and reduced dissemination, with several producing robust lung pathology. Based on these findings, we propose a set of pulmonary clinical isolates representing the major Mab subspecies for use in lung infection research. These isolates more accurately recapitulate the pathological features of human Mab lung disease and are expected to enhance the translational value of future mechanistic and therapeutic studies. SUMMARY STATEMENTUsing a mouse model, this study identified Mycobacterium abscessus clinical isolates whose infection profiles more closely resemble human pulmonary disease, establishing them as superior reference strains to ATCC 19977 for future translational and therapeutic research.

microbiology↗

bioRxiv2025

Harnessing cGAS-STING signaling to counteract the genotoxic-immune nexus in tauopathy

Tauopathies are progressive neurodegenerative disorders characterized by aberrant tau aggregation, cognitive decline, and persistent neuroinflammation, yet the mechanisms driving neuroinflammation and disease progression remain incompletely understood. Here, utilizing human postmortem AD brains and a mouse model of tauopathy, we report that genotoxic stress-induced cyclic GMP-AMP synthase (cGAS)-stimulator of interferon genes (STING) immune pathway form a self-amplifying loop that fuels neuropathology and cognitive deficits. Targeted disruption of this cycle through either genetic deletion of cGAS or pharmacological inhibition of STING restores immune homeostasis and attenuates tau pathology and cognitive deficits. Our results showed a significant accumulation of DNA double-strand breaks (DDSBs) and impaired DNA repair function, alongside elevated cGAS-STING signaling and type I interferon (IFN-I) responses in human AD brains compared to non-AD. In the PS19 transgenic (PS19Tg) mouse model of tauopathy, we found significantly elevated levels of DDSBs and altered expression of DNA repair proteins during early stages of disease, which preceded the dysregulation of cGAS-STING signaling and emergence of significant neuropathology in the later stage. Interestingly, genetic deletion of cGAS shifted microglial polarization from a pro-inflammatory M1 phenotype toward an anti-inflammatory M2 state, accompanied by a reduction in IFN-I signaling and improved cognitive performance in PS19Tg mice. Pharmacological STING inhibition reshaped the transcriptomic landscape, revealing selective regulation of pathways governing synaptic plasticity, and immune responses. This transcriptional reprogramming was accompanied by suppression of inflammatory responses, reduction in synaptic pathology, and attenuation of tau pathology in PS19Tg mice, underscoring STING as a therapeutic target for tauopathy. In conclusion, our findings reveal that genotoxic-immune crosstalk drives neuroinflammation and tau pathology and identify a conserved, druggable cGAS-STING axis that can be targeted to impede or slow disease progression in tauopathies. O_FIG O_LINKSMALLFIG WIDTH=184 HEIGHT=200 SRC="FIGDIR/small/678980v1_ufig1.gif" ALT="Figure 1"> View larger version (53K): org.highwire.dtl.DTLVardef@1339c89org.highwire.dtl.DTLVardef@16a1d5borg.highwire.dtl.DTLVardef@1ad4631org.highwire.dtl.DTLVardef@db4615_HPS_FORMAT_FIGEXP M_FIG C_FIG

neuroscience↗

bioRxiv2025

Practical considerations for accurate estimation of diffusion parameters from single-particle tracking in living cells

Advances in fluorescence microscopy have enabled high-resolution tracking of individual biomolecules in living cells. However, accurate estimation of diffusion parameters from single-particle trajectories remains challenging due to static and dynamic localization errors inherent in these measurements. While previous studies have characterized how such errors affect mean-squared displacement (MSD) analysis, practical guidelines for minimizing them during data acquisition and correcting them during analysis are still lacking. Here, we combine theoretical modeling and simulations to evaluate how exposure time and sampling rate influence the accuracy of MSD-based inference under fractional Brownian motion (FBM), a canonical model of anomalous diffusion. We demonstrate that decoupling exposure and sampling times enables escape from the error-prone regime, thus improving inference accuracy, and that incorporating an offset in nonlinear MSD fitting substantially improves the estimation of the anomalous diffusion exponent. We validate this framework using trajectories of cytoplasmic particles in Escherichia coli, recovering consistent diffusion parameters across multiple data sets. Our findings offer practical strategies to improve both experimental design and data analysis in single-particle tracking of live or synthetic systems.

biophysics↗

bioRxiv2025

Precise MRI-Histology Coregistration of Paraffin-Embedded Tissue with Blockface Imaging

Magnetic resonance imaging (MRI) provides 3D spatial information on tissue, yet it lacks at the molecular level. In contrast, histology provides cellular and molecular information, but it lacks the 3D spatial context and direct in vivo translation. Coregistering the two is key for the 3D-embedding of histological details, validating pathological MRI findings, and finding quantitative imaging biomarkers of neurodegenerative diseases. However, coregistration is challenging due to non-linear distortions of the tissue from histological processing and sectioning leading to microscopic and macroscopic nonlinear 3D deformations between specimen MRI and stained histology sections. To address this, we developed a novel pipeline, named Brewsters Blockface Quantification (BBQ), integrating robust optical approaches with innovative 2D and 3D registration algorithms to achieve precise volumetric alignment of specimen MRI data with histological images. On a variety of brain tissue specimens from distinct anatomical regions and across multiple species, our methodology generated blockface volumes with minimal distortion and artifacts. Using these blockface volumes as an intermediary, we achieve a precise alignment between MRI and histology slides, yielding registration results with an overlapping Dice score of [~]90% for whole tissue alignment between MRI and blockface volumes, and >95% for 2D MRI-histology registration. This correlative MRI-histology pipeline with robust 2D and 3D coregistration methods promises to enhance our understanding of neurodegenerative diseases and aid the development of MRI-based disease biomarkers.

bioengineering↗

bioRxiv2025

Coronary Artery Disease Risk Variant rs6903956 Links to Endothelial Dysfunction via PHACTR1 Regulation

Ischemic heart disease, particularly coronary artery disease (CAD), remain leading causes of mortality worldwide. The single nucleotide polymorphism rs6903956 on chromosome 6p24.1 has been identified as a susceptibility locus for CAD in East Asian populations through genome-wide association studies. However, its functional role has not been fully elucidated. This study investigates the mechanistic basis of rs6903956 and its contribution to CAD pathogenesis, focusing on endothelial cell dysfunction. We first conducted cohort studies, revealing an association between the rs6903956 A risk allele and blood pressure phenotypes, along with impaired endothelial responsiveness indicated by reduced flow-mediated dilation. Single-base editing of induced pluripotent stem cell-derived endothelial cells obtained from patients with CAD and expression quantitative trait loci analysis highlighted a cisacting impact of the A allele on PHACTR1 and EDN1 expression, suggesting allelespecific regulatory effects. Using in silico modeling by AlphaFold 3 platform, the A allele exhibited enhanced binding affinity for HOXA4 and MEIS1 transcription factors, forming a stable ternary complex that promoted transcriptional activation of PHACTR1. Functional assays demonstrated the enhancer role of rs6903956 A in PHACTR1 promoter activity, supporting its locus-specific regulatory function in endothelial cells. Under pathological flow conditions, endothelial cells harboring the A allele display elevated ICAM-1 expression and increased monocyte adhesion compared to the G allele, indicating allele-specific endothelial inflammatory activation. These findings propose a model in which rs6903956 influences PHACTR1 expression via HOX-MEIS cooperative binding, thereby modulating endothelial function and contributing to CAD susceptibility. This study provides mechanistic insights into the role of rs6903956 in endothelial dysfunction and CAD, informing potential therapeutic targets arising from genetic determinants in cardiovascular pathogenesis.

molecular biology↗