Biology subjects

Tong, M.

Publications and source records attributed to Tong, M..

At least 19 recordsLinked to original sources

bioRxiv2026

Marker-specific pixel attribution for single-cell protein quantification with PixAQ

Imaging mass cytometry (IMC) typically quantifies all markers within the same cell mask, although their signals have distinct spatial distributions. This quantification method may omit relevant signals or include signals from neighboring cells. We present PixAQ, which improves single-cell protein quantification in IMC through marker-specific probabilistic pixel attribution. PixAQ combines spatial proximity with anchor-group intensity models to assign pixel-marker observations among neighboring cells and an explicit residual component. Original intensities are averaged over marker-specific supports. Across three cohorts (680 regions), comparisons in 366 patients showed improved agreement with predefined population-marker relationships, with cohort-median patient-paired gains of 1.99 to 4.83 percentage points in expected-marker share over shared-mask quantification. Ablation analyses showed that both spatial and intensity evidence contributed to these gains. Conventional and high-resolution IMC images linked supports to visible local and intracellular signal structures. PixAQ improves single-cell protein quantification by determining which local signals contribute to each marker measurement without changing cell segmentation.

bioinformatics↗

bioRxiv2026

Continuous amyloid staging complements global amyloid burden in early Alzheimer's disease

Introduction: Early Alzheimer's disease (AD) staging remains challenging because clinical categories and global amyloid PET measures provide only coarse characterization of disease progression. We developed a continuous amyloid staging score from regional amyloid PET to complement global amyloid burden. Methods: Using longitudinal florbetapir PET from the Alzheimer's Disease Neuroimaging Initiative (ADNI), we trained an end-to-end deep learning model to directly estimate a continuous amyloid staging score. We evaluated longitudinal consistency, its relationship with global amyloid burden, regional contributors to early-stage variation, and whether the staging score stratified subsequent memory decline among participants matched for baseline global amyloid burden. Cognitive analyses were replicated in A4+LEARN. Results: The staging score better preserved longitudinal visit order than existing amyloid staging approaches. During early disease stages, it captured regional amyloid variation despite relatively stable global amyloid burden. Among participants with similar baseline global amyloid burden, higher staging scores were associated with faster subsequent memory decline in both ADNI and A4+LEARN. Discussion: A continuous amyloid staging score complements global amyloid burden for early AD staging and cognitive risk stratification. Keywords: Alzheimer's disease; amyloid PET; disease staging; cognitive decline

neuroscience↗

bioRxiv2026

Multi-omic profiling reveals impaired luteal resolution and CHK2-regulated inflammation in ovarian aging

The ovary ages earlier than most other organs, but the molecular mechanisms that drive this decline remain unclear. To gain more insight into this process, we profiled ovaries from young and reproductively aged mice after synchronized ovulation and found that aging was marked by the accumulation of luteal cells with increased steroidogenic, antigen presentation and inflammatory gene expression. DNA methylation was globally preserved in aged ovaries but showed focal age-associated changes at gene regulatory regions. Protein profiling identified an increase in checkpoint kinase 2 (CHK2), a DNA damage response protein encoded by the CHEK2 gene associated with the timing of natural menopause in humans. CHK2 and DNA damage markers were increased in aged mice and human ovarian somatic cells. Transient CHK2 inhibition in aged mice increased the proportion of mature oocytes and blastocyst formation and reversed inflammatory signatures in granulosa cells. These findings identify defective luteal resolution and inflammation as features of ovarian aging and suggest that CHK2 induces a reversible age-associated inflammatory signature. Our results show that reproductive decline may remain pharmacologically modifiable during ovarian aging via regulation of the inflammatory response.

molecular biology↗

bioRxiv2026

Divergent amyloid trajectories distinguish normal aging from Alzheimer's disease progression

Aging is the greatest risk factor for Alzheimer's disease (AD), yet how and when AD-related pathological progression diverges from aging remains poorly understood. This distinction is particularly difficult at early stages, when clinically and biomarker-defined populations contain individuals following fundamentally different trajectories. Here, we model AD progression as a deviation from aging using longitudinal amyloid PET and a self-supervised trajectory-learning framework. In ADNI, the learned trajectory revealed a shared early path that bifurcated into an aging branch and an AD-related branch. The two branches showed distinct profiles in amyloid burden, cognitive decline, risk of progression to AD dementia, and genetic risk. Unseen participants from the independent NACC cohort were projected onto the trajectory without retraining, further reproducing key branch-specific biological and genetic patterns. Together, the bifurcating trajectory provides a biologically grounded framework for resolving early-stage heterogeneity and enabling risk stratification beyond binary amyloid status.

neuroscience↗

bioRxiv2026

Cessation of diel vertical migration by an inshore dinoflagellate bloom under prey deprivation

Many dinoflagellates are mixotrophic and regulate their vertical position to navigate dynamic gradients in light, nutrients, and prey. Here, it is shown that the obligate kleptoplastidic mixotroph, Dinophysis acuminata, transitions from diel vertical migration to formation of a stationary, sub-surface thin layer in response to prolonged prey deprivation. An inshore bloom within a salt marsh kettle pond was recorded through continuous in-situ imaging, automated oxygen and fluorescence depth profiling, and targeted water chemistry measurements. During the blooms initial development, D. acuminata cells were photosynthetically active and divided vegetatively while vertically migrating. As photosynthesis and growth slowed, vertical migration ceased and cells formed a stable thin layer that promoted conditions for local acidification and nitrogen remineralization. Surface avoidance by the thin layer drove selective retention of cells within the relatively deep kettle hole. Together, these findings illustrate linkage of metabolic state and swimming behavior in D. acuminata and show how swimming behavior can drive development of toxic blooms within inshore systems. They also illustrate how D. acuminata and other eurytolerant bloom-forming species can exploit and shape physicochemical gradients associated with coastal eutrophication.

ecology↗

bioRxiv2026

Butyrate preserves cardiac health during aging

Aging promotes left ventricular hypertrophic remodeling and diastolic dysfunction, yet interventions that preserve cardiac function when initiated late in life remain limited. Butyrate, a gut microbiome derived short chain fatty acid, can act as both a metabolic substrate and an epigenetic modulator, but whether long term supplementation improves cardiac resilience in advanced age and through which multicellular programs is unclear. Here we treated male C57BL/6J mice with sodium butyrate beginning at 18 months of age and maintained treatment to 28 months. Compared with age matched controls, NaBu treated aged mice exhibited reduced left ventricular mass and improved cardiac diastolic function. Left ventricular bulk proteomics revealed a coordinated aging signature characterized by increased extracellular remodeling and innate immune/complement/hemostasis pathways and decreased metabolic and proteostasis networks; NaBu partially opposed age associated proteomic shifts and identified a subset of proteins that reversed direction between aging and NaBu axes. Single nucleus multiome profiling further resolved cellular and regulatory programs associated with aging and NaBu response. Aging induced extensive chromatin remodeling with the strongest coupled RNA and ATAC changes in cardiomyocytes. In aged hearts, NaBu was associated with a consistent reduction of endothelial IFN and MHCI antigen presentation and adhesion programs across endothelial subtypes, with concordant decreases in both RNA and chromatin accessibility/gene activity at key loci, consistent with reduced endothelial inflammatory priming. Together, physiological, proteomic, and multiome data support long term NaBu as a late life intervention that attenuates age-associated cardiac remodeling and reshapes multicellular molecular programs in the aged heart.

molecular biology↗

bioRxiv2026

Microporous Immune-Isolating Capsule with Improved Diffusion for Restored Dynamic Bidirectional Hormone Signaling in a Murine Model of Premature Ovarian Insufficiency

Pediatric cancer survivors treated with gonadotoxic chemotherapy or radiation face lifelong premature ovarian insufficiency (POI), leading to elevated risk of cardiovascular disease, osteoporosis, and metabolic dysfunction. Pharmacological hormone replacement therapy (HRT) cannot replicate the pulsatile, bidirectional signaling of the hypothalamic-pituitary-gonadal (HPG) axis, leaving a critical therapeutic gap. Immune-isolating hydrogel capsules offer a promising strategy for the implantation of donor ovarian tissue without immunosuppression yet they require optimization for human applications. Here, we engineer a microporous immune-isolating capsule by incorporating thermosensitive gelatin microgels as sacrificial porogens. Microfluidic fabrication yielded monodisperse microgels that dissolved at 37{degrees}C generating disconnected micropores within a non-degradable poly(ethylene glycol) (PEG) matrix. Critically, the diffusion of FSH-scale analogs (40 kDa) increased by almost two-fold through the microporous capsules relative to nanoporous controls, while antibody-scale molecules (150 kDa) were blocked, demonstrating size-discriminating permeability. In ovariectomized mice implanted with encapsulated ovarian xenografts for 20 weeks, microporous capsules restored dynamic HPG-axis signaling evidenced by elevated levels of estradiol and progesterone, FSH suppression, and fluctuating hormone levels that resembled physiological patterns. Microporosity also improved graft viability, increasing stromal cellularity and reducing follicular apoptosis. These findings support microporous immune-isolating capsules as a platform for physiologically authentic therapy for POI.

bioengineering↗

bioRxiv2026

TAR syndrome causal gene RBM8A is critical for embryonic bone development and proper Hedgehog signaling

RNA-Binding Motif Protein 8A (RBM8A) is a core component of the exon junction complex (EJC), which plays a fundamental role in post-transcriptional RNA regulation. Mutations in the RBM8A gene cause thrombocytopenia-absent radius (TAR) syndrome, characterized by reduced platelet number and radial defects. However, how its function contributes to bone development remains poorly understood. We found that RBM8A is highly expressed in human bone marrow-derived mesenchymal stem cells (MSCs) and mouse embryonic limb buds. To further investigate the role of RBM8A in bone development, we generated an Rbm8a conditional knockout mouse (cKO) model in the early limb bud and craniofacial mesenchyme using the Prx1-cre line, which recapitulates key features of TAR syndrome. Rbm8a deficiency in this mouse model resulted in significant defects in limb and craniofacial development, including shorter limb bones across developmental timepoints, delayed ossification, altered cranial morphogenesis, along with impairments in motor function and reduced brain size. To elucidate the molecular basis of these phenotypes, we performed RNA immunoprecipitation sequencing (RIP-seq) in embryonic limb tissue. Our analysis revealed that RBM8A preferentially associates with long, exon-rich transcripts with high isoform complexity. These transcripts are enriched in pathways related to primary cilia, RNA processing, and developmental signaling. Consistent with this, transcriptomic and enrichment analyses identified strong associations with ciliopathy-related and congenital skeletal disease gene sets. At the cellular level, Rbm8a deficiency resulted in altered expression of key chondrogenic and ossification markers in embryonic limb bone tissue, such as reduced SOX9 and increased IHH in the primary ossification center, suggesting impaired progression of endochondral ossification and disrupted Hedgehog pathway activity. Quantitative Reverse Transcription-PCR (qRT-PCR) analysis further validated selective downregulation of Hedgehog pathway targets, including Gli3 and Hhip, while other developmental genes remained unchanged, indicating pathway-specific attenuation. Together, these findings support a model in which Rbm8a regulates structurally complex transcripts that are enriched in ciliary and developmental signaling pathways, and that disruption of this process leads to selective impairment of Hedgehog signaling and skeletal development. These results provide a novel mechanistic link between RNA processing and developmental signaling and offer new insight into how defects in RNA regulation contribute to congenital skeletal abnormalities.

developmental biology↗

bioRxiv2026

Structural insights into antibody responses against influenza A virus in its natural reservoir

While influenza A virus undergoes rapid antigenic drift in humans, at least some subtypes, such as H3, have relatively stable antigenicity in natural waterfowl reservoirs, despite the presence of immune pressure. However, the underlying mechanisms remain poorly understood. This study identified and characterized 187 antibodies to H3 hemagglutinin from experimentally infected mallard ducks, 18 of which were further analyzed by cryo-EM. Compared with human H3 antibodies, duck H3 antibodies exhibited higher glycan-binding propensity, more balanced immunodominance hierarchy, and targeted distinct epitopes. Other unique features of duck H3 antibodies included a convergent CDR H3-independent heavy chain-only binding mode and an N-glycosylated CDR H3 as decoy receptor. By annotating duck immunoglobulin germline genes, we also demonstrated the importance of gene conversion in duck H3 antibodies. Overall, our findings provide insights into how millennia of coevolution have shaped the interplay between influenza A virus antigenic drift and antibody responses in the natural reservoir.

immunology↗

bioRxiv2026

Learning a Continuous Progression Trajectory of Amyloid in Alzheimer's disease

BACKGROUNDUnderstanding of early Alzheimer progression is critical for timely diagnosis and treatment evaluation, but traditional diagnostic groups often lack sensitivity to subtle early-stage changes. METHODSWe developed SLOPE, an unsupervised dimensionality reduction method that models the amyloid progression in AD on a continuous scale which preserves the temporal sequence of follow-up visits. Applied to longitudinal amyloid PET data, SLOPE generated a two-dimensional trajectory capturing global amyloid accumulation across the AD continuum. RESULTSSLOPE-derived pseudotime scores better preserved temporal consistency across diagnostic groups and longitudinal follow-up visits and can be generalized to held-out subjects. The learned trajectory revealed biologically consistent amyloid spreading patterns and greater sensitivity to early progression than global amyloid SUVR. DISCUSSIONSLOPE provides a continuous staging of amyloid pathology that complements global amyloid measures by capturing early localized progression.

bioinformatics↗

bioRxiv2025

Deep learning inference of universal dormancy pseudotime reveals the cellular targets of anti-cancer therapies

Controlled exit from and re-entry into the cell cycle is essential for multi-cellular life, while aberrant quiescent and senescent cell states have been implicated in age-related diseases and cancer treatment evasion. Recent molecular and imaging studies suggest non-cycling cellular states exist along a continuum of deepening dormancy, whereby the probability of cell cycle re-entry decreases with distance from the restriction point. We trained a probabilistic deep-learning model that enables mapping of heterogeneous single cell transcriptomic datasets into an interpretable latent space that encodes a common "dormancy pseudotime". We demonstrate that our model enables robust inference of active cell cycle states, and validate in diverse biological contexts that the inferred location along dormancy pseudotime represents a continuum from quiescence to durably arrested states. Applying dormancy pseudotime inference to pre- and post-treatment time points from patients undergoing anti-cancer treatment, we uncover new insights into the distinct tumour cell dormancy states targeted by immune checkpoint inhibitors and platinum-taxane chemotherapy. Given the ubiquity of single cell transcriptomics, we anticipate that dormancy pseudotime analysis will be widely applied to shed new light on the complex interplay between cycling and non-cycling cellular states in health and disease.

bioinformatics↗

bioRxiv2025

Vulnerability in the breadth evolution of an influenza broadly neutralizing antibody

The highly conserved influenza hemagglutinin (HA) stem domain is a major target for broadly neutralizing antibodies (bnAbs). However, despite being discovered more than a decade ago, the IGHV1-69-encoded CR9114 remains the only HA stem bnAb that cross-reacts with both influenza A and B viruses. To investigate the constraints on the breadth evolution of CR9114, this study performs four deep mutational scanning experiments to compare the binding affinity landscapes of the germline and somatic CR9114 against H1 HA, H3 HA, and influenza B HA. Many mutations that minimally affect or even improve the H1 HA binding are detrimental for binding to H3 HA and BHA. We further reveal the prevalence of epistasis in IGHV1-69 HA stem bnAbs. Overall, our findings provide a mechanistic explanation for the scarcity of HA stem bnAbs with cross-reactivity against both influenza A and B viruses, and have important implications for developing broadly protective influenza vaccines.

immunology↗

bioRxiv2025

Mining thermophile photosynthesis genes: a synthetic operon expressing Chloroflexota species reaction center genes in Rhodobacter sphaeroides

Photosynthesis is the foundation of the vast majority of life systems, and therefore the most important bioenergetic process on earth, and the greatest diversity in photosynthetic systems are found in microorganisms. However, understanding of the biophysical and biochemical processes that transduce light to chemical energy has derived from the relatively small subset of proteins from microbes that are amenable to cultivation, in contrast to the huge number of microbial DNA sequences encoding proteins that catalyze the initial photochemical reactions that has been deposited in databases, such as from metagenomics. We describe the use of a Rhodobacter sphaeroides laboratory strain for expression of heterologous photosynthesis genes to demonstrate the feasibility of mining this resource, focusing on hot spring Chloroflexota gene sequences. Using a synthetic operon of genes, we produced a photochemically active complex of reaction center proteins in our biological system. We also present bioinformatic analyses of anoxygenic type II reaction center sequences from metagenomic samples collected from hot (42-90{degrees} C) springs available through the JGI IMG database, to generate a resource of diverse sequences that potentially are adapted to photosynthesis at such temperatures. These data provide a view into the natural diversity of anoxygenic photosynthesis, through a lens focused on high-temperature environments. The approach we took to express such genes can be applied for potential biotechnology purposes as well as for studies of fundamental catalytic properties of these heretofore inaccessible protein complexes.

microbiology↗

bioRxiv2025

Embodied Intelligence Unlocks Autonomous Microscopy

Advanced microscopy is a cornerstone of modern science, yet its potential is constrained by a reliance on manual operation, which struggles with the complexity and reproducibility of long-term experiments protocols. While scripting offers a degree of automation, it lacks the generalizability to adapt to new samples or dynamic biological events. A fundamental challenge is the absence of an intelligent system that can interpret high-level scientific intent and ground it in the physical actions of the microscope. To bridge this gap, we introduce an Embodied Intelligent Microscope System (EIMS) with hierarchical reasoning structure that reimagines the microscope autonomous control. This system leverages the advanced reasoning of large models to interpret complex user commands and decompose them into actionable steps. To solve the critical grounding problem, we constrain the models output to the feasible action space, effectively serving as the models "hands and eyes" in the physical world. We demonstrate that our system achieves zero-shot generalization on complex, multi-step protocols and successfully automates challenging biological missions requiring expert-level judgment, such as capturing the sparse spatiotemporal events of cell mitosis and locating scatteredly distributed organoids. This work establishes a new paradigm for scientific instrumentation--fusing high-level intent understanding with grounded, dynamic execution--and provides a generalizable framework for deploying embodied intelligence to accelerate autonomous scientific discovery.

bioengineering↗

bioRxiv2025

Evolution of antibody cross-reactivity to influenza H5N1 neuraminidase from an N2-specific germline

The ongoing spread of highly pathogenic avian influenza H5N1 clade 2.3.4.4b virus in animals and its occasional spillover to humans have raised concerns about a potential H5N1 pandemic. Although recent studies have shown that pre-existing human antibodies can recognize H5N1 neuraminidase, the molecular basis of how this cross-reactivity develops remains poorly understood. In this study, we used a phage display antibody library derived from 245 healthy donors to isolate an antibody, HB420, that cross-reacts with neuraminidases of human H3N2 and avian H5N1 clade 2.3.4.4b viruses and confers protection in vivo. Cryo-EM analysis reveals that HB420 targets the neuraminidase active site by mimicking sialic acid binding through a single Asp residue. Furthermore, the inferred germline of HB420 is N2-specific but acquires cross-reactivity to H5N1 neuraminidase through somatic hypermutation. Overall, our findings provide insights into how neuraminidase antibody evolves breadth, which has important implications for the development of broadly protective influenza vaccines.

immunology↗

bioRxiv2025

Mouse Bio-behavioral Phenotyping Using a Digital Homecage Framework for Long-timescale, High-resolution, and Multi-factor Data Collection and Analytics

MOTIVATION Long-term monitoring of behavioral and physiological processes is critical for understanding complex brain-based phenomena and disorders that develop over extended periods, such as chronic stress, circadian disorders, and metabolic conditions. While digital phenotyping is available in humans using smart devices, there remains a deficit in rodent models. To address this lack of long-term rodent phenotyping, we introduce the "Digital Homecage" (DHC) system. Our system uses accessible components and is designed for seamless integration with brain recording technologies. The Digital Homecage (DHC) allows uninterrupted, long-timescale recording of more than 20 behavioral metrics in single-housed mice, captured at sub-second resolution via video, operant interactions, and wheel-running data. This report demonstrates the DHCs capacity to enable continuous, automated tracking of behaviors like actigraphy, sleep, grooming, and food choice options over weeks, thereby opening up new avenues for longitudinal analyses of chronic conditions. Data collected reveal circadian patterns in multiple spontaneous behaviors, aligning with known nocturnal tendencies. The systems potential to facilitate groundbreaking observations on the long-term behavioral correlates of various neuropsychiatric syndromes is aided by open-source software and relatively low cost. The DHC sets the stage for community-driven innovation, potentially transforming our approach to studying complex traits of brain function and behavior in laboratory settings.

animal behavior and cognition↗

bioRxiv2025

Rapid sexual reproduction in a mixotrophic dinoflagellate revealed through temporal partitioning of cellular processes

Dinophysis are specialist mixotrophs that must balance prey capture, cell division, and sexual recombination during blooms, yet relatively little is known about the occurrence and role of sex in their ecology. Here, mating was investigated in D. acuminata through continuous automated microscopy of cells in culture and during natural blooms. Both in culture and in situ, vegetative division and mating were phased on a diel timescale, occurring primarily at night and near dawn, regardless of prey availability. When prey were available, feeding occurred primarily during daylight hours. The sequencing of division and mating phases, the correlation of their daily amplitudes, and the timing of their establishment after a two-day light block suggests linkage of these processes. Mating, though frequent (up to 14% day-1 in culture), was also not associated with zygote accumulation or resting stage formation but rapid cell reproduction via meiosis. Confinement of division and mating to nighttime and early morning may minimize conflict with photosynthesis-related metabolism and/or predator exposure. Sexual reproduction was the dominant mode of proliferation during the observed blooms, accounting for 71% and 64% of new cell production in 2015 and 2021, respectively. Because it is dependent on encounter of a compatible gamete, sexual reproduction is increasingly accessible as blooms intensify. This sexual mode of proliferation may also alleviate populations susceptibility to pathogens, parasites, and other threats via genetic recombination, an example of Red Queen dynamics.

ecology↗

bioRxiv2025

The lysine side chain's swing motion in AapA1 toxin promotes water transport through transmembrane pores

The rising antibiotic resistance of Helicobacter pylori and the emergence of multi-drug-resistant strains pose significant challenges to its eradication. Understanding the molecular mechanisms driving this resistance is critical. This study investigates how AapA1, a toxin protein, may inhibit H. pylori growth and cause morphological changes by forming transmembrane pores in its inner membrane. Using experimental data and molecular dynamics simulations, we analyzed the pore structure and its formation mechanism. Results show that AapA1 forms pores on liposomes and exists stably in the membrane as -helices through different multimers, creating hydrophilic pores of varying sizes. Dynamic oscillation of K16/23 side chains in the membrane affects water transport through these pores, and restricting their movement significantly reduces transport. AapA1 monomers adopt three distinct conformations, with the K16/23 residues showing an up-down swing motion, potentially facilitating accelerated transport of substances. This study provides a new perspective on the dynamic structure-function relationship of AapA1-membrane interaction and highlights the role of positively charged residues in AapA1. Understanding the multimeric AapA1-membrane structure and the mechanism underlying AapA1-induced transmembrane pores is particularly crucial to develop new drugs and treatment options to reduce drug resistance emergence and combat any new emergence development.

biophysics↗