Biology subjects

Xu, S.

Publications and source records attributed to Xu, S..

At least 37 records · Page 2Linked to original sources

bioRxiv2026

A gene-augmentation platform for early-stage autosomal recessive retinitis pigmentosa

Most recessive inherited retinal dystrophies are, in principle, amenable to gene augmentation, yet only one such therapy has received regulatory approval. We sought to identify tractable autosomal recessive retinitis pigmentosa (RP) genes that could be treated using existing non-proprietary adeno-associated virus (AAV) components. More than 100 RP-associated genes were prioritized according to cellular expression, coding-sequence size, and the availability of cell type-specific promoters, yielding 19 candidate genes expressed predominantly in rods and/or retinal pigment epithelium (RPE). In rhesus monkey eyes, the human RHO and BEST1 promoters drove rod- and RPE-specific expression, respectively, whereas the GFAP and RLBP1 promoters were limited by absent Muller glial expression or dose-associated RPE toxicity. We selected PDE6B as a proof-of-concept gene and evaluated AAV8-RHO-PDE6B after neonatal subretinal delivery. The vector effectively preserved outer nuclear layer structure, electroretinography responses, and visually guided behaviors in rd1 and rd10 mice for at least 6 months. Dose-ranging studies identified retinal abnormalities at the higher doses, whereas the lower doses were comparatively well tolerated in eyes of mice and rhesus monkeys. Together, these findings define a preclinical framework for developing gene-augmentation vectors for a subset of early-stage autosomal recessive RP patients.

genetics↗

bioRxiv2026

A fast 2C-induction method reveals a barrier role of SP2 for totipotency

The acquisition of totipotency in vitro remains a major challenge, limiting our understanding of early embryogenesis and its clinical and translational applications. Here, we reported a robust and rapid chemical reprogramming system that converts mouse embryonic stem cells (mESCs) into totipotent-like cells (TLCs) within 36 hours, with efficiencies exceeding 70%. The induced cells exhibit transcriptomic, epigenomic, and functional features closely resembling 2-cell (2C) stage embryos, as confirmed by scRNA-seq, ATAC-seq, and chimeric embryo analyses. Single-cell trajectory construction revealed a branching reprogramming process that yields both successful totipotent-like and alternative, non-reprogrammed fates. Mechanistically, we identified the transcription factor SP2 as a barrier to totipotency acquisition, promoting lineage-specific gene expression while repressing totipotency networks. Collectively, our study has established a highly efficient in vitro model for totipotency induction and highlighted molecular barriers shaping cell fate decisions, providing a platform to dissect the mechanisms governing the pluripotency-to-totipotency transition.

developmental biology↗

bioRxiv2026

scWeave: A deep learning model that bidirectionally translates between gene expression and chromatin structure at single cell resolution

Chromatin structure and gene expression are intimately linked, yet characterizing how the two covary has proven challenging, primarily because the two modalities are rarely measured in the same cells. Recently, single-cell co-assay protocols have enabled simultaneous profiling of both modalities within the same cells, but these experiments remain costly and technically challenging. To better characterize the relationship between 3D chromatin architecture and gene expression and to enable cross-modality inference from single-modality measurements, we developed a model called scWeave that bidirectionally translates between gene expression (scRNA-seq) and 3D chromatin architecture (scHi-C) at single-cell resolution. The scWeave model employs dual autoencoders to extract separate cell-level latent representations and learns to translate between these representations using dedicated translation modules. We evaluate scWeave on six publicly available co-assay datasets spanning mouse embryonic development, mouse cortex, mouse olfactory epithelium, and human bone marrow. On held-out mouse cells, scWeave outperforms a nearest-neighbor baseline and existing methods adapted to single-cell resolution, achieving a 57% improvement in median Spearman correlation when predicting gene expression from chromatin structure and an 18.8% improvement in median HiCRep similarity in the reverse direction relative to the next-best baseline. We further show that scWeave learns cross-modally aligned latent representations at single-cell resolution, enabling cells profiled in one modality to be matched to their counterparts in the other. Finally, scWeave generalizes to entirely held-out developmental timepoints in mouse olfactory epithelium and performs well on held-out human bone marrow cells despite limited human training data. By predicting the unmeasured chromatin architecture or transcriptional state from a single measured modality, scWeave offers a route to extend the benefits of costly co-assays to the many cell types, developmental stages, and species that are currently profiled with only one modality.

genomics↗

bioRxiv2026

Contextualized phenovectors reveal a Drosophila serotonergic circuit controlling satiety

Distinguishing whether a neural circuit genuinely controls a holistic behavioral state, rather than merely producing isolated effects, is a central challenge in neuroscience. Feeding research illustrates the problem: decades of work have reached conflicting conclusions partly because individual behavioral features are assessed in isolation. Here we establish a contextualized ethomics approach that benchmarks optogenetic circuit manipulations against natural hunger-satiety transitions, using high-dimensional behavioral tracking in Drosophila and starvation as a ground-truth intervention. Applying this framework, we find that serotonergic neurons marked by the Tryptophan hydroxylase neuronal (Trhn) enhancer are both required for and instructive of a satiety-like state, whereas other feeding-related circuits produce only fragmentary behavioral changes. Intersectional dissection localized this control to Trhn neurons of the ventral nerve cord, which act through the sugar transporter Sut2 to sense nutrient state. This work strengthens our understanding of serotonergic feeding control and provides a generalizable framework for validating circuit-state relationships.

neuroscience↗

bioRxiv2026

Water as a thermal contrast agent for artificial-intelligence-enhanced in vivo mid-infrared thermography

In vivo infrared thermography is limited by the inherently poor spatial resolution at long wavelengths, low contrast, and the lack of biocompatible contrast agents. Here, we present 3-5 m mid-wave infrared (MWIR) thermography enhanced by an artificial intelligence (AI) network and cold phosphate-buffered saline (PBS) as a thermal contrast agent for noninvasive in vivo imaging with high contrast and resolution. MWIR imaging enabled high thermal sensitivity with microscale spatial resolution, strong relative thermal contrast, and facilitated visualization of the subcutaneous vasculature in the human arm, hand, ankle, the femoral artery and vein in rats, and the femoral vessels in mice, with image contrast further enhanced by AI networks. In a 4T1 tumor-bearing mouse model, AI-enhanced MWIR resolved early-stage tumors of [~]2.3 mm and metastases as small as [~]1.7 mm. Using cold PBS as a MWIR thermal contrast agent, we achieved precise tumor boundary visualization and real-time imaging-guided tumor resection. AI-enhanced MWIR offers a promising solution for early diagnosis and improved surgical precision.

bioengineering↗

bioRxiv2026

Near-Infrared II Scintillator for High-Resolution X-ray Imaging

Light scattering in scintillators is a pervasive problem and a key factor limiting X-ray imaging resolution. Here, we shift scintillator radioluminescence from the traditional visible range into the short-wave infrared (SWIR) or near-infrared II (NIR-II, 1000-3000 nm) window to mitigate light scattering and thereby enhance light penetration and X-ray imaging resolution. We present an NIR-II MgGa2O4:Ni2+ scintillator with peak emission at 1340 nm, achieving a threefold improvement in X-ray imaging resolution compared with visible scintillators owing to reduced light scattering. This heavy-metal-free NIR-II scintillator exhibits intense radioluminescence comparable to that of conventional visible-emitting CsI:Tl, achieving a detection limit of 56 nanograys per second, [~]100-fold lower than typical doses used in medical imaging. We show that this NIR-II scintillator enables high-resolution X-ray radiography of electronic circuit boards and biological tissues.

biophysics↗

bioRxiv2026

A CSF Proteomic Clock Reveals Opposing Brain-Aging Programs and Predicts Neurological Disease Progression

Aging is the risk factor for neurological disease, yet the molecular architecture of human brain aging remains poorly defined. Here, we analyzed more than 10,000 cerebrospinal fluid (CSF) proteomes across multiple cohorts and proteomic platforms to developed a 249-protein CSF aging clock that accurately predicts age and generalizes across independent datasets. CSF brain-age acceleration was increased across diverse neurological diseases, associated with blood-brain barrier (BBB) dysfunction, and predictive of longitudinal cognitive decline, neuroimaging progression, and dementia conversion. A simplified 30-protein panel retained similar prognostic performance. Biologically, clock proteins resolved two opposing programs: pro-aging activation of immune, vascular/BBB, ECM and coagulation pathways marked by CHI3L1, CD14, VWF, LRG1 and LTBP2, and anti-aging collapse of neuronal, synaptic, axonal and structural-maintenance programs marked by NPTX2, COL1A2, NID1, CDH8 and PENK. Brain-wide single-cell and regional mapping linked these programs to disease-vulnerable compartments. These findings establish a molecular framework for biological brain aging and neurological disease vulnerability.

neuroscience↗

bioRxiv2026

Near-future warming amplifies natural heatwave impacts and reorganizes freshwater communities

Future climate change may reshape ecological communities not only by increasing mean temperature, but also by altering the consequences of increasingly frequent heatwaves. Predicting these effects requires understanding how background warming interacts with short heatwaves in natural communities, where responses can arise through direct thermal stress and species interactions. We tested this using 32 outdoor freshwater mesocosms exposed to sustained near-future warming while capturing a documented natural heatwave. Warming raised temperature maxima that exceeded the thermal threshold of the pond snail, a main grazer in the community. Warmed communities showed lower grazer abundance, increased macrophyte and insect herbivore abundance, reduced phytoplankton biomass, and lower zooplankton density. Complementary assays showed that heatwave-level temperatures promoted macrophyte growth and reduced grazer survival, whereas reduced zooplankton performance mainly reflected indirect warming effects via food-web cascades. Thus, near-future warming can amplify natural heatwave impacts by exceeding consumer thermal thresholds and propagating through species interactions.

ecology↗

bioRxiv2026

Single-Cell Transcriptomic Analysis of the Immune Response to CHIKVInfection

BackgroundThe Chikungunya virus (CHIKV), a re-emerging mosquito-borne alphavirus, is responsible for acute febrile illness and severe polyarthralgia. Although both innate and adaptive immune responses influence the disease outcomes, the detailed cellular immunopathogenesis of CHIKV in peripheral blood is not yet fully elucidated. MethodsWe conducted single-cell RNA sequencing (scRNA-seq) on peripheral blood mononuclear cells (PBMCs) obtained from patients acutely infected with CHIKV and from healthy control subjects. Cellular interactions were inferred, and the transcriptomic results were orthogonally validated through quantitative real-time PCR (qPCR) and Enzyme-Linked Immunosorbent Assay (ELISA) to assess systemic interferon-stimulated responses. Furthermore, a comparative analysis was performed using publicly available single-cell data from Dengue virus (DENV) infections. ResultsCHIKV infection significantly altered the immune system, increasing monocytes and dendritic cells while reducing T and B lymphocytes. Monocytes and NK cells showed strong activation of interferon-stimulated genes (ISGs). Monocytes were identified as key in driving inflammatory and immune responses. In adaptive immunity, CHIKV led B cells to become plasmablasts with antiviral immunoglobulins and caused T cells and NK-like T cells to show signs of cytotoxicity and exhaustion. Validation showed increased levels of IFN-{gamma}, IFN-{beta}1, MX1, and ISG15. CHIKV triggered a more intense, monocyte-driven interferon response than DENV. ConclusionsAcute CHIKV infection induces a systemic interferon response predominantly centered on monocytes, accompanied by significant alterations in adaptive immunity. Circulating ISG products, including MX1 and ISG15, reflect the transcriptomic activation and may serve as potential biomarkers for assessing the early intensity of innate antiviral responses.

immunology↗

bioRxiv2026

Tirzepatide attenuates atherosclerosis through weight loss-independent anti-inflammatory mechanisms

BackgroundAtherosclerosis is a chronic inflammatory vascular disorder with persistent residual inflammation even after standard lipid-lowering therapy. Mounting evidence from bench to bedside suggests that diabetes and obesity accelerate atherosclerosis development. Tirzepatide (TZP), a dual Glucagon-Like Peptide-1 Receptor/Glucose-Dependent Insulinotropic Polypeptide Receptor (GLP-1R/GIPR) agonist approved for treating diabetes and obesity, has demonstrated proven cardiometabolic efficacy in large cardiovascular outcome trials. However, it remains largely uncertain whether TZP attenuates atherosclerosis independent of its anti-diabetic and anti-obese effects through direct actions on the vasculature. MethodsWe established atherosclerotic mouse models under diabetic, obese, and non-diabetic/non-obese conditions. Analysis of covariance (ANCOVA) and pair-feeding experiments were applied to experimentally decouple weight-dependent metabolic improvement from intrinsic vasculoprotection. Molecular and cell biological assays in human umbilical vein endothelial cells (HUVECs) and human aortic endothelial cells (HAECs) were performed to dissect the underlying signaling mechanisms. ResultsTZP markedly reduced aortic plaque burden and inflammation, restrained necrotic core enlargement, and improved plaque stability across all experimental mouse models. Both ANCOVA and pair-feeding experiments confirmed that these atheroprotective effects were independent of food intake and body weight loss. Furthermore, TZP attenuated systemic and vascular inflammation in Tumor Necrosis Factor- (TNF)-treated C57BL/6J mice, and this protection occurred without changes in body weight or blood glucose levels. Mechanistically, TZP directly targeted endothelial cells and activated the cyclic adenosine monophosphate (cAMP)/protein kinase A (PKA)/endothelial nitric oxide synthase (eNOS) pathway, increased eNOS phosphorylation and nitric oxide bioavailability, consequently downregulating the expression of the pro-inflammatory adhesion molecules Vascular Cell Adhesion Molecule-1 (VCAM-1) and Intercellular Adhesion Molecule-1 (ICAM-1). ConclusionsTZP arrests atherosclerosis progression through weight loss-independent anti-inflammatory mechanisms. These findings implicate TZP as a promising therapeutic drug for mitigating residual vascular inflammation in patients with atherosclerotic cardiovascular disease (ASCVD), irrespective of glycemic status or obesity. Clinical PerspectiveO_ST_ABSWhat Is New?C_ST_ABSO_LITirzepatide exerts direct anti-atherosclerotic effects in preclinical mouse models of atherosclerosis under diabetic, obese, and non-obese conditions. C_LIO_LITirzepatide directly targets endothelial GLP-1R/GIPR and downstream cAMP/PKA/eNOS signaling pathway to suppress NF-{kappa}B-driven vascular inflammation, thereby uncovering a previously unrecognized vasculoprotective mechanism underlying its cardiovascular benefits C_LI What Are the Clinical Implications?O_LITirzepatide exerts direct vascular protective effects independent of body weight reduction, suggesting that its cardiovascular benefits may extend beyond glycemic control and obesity management. C_LIO_LITirzepatide may represent a promising therapeutic drug for addressing residual vascular inflammation in ASCVD patients, including those without overt diabetes or obesity C_LI

physiology↗

bioRxiv2026

The Two-Component System CrdRS Mediates Sub-inhibitory Antibiotic-Induced Biofilm Formation in Helicobacter pylori via Bidirectional Regulation of Transporters PlpA and GlnP

Biofilm formation by Helicobacter pylori is a major driver of antibiotic tolerance and treatment failure, yet the signaling pathways that trigger biofilm development under sub-inhibitory antibiotic pressure remain poorly understood. Here, we show that sub-MIC levels of metronidazole, amoxicillin, and ciprofloxacin potently induce dense H. pylori biofilms. Through transcriptomic and genetic analyses, we identify the two-component system CrdRS as a central signaling hub that orchestrates this response via the bidirectional regulation of two ATP-binding cassette (ABC) transporters. Phosphorylated CrdR binds to AC-rich promoter motifs to directly activate plpA, which encodes a substrate-binding protein that drives exopolysaccharide secretion and matrix assembly. Concomitantly, CrdR represses glnP, which encodes an inner-membrane permease, thereby relieving transcriptional inhibition of the L-asparaginase gene ansB. This derepression triggers aberrant reactive oxygen species (ROS) accumulation, which promotes oxidative stress-dependent biofilm maturation. Through phenotypic analysis of crdRS deletion mutants, phosphorylation-defective point mutants (CrdRD53A and CrdSH173A), and exogenous hydrogen peroxide (H2O2) induction, we demonstrate that CrdRS specifically responds to antibiotic-induced stress signals in a manner genetically separable from ROS sensing. Collectively, our findings establish a dual-mechanism model in which CrdRS orchestrates antibiotic-induced biofilm formation by simultaneously controlling matrix production and intracellular ROS generation. Notably, glnP expression was significantly lower in clinical multidrug-resistant isolates than in drug-sensitive ones, whereas plpA showed the opposite trend. These findings provide a mechanistic foundation for developing CrdRS-targeted strategies to combat biofilm-associated H. pylori infections.

microbiology↗

bioRxiv2026

A GPX4 phosphorylation switch by FGFR1 guards against ferroptosis

Ferroptosis is driven by lipid peroxidation, yet the mechanisms by which cells rapidly adjust their sensitivity to ferroptosis in response to extracellular cues remain elusive. We identify a direct phosphorylation switch controlling the activity of glutathione peroxidase 4 (GPX4), the core ferroptosis regulator. The receptor tyrosine kinase FGFR1 directly binds and phosphorylates GPX4 at Tyr180/Tyr196 in a kinase-dependent manner, requiring its Tyr730 as a docking site. This phosphorylation enhances GPX4s catalytic activity and suppresses ferroptosis. In cardiac ischemia/reperfusion injury, the FGFR1-GPX4 axis is suppressed, and a selective FGFR1 agonist (FGF-1{Delta}NT) reactivates it to protect against ferroptosis-mediated damage. Critically, a phosphorylation-deficient GPX4 knock-in mouse exhibits hypersensitivity to injury and non-responsive to this agonist, proving that GPX4 phosphorylation is essential. Our findings reveal a rapid mechanism for regulating ferroptosis via GPX4 tyrosine phosphorylation, directly linking receptor tyrosine kinase signaling to ferroptosis, and offering new strategies for treating ischemic-and other ferroptosis-associated diseases.

Cell Biology↗

bioRxiv2026

A human intracranial map of consciousness returning from anesthesia

How consciousness arises from the brain remains a central yet unsolved question. Although substantial effort has been devoted to characterizing the neural correlates of conscious perception during wakefulness, how consciousness re-emerges from unconscious states remains poorly understood. Here, we took advantage of the rare opportunity to record human intracranial local field potentials from propofol-induced general anesthesia through the transition to behavioral engagement and subsequent full wakefulness, capturing both the stable states of unconsciousness and consciousness and the reconstituting neural dynamics that bridge them. Anesthesia cannot be defined by any single electrophysiological signature; rather, it is an organized low-frequency regime characterized by a constellation of coordinated neural phenomena, including aperiodic slow waves, alpha/beta periodicity, global alpha synchronization, and slow-wave-alpha/beta phase-amplitude coupling. Following anesthetic cessation, this regime progressively dissolved as neural excitability and dynamical complexity climbed. The appearance of conscious behavior coincided with a fast and dramatic transformation in high-gamma activity, from stochastic and unpredictable bursts to structured responses that were distributed, task-selective, and event-locked. These findings suggest that the recovery of consciousness is a multiscale reorganization in which distinct neural underpinnings rise and fall. Together, this work charts an electrophysiological map of how conscious cognition is extinguished, reconfigured, and restored in the human brain.

neuroscience↗

bioRxiv2026

Generative design of antigen-specific T-cell receptor sequences with a conditional diffusion model

T cell receptor (TCR)-based immunotherapy holds immense potential for treating cancers, autoimmunity, and infectious diseases, where antigen-specific TCR recognition is crucial for adaptive immune responses. Engineering or de novo generation of the complementarity-determining region 3 (CDR3) loops of TCRs using artificial intelligence offers a powerful alternative to designing antigen-specific TCRs rather than laborious experimental screening. However, current in silico approaches are constrained by weak conditional guidance, limited flexibility, and a lack of rigorous functional validation. To address these limitations, we introduce TCRDiff, a generative diffusion framework for designing antigen-specific TCRs conditioned on peptide-MHC (pMHC) targets and germline-encoded TCR variable genes. By leveraging pre-trained knowledge from massive T-cell repertoires and TCR-pMHC recognition data, TCRDiff generates CDR3{beta} sequences that closely resemble native-binding TCRs via a denoising diffusion process. Furthermore, incorporating interface geometry features generated TCR-pMHC complexes with superior structural plausibility than models relying solely on sequence-based diffusion or structure-based modeling. As a proof of concept, we deployed TCRDiff in a systematic pipeline to design candidate TCRs against a clinically validated cancer antigen. In vitro activation assays validated that TCRDiff-generated TCRs efficiently recognize the MAGE-A3 epitope with minimal off-target reactivity. Thus, TCRDiff establishes a powerful, validated computational paradigm to accelerate the development of TCR-based immunotherapies.

bioinformatics↗

bioRxiv2026

Desert hedgehog enhances endothelial resilience and prevents atherosclerosis by mitigating PAI-1 signaling

BACKGROUNDAtherosclerotic cardiovascular disease remains the leading cause of death worldwide. Most current pharmacotherapies target conventional risk factors that promote atherosclerosis (e.g., hyperlipidemia) rather than intrinsic resilience factors that protect against atherosclerosis in the face of risk factors. Here, we investigated the role of desert hedgehog (DHH), a canonical ligand of the hedgehog signaling pathway, as a novel resilience factor that restrains endothelial mesenchymal transition (EndoMT) and protects against atherosclerosis. METHODSSingle-cell RNA sequencing (scRNA-seq) was performed on atheroprone and atheroprotective regions of the ApoE knockout mouse aorta to identify mechanoresponsive genes associated with atherosclerosis. Endothelial cell-specific Dhh knockout mice were subjected to partial carotid ligation and hypercholesterolemic conditions to investigate the role of endothelial Dhh in atherosclerosis progression. scRNA-seq, bulk RNA sequencing, endothelial lineage tracing, immunoprecipitation-coupled mass spectrometry, and surface plasmon resonance were used to in-vestigate the role and mechanism of DHH in EndoMT. Pharmacological interventions and recombinant DHH administration were performed in vivo to evaluate therapeutic potential of DHH targeting. DHH expression was also examined in human atherosclerotic arteries and serum samples from patients with coronary artery disease. RESULTSDHH protein expression was enriched in arterial endothelium from mice, porcine, and humans. However, DHH expression was significantly reduced in atherosclerotic arteries and serum from patients with coronary artery disease. scRNA-seq of atheroprone and atheroresistant region of mouse aorta identified Dhh as a novel mechanoresponsive gene enriched in aortic regions exposed to unidirectional laminar flow. Endothelial cell-specific Dhh knockout (DhhecKO) mice exhibited increased atherosclerotic lesion area, large necrotic cores, and reduced collagen content following partial carotid ligation. Similarly, under hypercholesterolemic conditions, both male and female DhhecKO mice showed aggravated atherosclerosis progression. scRNA-seq of DhhecKO mouse aortas revealed an increased proportion of endothelial cells undergoing mesenchymal transition, indicating enhanced EndoMT. These findings were corroborated by bulk RNA-sequencing of DHH depleted HUVECs and endothelial lineage tracing in inducible DhhecKO mice. Mechanistically, DHH directly interacted with plasminogen activator inhibitor type 1 (PAI-1) and suppressed PAI-1-induced EndoMT. PAI-1 promoted EndoMT in ECs through activation of canonical TGF-{beta} signaling (SMAD2/3) and noncanonical AKT/ERK1/2 signaling via interaction with low-density lipoprotein receptor-related protein (LRP1). Neutralization of PAI-1 or inhibition of LRP1, AKT/ERK1/2, or SMAD3 signaling abolished DHH deficiency-induced EndoMT. DHH competitively inhibited PAI-1 binding to LRP1, thereby attenuating downstream pro-EndoMT signaling. Intriguingly, treatment with PAI-1 inhibitor TM5275 mitigated endothelial Dhh deficiency induced EndoMT in vivo. Of translational relevance, recombinant mouse DHH protein administration reduced atherosclerosis progression, stabilized plaque, and decreased the expression of EndoMT markers in ApoE knockout mice. CONCLUSIONSDesert hedgehog (DHH) is an intrinsic endothelial cell-enriched resilience factor that protects against EndoMT and atherosclerosis by preventing PAI-1 signaling. The present study implicates endothelial DHH as a potential therapeutic target for atherosclerotic cardiovascular disease. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=163 SRC="FIGDIR/small/729715v1_ufig1.gif" ALT="Figure 1"> View larger version (50K): org.highwire.dtl.DTLVardef@194c8a3org.highwire.dtl.DTLVardef@1004d52org.highwire.dtl.DTLVardef@d47c73org.highwire.dtl.DTLVardef@368866_HPS_FORMAT_FIGEXP M_FIG C_FIG Desert hedgehog (DHH) is an intrinsic endothelial cell-enriched resilience factor that protects against EndoMT and atherosclerosis by preventing PAI-1 binding to LRP1 and downstream AKT/ERK, as well as SMAD2/3 signaling. Clinical PerspectiveO_ST_ABSWhat Is New?C_ST_ABSO_LIDHH was identified as a flow-responsive resilience gene that is downregulated by disturbed flow in endothelial cells. C_LIO_LIEndothelial-specific deletion of Dhh promoted endothelial-to-mesenchymal transition (En-doMT) and atherosclerosis. C_LIO_LIDHH directly interacts with PAI-1 and preclude PAI-1 mediated pro-EndoMT signaling. C_LI What Are the Clinical Implications?O_LIDHH maintains endothelial homeostasis during atherosclerosis. C_LIO_LITargeting endothelial DHH-PAI-1 axis may represent a potential therapeutic strategy to reduce EndoMT and limit plaque progression. C_LIO_LILower circulating DHH level may serve as a potential biomarker of endothelial dysfunction and plaque vulnerability in atherosclerotic disease. C_LI

biochemistry↗

bioRxiv2026

Efficient base editing and development in human embryos without chromosomal alterations

Cas9-based tools enable the introduction of genetic lesions to investigate DNA repair outcomes and edit the genome at disease-relevant loci. DNA double-strand breaks (DSBs) induced by CRISPR/Cas9 result in frequent aneuploidy and large deletions, revealing a repair deficiency in early human embryos and limiting the clinical application of this technology. Here we evaluated the DNA repair outcomes of DNA nicks and mismatches introduced using base editors in human embryos at two targets, PCSK9 and HBG. Editing was efficient and, unlike Cas9-induced DSBs, did not result in either chromosomal abnormalities or large deletions. Small insertions or deletions after base editing were rare, and off-target activity was dependent on the guide RNA. Delivering the base editor as a protein at fertilization or at the pronuclear stage allowed normal development to the blastocyst stage and the derivation of edited stem cell lines. In stark contrast, introduction of the editor as RNA resulted in early embryo arrest. Our results demonstrated that, unlike DSBs, DNA nicks and mismatches are efficiently repaired in human embryos, allowing specific on-target changes without genotoxic consequences.

genetics↗

bioRxiv2026

Inhibition of hexokinase 2 undermines cartilage health and accelerates osteoarthritis

ObjectiveEnhanced glycolysis is a metabolic hallmark of chondrocytes in osteoarthritis (OA); however, the roles of the glycolytic rate-limiting enzyme hexokinase 2 (HK2) in cartilage remain poorly understood. MethodsPharmacological approach (3-bromopyruvate (3-BrPA) treatment) and mice model involving HK2 knockout in Col2a1-expressing chondrocytes are utilized to access the impact of HK2 blockage on cartilage ex vivo and in vivo. The in vivo effects of HK2 inhibition on OA progression were evaluated using a destabilization of the medial meniscus (DMM)-induced OA mouse model, through both intra-articular 3-BrPA administration and chondrocyte HK2 deletion. Additionally, we analyzed published single-cell RNA sequencing (scRNA-seq) datasets from human articular cartilage and integrated these with bulk RNA-seq data from HK2-deficient chondrocytes to characterize HK2 expression features across conditions. ResultsBoth pharmacological inhibition and genetic deletion of HK2 impair cartilage formation ex vivo. Bulk RNA-seq analysis and ex vivo studies demonstrated a promoted ossification-like process due to HK2 ablation in chondrocytes. Through pseudotime analysis of published single-cell RNA sequencing (scRNA-seq) datasets from human articular cartilages, we further identified that HK2 is differentially expressed across conditions, with a feature of a relatively high expression level at terminal stages of chondrocyte differentiation in the context of OA. We next confirmed HK2 deficiency in chondrocytes significantly exacerbated OA progression but having no impact on skeletal development in mice. ConclusionsHK2 plays a critical role in maintaining cartilage health, likely through the regulation of calcification, thereby highlighting the potential risks associated with targeting glycolytic enzymes as a therapeutic strategy for OA.

pathology↗

bioRxiv2026

Objective curriculum-guided design of multi-property proteins

Designing functional proteins that simultaneously possess multiple bio-chemical properties remains a significant challenge, as key protein properties, such as solubility, stability, binding affinity, and chemical resistance, are often interdependent or even conflicting. Current approaches typically attempt to jointly optimize multiple functional objectives in one shot, followed by extensive screening to identify rare feasible designs. Here, we introduce OCDesign, an objective curriculum-guided framework for multi-property protein design. OCDesign is based on the objective curriculum principle, i.e., the order in which objectives are introduced can shape the accessibility of functional solutions. In each design round of OCDesign, candidate sequences are generated in silico, assessed across multiple properties, selected based on Pareto-optimal trade-offs, and experimentally validated, with each experimental stage testing the role of the newly introduced objective within the curriculum. Using antibody-binding protein A as a model system, we show that one-shot optimization fails to yield functional designs, whereas a staged curriculum--progressing from solubility and structural consistency to binding affinity, and then to alkaline resistance--enables the design of proteins possessing multiple desired properties through substantially fewer wet-lab experiments. These results establish OCDesign as a practical computational-experimental strategy for organizing and integrating multiple objectives in protein design, and suggest that objective ordering is a key determinant of accessibility in high-dimensional design spaces.

bioengineering↗