Biology subjects

Shan, Z.

Publications and source records attributed to Shan, Z..

At least 19 recordsLinked to original sources

bioRxiv2026

Exploratory Proteomics of Brain Extracellular Vesicle Enriched Preparations After LPS Challenge and miRNA Associated Vesicle Administration

We used brain extracellular vesicle (EV) proteomics to identify protein responses to acute neuroinflammation and an EV-based microRNA intervention. Rats received vehicle, intracerebroventricular lipopolysaccharide (LPS), or LPS followed by an intravenous EV formulation containing anti-inflammatory microRNAs (miRNAs). Brains were collected 24 h after LPS or vehicle administration, and three independently processed animal-derived EV-enriched preparations per group were analyzed by label-free proteomics. Across all nine samples, 953 protein groups had complete normalized abundance measurements. Compared with vehicle controls, LPS exposure was associated with 227 exploratory protein changes, comprising 95 increases and 132 decreases at nominal P < 0.05 and an absolute log2 fold change >= log2(1.5). Increased proteins were enriched for translation-related annotations, whereas decreased proteins were enriched for plasma-membrane annotations. These LPS-associated changes did not remain significant after Benjamini-Hochberg correction at a false discovery rate (FDR) threshold of q < 0.05. Following treatment, 119 of the 227 candidates were closer to control abundance on the log2 scale; 23 candidates also showed nominal treatment-versus-LPS differences. Lamb2 and Ptgfrn were the two proteins that differed significantly between treated and LPS-only animals after FDR correction, identifying them as priorities for further validation. Nevertheless, 174 proteins differed significantly between treated animals and vehicle controls, and analysis across the measured proteins did not demonstrate an overall shift toward control values. Thus, the intervention was associated with specific protein responses, but treated animals retained substantial differences from controls. This study identifies candidate markers for investigating treatment responses in brain EV-enriched preparations and demonstrates the value of examining individual proteins alongside broader proteomic changes. Independent validation is needed to determine their functional relevance and distinguish miRNA-specific effects from contributions of the administered EV carrier.

neuroscience↗

bioRxiv2026

Defense-associated reverse transcriptases synthesize DNA in a telomerase-like manner

Defense-associated reverse transcriptases (DRTs) constitute an emerging class of bacterial defense systems that harness reverse-transcriptase activity to generate cDNA products and counter phage infection. A distinct subset of DRT10 systems evolutionarily related to telomerase generates tandem-repeat DNA to confer antiphage immunity. Here, we biochemically characterize the reverse transcription activity of DRT10 and determine cryo-EM structures of DRT10-ncRNA complex in three functional states, revealing an unanticipated 2:1 RT:ncRNA assembly mode. Two DRT10 protomers dimerize through a central pair of interlocked {beta}-hairpins and are further embraced by a single ncRNA. This architecture distinguishes DRT10 from previously characterized DRT complexes, which either lack an ncRNA component or exhibit an equivalent stoichiometry of RT:ncRNA. Strikingly, despite the nearly identical conformations of the two DRT10 protomers, cDNA product density was resolved in only one catalytic pocket, revealing different product occupancy within the dimer. Moreover, along with flanking linkers, the stem loops of the ncRNA as fixed points to supply steric barriers help define the template boundary. These findings reveal a distinctive ncRNA-mediated DRT10 dimeric assembly and establish a mechanistic framework for protein-primed reverse transcription constrained by the length of the RNA flanking linker.

biochemistry↗

bioRxiv2026

Bacterial Peptidoglycan Extends Lifespan by Activating Lysosomal Activity through V-ATPase Binding

Lysosomal dysfunction is a hallmark of aging, yet whether microbial components actively regulate this organelle to influence longevity remains unknown. Here, we identify bacterial peptidoglycan (PGN), a major cell wall component degraded by host lysozyme, as an evolutionarily conserved activator of lysosomal function that extends lifespan in both C. elegans and mice. We show that aging leads to an intestinal decline in lysozyme expression, which impairs bacterial cell-wall digestion and results in systemic PGN deficiency. Late-life PGN supplementation (starting at 18 months of age) significantly prolongs mouse lifespan and improves healthspan. Mechanistically, PGN localizes to lysosomes and directly binds V-ATPase subunits, enhancing ATP hydrolysis activity and promoting lysosomal acidification. This effect is abolished by V-ATPase inhibition (bafilomycin A1) or genetic disruption of lysosomal components (cup-5 and vha-12 mutants), confirming that functional V-ATPase is strictly required for lysosomal function and the longevity benefit. Importantly, PGN restores lysosomal acidification in aged cells, alleviates cellular senescence markers, and improves multiple hallmarks of aging including locomotion and muscle integrity. Collectively, these findings reveal an evolutionarily conserved mechanism whereby hosts exploit bacterial cell wall components to maintain cellular homeostasis, establishing a gut microbiome-lysosome-longevity axis with implications for microbiome-based anti-aging interventions.

physiology↗

bioRxiv2026

Spatial MALDI-MSI Reveals a Coordinated Vicious Cycle of Oxidative Membrane Damage and Ceramide-Driven Sphingolipid Dysregulation in the Chronically Neuroinflamed Brain

BackgroundChronic neuroinflammation is a major driver of cognitive decline, vascular cognitive impairment, and Alzheimers disease. However, the spatial lipidomic alterations underlying neuroinflammatory brain injury remain poorly defined. Oxidative stress and sphingolipid dysregulation have been implicated, but their regional distribution and interplay in the brain are not well characterized. MethodsWe performed positive-ion mode matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI) on coronal brain sections from middle-aged spontaneously hypertensive rats (SHR), a model of chronic neuroinflammation, and normotensive Wistar-Kyoto (WKY) controls. Spatial distributions and relative abundances of multiple lipid classes, including phosphatidylcholines (PCs), sphingomyelins (SMs), hexosylceramides (HexCers), ceramides, phosphatidylserines (PSs), phosphatidylinositols (PIs), phosphatidylethanolamines (PEs), phosphatidic acids (PAs), and sulfatides, were mapped and compared between genotypes. Region-of-interest analysis was used to quantify changes across cortex, hippocampus, and white-matter tracts. ResultsSHR brains exhibited a coordinated lipidomic signature characterized by pronounced oxidative stress and membrane remodeling. Oxidized and short-chain PCs were markedly upregulated (up to 11.6-fold), while major structural diacyl PCs were broadly downregulated. Concurrently, sphingolipids were significantly altered, with robust upregulation of SM(d36:1) (7.5-fold) and multiple HexCer species (1.5-1.9-fold), accompanied by accumulation of ceramides. These changes were accompanied by heterogeneous redistribution of PS, PI, and PE species, particularly within the hippocampus. Sulfatide patterns in white-matter tracts were also altered, suggesting myelin remodeling. Region-of-interest analysis confirmed that the most pronounced lipid alterations were concentrated in the hippocampus and white-matter regions. ConclusionsChronic neuroinflammation induces a spatially organized, multi-class lipid remodeling response in the brain, driven by advanced oxidative membrane damage and a shift toward a pro-apoptotic sphingolipid profile. The convergence of these pathways creates a vicious cycle of membrane injury, mitochondrial dysfunction, and sustained neuroinflammation that is especially prominent in the hippocampus and white matter. These spatially resolved findings provide direct evidence that oxidative stress and sphingolipid dysregulation are central, interrelated mechanisms contributing to neurovascular injury and increased risk of cognitive impairment. The study highlights the power of MALDI-MSI to uncover region-specific lipid pathology and identifies potential lipid-based targets for therapeutic intervention in neuroinflammatory brain disease. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=140 SRC="FIGDIR/small/738110v1_ufig1.gif" ALT="Figure 1"> View larger version (47K): org.highwire.dtl.DTLVardef@853af9org.highwire.dtl.DTLVardef@e2d079org.highwire.dtl.DTLVardef@e4e065org.highwire.dtl.DTLVardef@480d80_HPS_FORMAT_FIGEXP M_FIG C_FIG

neuroscience↗

bioRxiv2026

Structural Mechanism of Filamentation Induced Dampening of GTP Inhibition of Glutamate Dehydrogenase

Glutamate dehydrogenase (GDH) is a highly regulated key enzyme that catalyzes the reversible oxidative deamination of glutamate to alpha-ketoglutarate, positioning it at a critical hub linking amino acid catabolism to energy production while supplying ammonia for urea synthesis and other nitrogen pathways. Early investigations have shown that bovine GDH (bGDH), which shares 98% sequence identity with its human homolog, assembles into polymeric filaments with altered allosteric responses. Filamentation has only relatively recently been appreciated as a widespread mechanism of enzyme regulation, prompting a reevaluation of these early observations in GDH. Here, we use high resolution cryogenic electron microscopy (cryo EM) to show that bGDH hexamers assemble via reciprocal antenna interactions that oppose the conformational changes associated with GTP inhibition, revealing how filamentation reshapes GDH allostery and with implications for the treatment of human disease.

biochemistry↗

bioRxiv2026

Age-Dependent Spatiotemporal Remodeling of Brain Sphingolipids During LPS-Induced Neuroinflammation: MALDI-MSI Reveals Accelerated Sphingomyelin Depletion and Sulfatide Accumulation Linked to Mitochondrial Oxidative Stress

Aging is a major risk factor for exacerbated neuroinflammation and neurodegenerative diseases, yet the underlying lipid metabolic mechanisms remain incompletely understood. Here, we employed high-resolution matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI) combined with quantitative peak-area analysis and conceptual kinetic modeling to investigate age-dependent sphingolipid remodeling in the rat brain following intracerebro-ventricular (ICV) LPS challenge. In old rats, MALDI-MSI revealed pronounced and progressive sphingolipid dysregulation compared with young animals. Quantitative analysis showed a dramatic [~]10-fold reduction in SM(d36:1) and [~]4-fold reduction in SM(d42:2), accompanied by significant accumulation of long-chain sulfatides (2.12-fold increase in C24:1-sulfatide and 1.45-fold increase in C24(OH)-sulfatide) at both 24 h and 72 h post-LPS. Spatial imaging demonstrated that these changes were markedly amplified in white matter regions and became more widespread and intense at 72 h. A simplified Michaelis-Menten kinetic model successfully recapitulated the experimental data, identifying increased nSMase2 activity (higher Vmax) as the primary driver of accelerated sphingomyelin hydrolysis and subsequent ceramide rerouting into sulfatide synthesis. This metabolic shift generates excess ceramide that promotes Drp1-mediated mitochondrial fission, elevates mitochondrial ROS production, and disrupts bioenergetics, establishing a feed-forward loop linking sphingolipid remodeling to mitochondrial oxidative stress and white matter vulnerability in the aged brain. These findings provide the first spatially and temporally resolved demonstration of age-dependent sphingolipid metabolic reprogramming during neuroinflammation. By integrating multimodal MALDI-MSI, quantitative lipidomics, and kinetic modeling, this study reveals a previously underappreciated nSMase2-ceramide-mitochondrial axis in neuroinflammaging. Graphic Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=179 SRC="FIGDIR/small/734865v1_ufig1.gif" ALT="Figure 1"> View larger version (50K): org.highwire.dtl.DTLVardef@be7a9eorg.highwire.dtl.DTLVardef@164499dorg.highwire.dtl.DTLVardef@11cea50org.highwire.dtl.DTLVardef@15b2ac2_HPS_FORMAT_FIGEXP M_FIG C_FIG

neuroscience↗

bioRxiv2026

Platelets promote acute liver injury via extracellular vesicles-mediated Aldolase A

Platelets have emerged as active regulators of acute liver injury (ALI), yet the molecular mechanisms underlying their pathogenic functions remain poorly defined. Here, we demonstrate that platelets contribute to liver injury, in part, by metabolically reprogramming Kupffer cells (KCs). We show that platelets are actively recruited to the injured liver and communicate with KCs through extracellular vesicles (EVs), which deliver the glycolytic enzyme aldolase A (ALDOA) into recipient cells. This EV mediated transfer promotes glycolytic reprogramming in KCs, contributing to their proinflammatory activation and amplification of hepatic injury. Genetic ablation of Aldoa in platelets attenuated liver injury, demonstrating a pathogenic role for platelet derived ALDOA in vivo. Pharmacological inhibition of ALDOA with Aldometanib markedly attenuated liver injury. Circulating ALDOA levels were elevated in patients with ALI and correlated with disease severity. These findings uncover a platelet macrophage metabolic axis that drives ALI and nominate ALDOA as a therapeutic target and biomarker.

pathology↗

bioRxiv2026

Chronic Paraventricular OX1R Overexpression Induces Oxidative Stress and Hypertension in Rats

BackgroundHyperactivity of the orexin system has been implicated in hypertension, yet the long-term impact of orexin 1 receptor (OX1R) overexpression in the paraventricular nucleus (PVN), a key brain region involved in arterial blood pressure (ABP) regulation, remains unclear. This study examined whether chronic OX1R overexpression in the PVN alters cardiovascular, neuroendocrine, and oxidative functions. MethodsAdult male Sprague-Dawley rats received bilateral PVN injections of AAV2-OX1R or control virus AAV2-GFP. Their ABP and heart rate (HR) were monitored for eight weeks via radiotelemetry. Renal sympathetic nerve activity (RSNA) and ABP responses to PVN orexin A administration were assessed using in vivo recordings. Reactive oxygen species (ROS) were quantified using MitoProbe, and gene expression was analyzed by qPCR. Primary neuronal cultures were used to explore underlying mechanism involved in increased ROS production and neuronal activity. ResultsPVN OX1R overexpression increased mean ABP ([~]10 mmHg), water intake, and ROS levels in both the PVN and peripheral organs. In addition, orexin A evoked exaggerated RSNA (155% vs. 73%) and pressor responses (27 vs. 16 mmHg) in PVN OX1R overexpression rats compared to control rats, these effects were attenuated by the OX1R antagonist SB408124. Plasma vasopressin levels were elevated, and brain sections confirmed increased ROS in OX1R-positive neurons. In vitro experiments further demonstrated that orexin A upregulated ROS-generating enzymes, suppressed antioxidant genes, and enhanced neuronal excitability. ConclusionChronic PVN OX1R overexpression induces oxidative stress, sympathetic overactivity, and increased plasma vasopressin, contributing to hypertension development. Targeting PVN OX1R signaling may provide therapeutic benefit for hypertension.

physiology↗

bioRxiv2025

Distinct neural processing of real versus unreal optic flow in the human brain

Visual perception of self-movement relies on optic flow, a global motion pattern generated by self-movement in a rigid 3D environment. Such optic flow conveys both 2D features (e.g., radial velocity fields and centers of motion) and 3D structural information (e.g., motion perspective). However, many previous neuroimaging studies have used simplified 2D radial motion as "real" optic flow, leaving it unclear whether such "unreal" stimuli adequately capture how the brain processes ecologically valid self-movement information. Previous behavioral work has shown that sensitivity to expansion and contraction reverses depending on whether optic flow is ecologically valid (L. Li et al., 2025), raising the question of how such differences are represented in the human brain. We conducted two functional magnetic resonance imaging (fMRI) experiments to systematically compare cortical processing of real versus unreal optic flow. The two stimulus types were strictly matched in 2D global features and local motion signals, differing only in whether they preserved 3D structure consistent with self-movement. To minimize task-related influences, participants performed a task-irrelevant fixation point color-change detection task during scanning. Experiment 1 used a block design in which participants viewed expanding and contracting optic flow at different motion coherence levels. Experiment 2 presented real and unreal optic flow within the same scanning session to directly test neural sensitivity to ecological validity. Multivoxel pattern analysis (MVPA) revealed that, among multiple visual and optic flow-responsive regions, the human dorsal medial superior temporal area (MST) exhibited robust and consistent sensitivity to the ecological validity of optic flow. Specifically, under real optic flow, decoding accuracy for motion coherence in MST was significantly higher for expansion than for contraction, whereas this preference was reversed under unreal optic flow. Neural threshold estimates derived from fMR-metric functions further supported this pattern reversal, indicating systematic modulation of MST sensitivity by ecological structure. Experiment 2 further demonstrated that, regardless of motion pattern, MST reliably distinguished real from unreal optic flow, with decoding accuracy increasing as motion coherence increased. These findings indicate that MST encodes not only motion patterns themselves but also whether optic flow conforms to the ecological structure of self-movement. Together, these results indicate that MST responses are not determined solely by 2D radial motion features, but are modulated by whether optic flow preserves 3D structure consistent with self-movement. This work clarifies a long-standing conflation between simplified radial motion and real optic flow and highlights the importance of ecological structure in shaping dorsal-stream motion representations related to self-movement perception.

neuroscience↗

bioRxiv2025

IgG-Bridging-Seeded Synergistic Aggregation of SARS-CoV-2 Spikes Underlies Potent Neutralization by A Low-Affinity Antibody

Mechanistic studies of viral neutralization typically prioritize high-affinity antibodies, relegating low-affinity binders to the sidelines. We report P5-1C8, a Class 1 SARS-CoV-2 antibody that exemplifies this underexplored "low-affinity yet high-potency" phenotype, retaining strong neutralization of Omicron JN.1 despite markedly weakened trimer binding (KD = 225 nM; IC50 = 0.06 nM). Structural and biophysical analyses reveal that P5-1C8 engages WT and BA.1 spikes through canonical intra-spike bivalency, but with JN.1 it induces aggregation. Using virion-like nanoparticles displaying multiple spikes, we show that IgG remains bound with no detectable dissociation and triggers pronounced aggregation. Coarse-grained molecular dynamics delineate the stepwise pathway in which weak IgG-spike contacts seed aggregation via transient inter-spike bridging. Together, these findings establish the first mechanistic framework demonstrating how weak-binding antibodies can nonetheless achieve potent neutralization through higher-order aggregation, thereby expanding the conceptual landscape of antibody function and opening new directions for antibody evaluation and design. Graphical AbstractLow-affinity antibodies are frequently disregarded in discovery pipelines. This work reports P5-1C8, a Class 1 SARS-CoV-2 antibody with weak trimer binding (KD-to-IC50 > 3,700-fold) yet potent neutralization of Omicron JN.1. Structural, biophysical, functional and coarse-grained simulations collectively demonstrate that transient inter-spike IgG bridging seeds higher-order aggregation, which in turn drives neutralization and provides a mechanistic framework. O_FIG O_LINKSMALLFIG WIDTH=199 HEIGHT=200 SRC="FIGDIR/small/691454v1_ufig1.gif" ALT="Figure 1000"> View larger version (79K): org.highwire.dtl.DTLVardef@1d4f132org.highwire.dtl.DTLVardef@1274400org.highwire.dtl.DTLVardef@e186a6org.highwire.dtl.DTLVardef@4f1870_HPS_FORMAT_FIGEXP M_FIG C_FIG

cell biology↗

bioRxiv2025

PRSNet-2: End-to-end genotype-to-phenotype prediction via hierarchical graph neural networks

The emergence of large-scale biobanks has opened unprecedented opportunities for the development of data-driven approaches, especially deep learning-based methods, for genotype-to-phenotype (G2P) prediction. However, designing an end-to-end framework capable of directly leveraging extremely high-dimensional genotypic data while simultaneously mitigating overfitting and ensuring robust prediction remains a significant challenge. In this study, we introduce PRSNet-2, an interpretable end-to-end deep learning framework designed to predict complex phenotypes directly from large-scale genotypic data. PRSNet-2 proposes a novel hierarchical graph neural network (GNN) architecture, which first employs a multi-kernel aggregator to map high-dimensional genotypic features to gene-level representations. Next, it models gene-gene interactions through message-passing operations and uses an attention-based readout module to generate interpretable phenotypic predictions. We further introduce significance-guided regularization strategies to boost models generalizability based on prior genetic associations. Extensive empirical evaluations across multiple complex traits and diseases demonstrate that PRSNet-2 consistently outperforms a variety of baseline methods and exhibits superior capabilities in overcoming overfitting, even when modeling around half a million single nucleotide polymorphisms (SNPs). Moreover, PRSNet-2 is shown to be easily extendable to integrate multiple genome-wide association study (GWAS) datasets into a single model, thereby enhancing predictive performance for both single-phenotype and multi-phenotype prediction tasks. The inherent interpretability of PRSNet-2 further facilitates the identification of disease-relevant genes, functional gene modules, and potential therapeutic targets. In summary, PRSNet-2 offers a powerful, versatile tool for both genetic risk stratification and the discovery of biological insights.

genetics↗

bioRxiv2025

Copper-Free Click Chemistry Enables High-Fidelity Engineering of Mitochondria Targeted Brain-Derived Exosomes

Mitochondrial dysfunction is a hallmark of neurodegenerative and neuroinflammatory disorders, including hypertension and cardiovascular disease, yet strategies for safe and precise mitochondrial-targeted delivery remain limited. Here, we establish strain-promoted azide-alkyne cycloaddition (SPAAC) as a biocompatible, high-fidelity chemical platform for engineering mitochondria-targeted brain-derived exosomes (BR-EVs). Copper-free click conjugation of a mitochondrial-targeting ligand (e.g. Cy5-DBCO) under mild aqueous conditions preserved vesicle morphology (30-150 nm core; 120-200 nm hydrodynamic), proteomic composition, and uptake dynamics. Time-course imaging and fluorescence recovery after photobleaching (FRAP) revealed unaltered endocytic kinetics, >75 % mitochondrial colocalization, and intact organelle architecture. In vivo neuroinflammation and biodistribution analyses demonstrated immunological neutrality, strong central nervous system retention, and minimal peripheral dispersion following intracerebroventricular administration. Proteomic profiling of unlabeled Sprague-Dawley (SD) and hypertensive Dahl salt-sensitive (DSS) BR-EVs uncovered hypertension-driven enrichment of oxidative and complement pathways correlating with mitochondrial fragmentation and reactive oxygen species generation in neuronal cultures. These findings establish SPAAC-mediated ligand conjugation as a biocompatible and chemically precise approach for generating mitochondria-targeted exosomes that preserve exosomal identity, biodistribution, and signaling fidelity--advancing a foundational platform for organelle-specific delivery and mechanistic imaging in the central nervous system.

neuroscience↗

bioRxiv2025

A Reversible Mitochondrial ROS Probe for Monitoring Mitophagy Dynamics: Development and Application of MitoFlare

Mitochondrial dysfunction and defective mitophagy are defining features of numerous neurodegenerative and metabolic disorders, yet existing tools provide limited ability to quantify mitophagy dynamics in real time within living, post-mitotic cells. Here we present MitoFlare, a mitochondria-targeted, reversible mtROS-responsive fluorogenic probe that enables continuous, non-genetic visualization of mitochondrial oxidative activation and turnover. MitoFlare incorporates dual TEMPO nitroxide quenchers into a long-wavelength rhodamine scaffold, producing >95% basal quenching and rapid, fully reversible fluorescence activation in response to mitochondrial superoxide, hydroxyl radicals, lipid-derived peroxyl species, and peroxynitrite. When combined with LysoTracker Green, MitoFlare forms a dual-probe imaging platform that resolves the entire mitophagy cascade with high spatial and temporal fidelity in intact PC12 neuronal cells. Using this platform, we established a quantitative framework comprising three mechanistically distinct metrics: (i) a proximity index that reports early mitochondrial engagement with lysosomes, (ii) Manders M1 coefficient that captures mid-stage mitochondria-lysosome fusion and mitophagosome formation, and (iii) a quenching/swelling index that resolves terminal lysosomal degradation. Nutrient deprivation induced a complete, temporally ordered mitophagy program, including mtROS priming, Parkin-OPTN-associated fusion, and efficient acidification-dependent cargo degradation. In contrast, inhibition of v-ATPase with bafilomycin A1 arrested mitophagy at the fusion stage, resulting in persistent redox-active mitochondrial cargo that failed to undergo lysosomal digestion. Importantly, MitoFlares reversible redox chemistry uniquely revealed accumulation of undegraded, oxidatively active mitochondrial remnants within non-acidified vesicles--pathological intermediates that are undetectable using irreversible ROS dyes or genetically encoded reporters. These findings demonstrate that mitophagy proceeds through discrete, redox-regulated and lysosome-dependent phases that can be quantitatively mapped in real time. By enabling synchronized measurement of oxidative activation, organelle trafficking, fusion, and degradation, the MitoFlare-LysoTracker system establishes a new benchmark for dynamic mitophagy analysis in physiologically relevant models. This platform provides a powerful foundation for mechanistic interrogation of mitochondrial quality control and for accelerating the discovery of therapeutic strategies aimed at restoring mitophagic fidelity in neurodegenerative, cardiovascular, and metabolic diseases.

neuroscience↗

bioRxiv2025

Hyperactivated Glycolysis Drives Spatially-Patterned Kupffer Cell Depletion in MASLD

Metabolic dysfunction-associated steatotic liver disease (MASLD) progression is characterized by hepatic inflammation and cell death, yet the mechanisms underlying Kupffer cells (KCs) loss remain poorly understood. Here, we sought to elucidate the metabolic basis of KCs death during MASLD. Using metabolomics, immunostaining, and flow cytometry, we evaluated metabolic alterations and KCs death throughout early MASLD progression. We found that KCs death is an early hallmark of MASLD, exhibiting greater susceptibility and displaying a spatial distribution consistent with KCs zonation. Moreoever, KCs undergo progressive metabolic reprogramming toward enhanced glucose utilization during MASLD development, which is correlated with KCs death. In combination of biochemical agonist, isotope tracing and primary KCs culture, we further demonstrated that augmented glycolytic metabolism directly drives KCs death in vitro. Consistently, using Chil1-deficient mice, we further demonstrated that increased glucose utilization accelerates KCs death in vivo. Together, these findings establish a causal link between glycolytic activation and KCs loss during MASLD progression, highlighting glucose metabolic pathways as potential therapeutic targets to preserve KCs homeostasis and mitigate MASLD.

pathology↗

bioRxiv2025

Mid-zone hepatocytes trade proliferation for survival via Atf4-Chop axis in early acute liver injury

Hepatocytes undergo extensive proliferation to facilitate liver repair after injury, yet early adaptive changes prior to proliferation remain unclear. Here, we report that during early acetaminophen (APAP)-induced liver injury, hepatocytes exhibit transient proliferation suppression that is most pronounced in mid-zone hepatocytes, consistent with zonal APAP metabolism. While spatial transcriptomics (ST) provided robust evidence for this arrest in the mid-zone, support for a similar arrest in pericentral hepatocytes was limited. Integrating ST with immunohistochemistry and functional studies, we identified a unique mid-zone stress-response program centered on the Atf4-Chop axis, which suppresses proliferation via the cell cycle inhibitor Btg2. Together, our findings support a model in which mid-zone hepatocytes transiently prioritize stress adaptation over proliferation, thereby preserving regenerative capacity for subsequent liver repair.

pharmacology and toxicology↗

bioRxiv2025

Microbial Tryptophan Metabolism Activates Host Lysosomal Activity to Facilitate Lipid Breakdown and Ameliorate Hepatic Steatosis

Lysosomes are central to lipid metabolism, yet how gut microbiota-derived metabolites regulate lysosomal function to influence host lipid homeostasis remains unknown. Here, we identify an evolutionarily conserved mechanism in which bacterial tryptophan metabolism activates lysosomal activity to promote lipid breakdown. By developing a lysosomal-responsive lipid reporter in C. elegans to screen for bacterial metabolic states that modulate host lipid storage, we discover that E. coli tryptophan catabolism via tryptophanase TnaA induces lysosomal lipid chaperone LBP-8, driving lipid mobilization. Moreover, tryptophan metabolites enhanced lysosomal acidification and degradation capacity, while genetic disruption of lysosomal regulators reversed these effects. Strikingly, bacterial tryptophan metabolism further promoted mitochondrial {beta}-oxidation through lysosomal lipase activity. This pathway was conserved in mammalian hepatocytes, where E. coli-derived tryptophan metabolites enhance lysosomal function and reduce lipid accumulation. In high-fat diet mice, restoring gut bacterial tryptophan metabolism alleviated hepatic steatosis. Our work uncovers microbiota-regulated lysosomal activation as a critical axis in lipid homeostasis, highlighting its potential as a therapeutic target for metabolic disorders linked to lysosomal dysfunction. SignificanceWe uncover a conserved mechanism by which microbial tryptophan metabolism enhances lysosomal function to maintain host lipid homeostasis. Specifically, we demonstrate that bacterial tryptophan catabolism--via the enzyme TnaA--promotes lysosomal acidification, proteolytic capacity, and structural remodeling in C. elegans, driving lipid breakdown through the lysosomal chaperone LBP-8. This activation boosts mitochondrial {beta}-oxidation and reduces lipid storage. Importantly, the same pathway operates in mammalian hepatocytes and in a high-fat diet mouse model, where restoring bacterial tryptophan metabolism markedly alleviates hepatic steatosis. Our findings bridge microbial metabolism and lysosomal dynamics, offering fresh insights into host-microbe crosstalk and metabolic regulation. HighlightsTnaA-mediated bacterial tryptophan catabolism promotes lipid mobilization via lysosomal chaperone. Bacterial tryptophan metabolites boost lysosomal function, lipid breakdown, and mitochondrial {beta}-oxidation. Conserved microbiota-lysosome-lipid axis from worms to mammalian liver. Restoring gut bacterial tryptophan metabolism alleviates hepatic steatosis in high-fat diet mice.

cell biology↗

bioRxiv2025

Microbial Peptidoglycan Engages Autophagy Receptor P62 to Induce Protective Mitophagy in the Liver

Although mitophagy is critical for maintaining mitochondrial integrity and hepatic homeostasis, the microbial-derived signals controlling this process remain unknown. Given the gut microbiotas profound influence on liver pathophysiology, identifying specific bacterial factors that directly regulate hepatocyte mitophagy could unlock novel therapeutic strategies. In this study, we identify bacterial peptidoglycan (PGN)--a conserved cell wall component--as a key activator of mitophagy that protects against hepatocyte death. Through both in vivo and in vitro studies, we demonstrate that either heat-killed Escherichia coli or purified PGN attenuates hepatocyte death. Mechanistically, PGN is internalized by hepatocytes, localizes to mitochondria, and initiates mitophagy via direct interaction with the autophagy adaptor p62/SQSTM1. Genetic ablation of p62 in hepatocytes completely abolishes PGN-induced mitophagy, underscoring the pathways essential role. Strikingly, therapeutic administration of PGN markedly alleviates carbon tetrachloride (CCl4)-induced hepatic fibrosis, reducing collagen deposition and suppressing hepatic stellate cell activation through enhanced autophagic flux. Our work unveils a previously unrecognized host-microbe crosstalk in which PGN acts as a mitophagy inducer, offering a potential therapeutic avenue for liver diseases driven by mitochondrial dysfunction.

cell biology↗

bioRxiv2025

Differential Regulation of Hepatic Macrophage Fate by Chi3l1 in MASLD

Metabolic dysfunction-associated steatotic liver disease (MASLD) progression involves the replacement of protective embryo-derived Kupffer cells (KCs) by inflammatory monocyte-derived macrophages (MoMFs), yet the regulatory mechanisms remain unclear. Here, we identify chitinase 3-like 1 (Chi3l1/YKL-40) as a critical metabolic regulator of hepatic macrophage fate. We observed high expression of Chi3l1 in both KCs and MoMFs during MASLD development. Genetic deletion of Chi3l1 specifically in KCs significantly exacerbated MASLD severity and metabolic dysfunction, whereas MoMF-specific Chi3l1 deletion showed minimal metabolic effects. Mechanistic studies revealed that this cell type-specific regulation arises from differential metabolic requirements: KCs display elevated glucose metabolism compared to MoMFs. Chi3l1 directly interacts with glucose to inhibit its cellular uptake, thereby selectively protecting glucose-dependent KCs from metabolic stress-induced cell death while having negligible effects on less glucose-dependent MoMFs. These findings uncover a novel Chi3l1-mediated metabolic checkpoint that preferentially maintains KCs populations through glucose metabolism modulation, providing important new insights into the pathogenesis of MASLD and potential therapeutic strategies targeting macrophage-specific metabolic pathways.

molecular biology↗