Biology subjects

Wu, X.

Publications and source records attributed to Wu, X..

At least 163 records · Page 9Linked to original sources

bioRxiv2025

Glycemia Shift Pancreatic Islets Rhythmicity via δ-α Cell in vivo, Impairment in Diabetes

Blood glucose homeostasis relies on the well-coordinated rhythmic activity of millions of islets throughout the pancreas. Islet rhythmicity is triggered by glucose elevation and mediated by paracrine interactions. However, the dynamics of islet population rhythmicity in healthy and diabetic pancreases in vivo remain poorly understood. Using simultaneous multi-islet Ca2+ imaging (20-100 islets per experiment) in both live mice and pancreatic tissue slices, we systematically studied how glycemia fluctuations and intra-islet paracrine signaling collectively shape the islet rhythmicity. In this study, we report that a transition from Hyperglycemia to Euglycemia induces a coordinated shift from slow to fast islet Ca2+ oscillations (HESF) in vivo. HESF is conserved in pancreatic tissue slices and isolated islets, however, not dispersed single cells in vitro, suggesting a mechanistic link with paracrine interactions. We found HESF arises from -cell activation, which is inhibited by {delta} cells upon glucose elevation. The autonomous islets mostly differ in phase and period at high glucose level. Diabetic mice with disrupted glycemic stability lost HESF both in vivo and in vitro. Interestingly, HESF is preserved in {beta}-cell knockout Gcgr transgenetic mice, both in vivo and in vitro, suggesting HESFs dependence on Glp1r. Indeed, HESF was restored in semaglutide-treated diabetic mice with stabilized glycemic stability. These findings offer a comprehensive understanding of how {delta} and cells influence islet rhythmicity and precisely maintain the stability of blood glucose.

physiology↗

bioRxiv2025

Beyond RECIST: mathematical modeling and Bayesian inference reveal the importance of immune parameters in metastatic breast cancer

Successful immunotherapies must overcome patient- and organ-specific tumor heterogeneities to mount an effective response. Yet tumor dynamics remain poorly characterized in organ-specific contexts and associated immune environments. To quantify heterogeneous tumor responses, we developed methods to fit mathematical models of the tumor-immune dynamics to patients undergoing combination therapy for metastatic breast cancer: checkpoint inhibition via nivolumab + ipilimumab combined with entinostat, as measured by RECIST criteria. In a subset of patients additional immune dynamics were quantified by spatial proteomics. Bayesian parameter inference revealed that only immune-modulatory parameters controlled response; parameters controlling cytotoxicity were uninformative. Through posterior parameter sampling and simulation, we created virtual tumor cohorts, enabling extrapolation beyond the data to predict probabilities of response in metastatic lesions where no data exist. We validated pre-dictions from our virtual tumor population using held-out data characterizing off-target lesions from the patient cohort. Profile-wise likelihood analysis revealed that scans in the week immediately following treatment hold particularly high value in identifying the tumor dynamics. Overall, we demonstrate how through modeling & inference cohort size limitations can be over-come through the creation of virtual tumor populations, giving insight into the site-specific mechanisms of disease progression and response.

systems biology↗

bioRxiv2025

Signatures of adaptive self-organization detected in swidden societies worldwide

Quantitative signatures of adaptive self-organization and the spatial scale of human management can be detected in anthropogenic landscapes. Yet, while abundant evidence for self-organization in physical and natural systems exists, there are few adaptive self-organization models that can explain coupled dynamics of human subsistence systems that have also been rigorously validated. Here, we analyze over 18,000 forest patches extracted from 18 remotely-sensed swidden mosaics from tropical and subtropical regions across the planet. We discover a power law signature of adaptive self-organization in the size of the patches and correlation distances that indicate the spatial scale of socioecological adaptation. To explain this pattern, we propose a mathematical model of swidden labor reciprocity, normative reasoning, field placement, forest disturbance and regrowth dynamics that explains the emergent spatial structure of swidden mosaics. Social self-organization can promote sustainable levels of forest disturbance when ecological dynamics such as facilitation, seed-dispersal and competition are conditioned by social dynamics, giving rise to power-law land-use patterns. We use simulations to explore the emergent properties of the model and identify two regimes: deforestation and sustainable swidden. When we analyze the relationship between the simulation results in the remote sensing data, we find that 16/18 of the study areas exhibit power-law distributions and spatial correlation patterns that match the sustainable swidden regime in the model. These findings support the hypothesis that adaptive self-organization may be a general characteristic of coupled human and natural systems.

ecology↗

bioRxiv2025

Hi-Cformer enables multi-scale chromatin contact map modeling for single-cell Hi-C data analysis

Single-cell Hi-C captures the three-dimensional organization of chromatin in individual cells and provides insights into fundamental genomic processes such as gene regulation and transcription. While analyses of bulk Hi-C data have revealed multi-scale chromatin structures like A/B compartments and topologically associating domains, single-cell Hi-C data remain challenging to analyze due to sparsity and uneven distribution of chromatin contacts across genomic distances. These characteristics lead to strong signals near the diagonal and complex multi-scale local patterns in single-cell contact maps. Here, we propose Hi-Cformer, a transformer-based method that simultaneously models multi-scale blocks of chromatin contact maps and incorporates a specially designed attention mechanism to capture the dependencies between chromatin interactions across genomic regions and scales, enabling the integration of both global and fine-grained chromatin interaction features. Building on this architecture, Hi-Cformer robustly derives low-dimensional representations of cells from single-cell Hi-C data, achieving clearer separation of cell types compared to existing methods. Hi-Cformer can also accurately impute chromatin interaction signals associated with cellular heterogeneity, including 3D genome features such as topologically associating domain-like boundaries and A/B compartments. Furthermore, by leveraging its learned embeddings, Hi-Cformer can be extended to cell type annotation, achieving high accuracy and robustness across both intra- and inter-dataset scenarios.

bioinformatics↗

bioRxiv2025

Low-frequency brain oscillations reflect the dynamics of the oculomotor system: a new perspective on subsequent memory effects

Neural activity and eye movements are two well-established predictors of memory performance in humans. Successful memory formation is typically associated with reduced alpha/beta power (i.e., the alpha/beta subsequent memory effect) and an increased number of eye movements. However, the functional relevance of these two memory correlates has primarily been investigated in isolation, leaving their coupling and combined contribution to memory formation largely unknown. Here, we address this gap through four experiments involving simultaneous eye-tracking and scalp or intracranial electroencephalography recordings while participants viewed scenes either freely or under varying levels of visual constraint. Across experiments and cohorts, we identified the degree of visual exploration, rather than subsequent memory performance, as the consistent and robust modulator of alpha/beta power reductions. Going beyond correlations, manipulating eye movements dissociated alpha/beta power reductions from memory. Combined with the finding that saccade parameters predicted the dynamics of alpha/beta activity, our results directly link alpha/beta activity to the oculomotor system. Moreover, we show that the accumulation of saccades over time accounted for the reduced alpha/beta power, resembling the pattern typically observed after stimulus onset. Our work demonstrates that neglecting eye movements results in an incomplete understanding of the alpha/beta subsequent memory effect. Together, the evidence points at alpha/beta activity directly reflecting eye movements, and only indirectly, memory formation. The current study thus bridges two lines of previous research and advocates for new perspectives on the specific interpretation of the alpha/beta subsequent memory effect and on the function of alpha/beta activity in general.

neuroscience↗

bioRxiv2025

Construction, Phenotypic Characterization, and Immunomodulatory Function Study of BMSC-Macrophagy Hybrid in vitro

ObjectiveThe aim of our study was to integrate bone marrow-derived stem cells (BMSCs) with macrophages and to investigate the performance of this hybrid in IgG clearance, immunosuppression, and the mitigation of inflammatory injury. MethodsBMSCs and RAW264.7 cells were fused through transient transfection using the COVID-19 spike glycoprotein and human angiotensin-converting enzyme 2 (hACE2), respectively. The resulting hybrids were purified using magnetic sorting, and their characteristics were evaluated by examining morphology, glucose uptake rates, phagocytic capacity, cytoskeletal morphology, and the levels of Fc{gamma} receptors, as well as anti-inflammatory, antioxidant, adhesion, and complement inhibitory factors. The IgG clearance capability of the hybrids was assessed by measuring internalization efficiency and infiltration into renal organoids. Additionally, the reparative effects of the hybrids were evaluated using doxorubicin-treated MPC5 damage models. Live cell imaging techniques were employed to investigate the interactions between the hybrids and immature dendritic cells (iDCs). Hybrid-derived nanovesicles were isolated, and their abilities to target IgG and clear IL-6 were evaluated using immunofluorescence and ELISA. Furthermore, chloroplasts and C-dots were co-incubated with the hybrids, and their benefits were assessed by measuring cell viability, oxygen content, ROS levels, mitochondrial function, and the levels of anti-inflammatory, antioxidant, and pro-repair factors. A scratch assay was conducted to determine the impact of chloroplast transplantation on the migration ability of 3T3 cells. ResultsHigh-purity hybrids were successfully produced that cleared IgG while maintaining their anti-inflammatory and antioxidant effects. They promoted the recovery of podocyte injury through vesicular and mitochondrial delivery, as well as efferocytosis. Furthermore, the hybrids inhibited iDCs primarily through migrasomes, intercellular nanomicrotubule connections, and phagocytosis. The derived nanovesicles were capable of residing in IgG-enriched regions and adsorbing IL-6. Interestingly, the transplanted chloroplasts enabled the hybrids to utilize light energy to enhance their antioxidant capacity and promote the migration of 3T3 cells, which contributes to wound repair. Subsequently, the loading of C-dots was beneficial for enhancing resistance to oxidative damage. ConclusionsOur results suggest that hybrids-mediated therapy is a new and creative therapeutic approach for managing immune-mediated nephropathies.Hybrids demonstrated effective immunomodulatory and promoted injury recovery. The derived nanovesicles have the potential to alleviate the inflammatory burden at sites of antibody immune complex deposition. Additionally, the incorporation of chloroplasts and C-dots enhanced the hybrids adaptability to ischemic-hypoxic microenvironments.

cell biology↗

bioRxiv2025

Identification of cellular and molecular risk signatures for progression to late-stage age-related macular degeneration using the 9-step Minnesota Grading System

Age-related macular degeneration (AMD) is a complex multifactorial disease, and the molecular mechanisms underpinning the progression of intermediate AMD to geographic atrophy are not fully understood. To better understand mechanisms driving progression, we performed bulk RNA sequencing on dissected macular and peripheral RPE/choroid and neural retina tissue from postmortem human eyes graded using the 9-step Minnesota Grading System (MGS). Binning of intermediate AMD cases into three distinct groups (AMD3L, AMD3M, AMD3H) based on the 5-year risk of progression enabled identification of distinct gene and pathway changes associated with progression to late-stage disease. Identified changes in gene expression were validated using ELISA or histological methods. RPE-specific genes and lipid metabolic pathways showed a transient increase in AMD3L followed by a pronounced decrease in AMD3H. In AMD3H, immune response genes such as C3, TREM2, and OLR1 were upregulated when compared to AMD3L samples, as well as genes specific to Muller glia/astrocytes (NGFR, SPP1, GPX3). Our findings support complement inhibition as a promising therapeutic option for slowing conversion to advanced AMD and identify macrophage and Muller/astrocyte genes as potential cell types to target in AMD. Further, we demonstrate the value of combining emerging, outcomes-based, clinically relevant grading systems with profiling technologies to generate new insights into ocular diseases. HighlightsO_LIIntermediate AMD (AMD3) can be further divided into 3 stages - AMD3L (low risk), AMD3M (intermediate risk), AMD3H (high risk) - using the MGS9 grading system based on the risk of disease progression to late AMD. C_LIO_LIRNA sequencing of the macular RPE/choroid shows opposing changes in gene expression of multiple biological pathways for both RPE and immune cells between AMD3L and AMD3H stages. C_LIO_LIIn the macular neural retina, most biological pathways were downregulated in AMD2 (early AMD) but upregulated in AMD4 (late AMD) compared to AMD1 (non-AMD control). C_LIO_LIMolecular and cellular signatures associated with a high risk of progression to AMD4 include activation of complement C3, two subtypes of macrophages expressing either TREM2 or OLR1, and Muller glia/astrocytes as evidenced by the upregulation of GFAP and NGFR. C_LIO_LIUnderstanding the roles of these high-risk associated genes in AMD progression will facilitate the development of new treatments that prevent or delay the irreversible central vision loss in AMD patients. C_LI

neuroscience↗

bioRxiv2025

A Paracrine-to-Autocrine Shunt of GREM1 Fuels Colorectal Cancer Metastasis via ACVR1C

Tumor cells typically rely on paracrine stromal signals to guide malignant behaviors, but whether they acquire signaling autonomy to support metastasis remains unclear. We elucidate this in colorectal cancer (CRC) by uncovering a paracrine-to-autocrine shunt of Gremlin1 (GREM1), a canonical stromal-secreted antagonist of bone morphogenetic proteins (BMPs). We demonstrate that while GREM1 remains restricted to stromal cells in earlier-stage (I-III) CRC, its ectopic expression in tumor epithelium increases markedly in stage IV. Mechanistically, we identify ACVR1C as a novel, high-affinity epithelial receptor for GREM1. Their interaction activates SMAD2/3 signaling, which upregulates SNAI1 and GREM1, establishing a feedback loop that amplifies epithelial-mesenchymal transition (EMT). Disrupting this loop impairs CRC metastasis in vivo. Clinically, epithelial GREM1 or ACVR1C expression predicts metastasis and poor survival. These findings define a paradigm in which tumor cells hijack stromal GREM1 to establish a GREM1-ACVR1C autocrine loop that sustains EMT and metastasis, marking a shift toward signaling autonomy and revealing a targetable vulnerability in advanced CRC.

cancer biology↗

bioRxiv2025

Methyl gallate Attenuates Post-Stroke Emotional and Cognitive Symptoms by Promoting Hippocampal Neurogenesis via PI3K/GSK3 and AMPK Signaling

Restoring hippocampal neurogenesis is an effective strategy for post-stroke recovery. Methyl gallate (MET) exhibits neuroprotective properties. However, the effect of MET in improving brain functional recovery in the post-stroke depression (PSD) model and its underlying mechanism remains unknown. Single-cell data analysis showed that the cell types and molecular characteristics of PSD are similar to those of primary depression but exhibit weaker synaptic plasticity and stronger inflammatory signals. In addition, molecular docking studies revealed that MET exhibits a significant binding capacity with AMPK/GSK3, suggesting that MET mediates the neuroprotective effects of both. In this study, we created a post-stroke depression (PSD) model by performing physical restraint after ischemia and tested the treatment effects of MET. We observed that MET significantly attenuated PSD-induced depressive-/anxiety-behaviors associated with a reduction of stress hormone corticosterone and ACTH levels. Morris water maze and recognition task results indicate that MET can also alleviate cognitive impairments in the PSD model. In the hippocampus of the PSD model, MET improved the proliferation and differentiation of neural stem/progenitor cells (NSPCs). MET treatment significantly enhanced the activity of AMPK and decreased the activity of GSK3{beta}. Furthermore, in primary neural progenitors under hypoxia, both the PI3K inhibitor LY294002 and the AMPK inhibitor compound C blocked the effects of MET to promote neural development. Animal experiments also confirmed that LY294002/compound C treatment could reduce the effects of MET in antidepressant behaviours. Taken together, our results indicate that PI3K, as well as AMPK-mediated adult neurogenesis, is restored by MET to improve brain functions in the PSD model.

neuroscience↗

bioRxiv2025

Measure Sodium Transport in Cells with NMR

Sodium ions (Na+) are fundamental to numerous physiological functions, such as maintaining electrolyte balance, enabling nerve impulse transmission, and facilitating muscle contraction. Dysregulation of Na+ transport across cell membranes is implicated in a range of health issues, including metabolic syndromes, neurological conditions, and cardiovascular diseases. However, current methods for assessing cellular Na+ activity often face limitations; they can be invasive or fail to capture dynamic changes. In this study, we introduce a non-invasive 23Na nuclear magnetic resonance (NMR) methodology designed to directly quantify the transport rate of sodium ions in living cells. Our technique integrates relaxation exchange spectroscopy (REXSY) with a multi-site exchange model, enabling the investigation of Na+ transport dynamics on a timescale of sub-seconds. A key advantage is its ability to differentiate between intracellular and extracellular Na+ pools based on the endogenous NMR relaxation difference, thereby avoiding the need for potentially disruptive exogenous reagents. Experiments conducted on human cell lines successfully demonstrated the techniques capacity to distinguish between various physiological states, such as when ion channels are pharmacologically blocked or activated. The resulting measurements of Na+ transport rates and intracellular Na+ fractions show a clear correlation with cellular metabolic activity, offering valuable quantitative markers for monitoring transmembrane ion dynamics in vitro.

cell biology↗

bioRxiv2025

Stage- and tissue-specific gene editing using 4-OHT inducible Cas9 in whole organism

Vertebrate genes usually function in specific tissues at specific stages, and thus their functional studies require conditional knockout or editing. In the zebrafish, spatiotemporally inducible genome editing, especially during early embryonic development, is still challenging. Here, we report inducible Cas9-based gene editing in specific cell types at desired stages in zebrafish. We show that the nCas9ERT2 fusion protein consisting of Cas9 and estrogen receptor flanked by two nuclear localization signals is normally located in the cytoplasm and efficiently translocated into the nucleus upon 4-hydroxytamoxifen (4-OHT) treatment in cultured cells or embryos. As a proof-of-concept, we demonstrate that target genes in primordial germ cells or germ cells of transgenic zebrafish embryos or adults with stable expression of nCas9ERT2 and gene-specific guide RNAs (gRNAs) can be induced to mutate by application of 4-OHT. This inducible nCas9ERT2 system also works in mouse early embryos. Thus, this inducible gene editing approach will be another choice for studying the temporospatial function of genes at the organismal level. Hence, the inducible nCas9ERT2 system expands the tissue- and stage-specific gene editing toolkit.

developmental biology↗

bioRxiv2025

ODFormer: a Virtual Organoid for Predicting Personalized Therapeutic Responses in Pancreatic Cancer

Pancreatic cancer (PC) patient-derived organoids (PDOs) faithfully recapitulate therapeutic responses but face clinical translation barriers, including high costs and technical complexity. To address these problems and the lack of frameworks for PDO-based drug-response assays, we developed ODFormer, a computational framework that simulates PC PDOs to predict clinically actionable, patient-specific drug responses by integrating transcriptomic and mutational profiles. ODFormer first employed two encoders, pretrained on 30,000 pan-cancer bulk transcriptomics and 1 million PC single-cell profiles respectively, to distil tissue-and organoid-specific representations. Then, trained on our curated 14,000 PDO drug-response assay (across 183 PDOs and 98 drugs) using a transformer-augmented hybrid contrastive network, ODFormer significantly outperformed state-of-the-art methods, notably achieving a PCC >0.9 in predicting standardized drug response. Multi-cohort retrospective analyses further demonstrated that ODFormer-guided personalized therapy significantly improves clinical outcomes, without requiring physical organoid assays. Furthermore, ODFormer identified novel clinico-biological PC subtypes and revealed therapy resistance biomarkers by stratifying predicted responders and non-responders. These were validated using independent datasets including TCGA-PDAC. Notably, ODFormer-guided treatment efficacy showed high concordance with prospective clinical responses by CA19-9.

bioinformatics↗

bioRxiv2025

CRISPR-Based Gene Dependency Screens reveal Mechanism of BRAF Inhibitor Resistance in Anaplastic Thyroid Cancer

Anaplastic thyroid cancer (ATC) is the most aggressive form of thyroid cancer. Despite recent advances in treating BRAFV600E-driven ATC, therapy resistance remains a significant challenge, often resulting in disease progression and death. Leveraging a focused CRISPR/KO screen in parallel with a CRISPR/activation screen, both tailored on response to BRAFV600E inhibitor treatment, we identified TAZ (encoded by the WWTR1 gene) deficiency as synthetically lethal with BRAF inhibitor in ATC. TAZ is overexpressed in ATC compared to well-differentiated thyroid tumors. We demonstrate that TAZ-deficient ATC cells display heightened sensitivity to BRAF inhibitors both in vitro and in vivo. Using gene essentiality score across a large panel of cancer cell lines, we found that BRAFV600E-driven cancers are highly sensitive to TAZ loss, unlike their counterparts with wild-type BRAF and non-BRAFV600E. Mechanistically, we demonstrate that dabrafenib triggers the Unfolded Protein Response (UPR) under ER stress and suppresses protein synthesis. TAZ loss represses the UPR, reverses the inhibition of protein synthesis, and triggers increased cell death by ferroptosis in dabrafenib-treated ATC. Collectively, our findings unveil TAZ as a new target to overcome resistance to BRAF inhibitors in undifferentiated thyroid cancer.

cancer biology↗

bioRxiv2025

Chromosomal-level genome of the Pinus massoniana 'Minlin WY36'

Owing to conifers complex and large genomes, it is challenging to construct a comprehensive reference genome and conduct evolutionary and genomic research. Here, we present the chromosome-level assembly of Pinus massoniana Minlin WY36, a pioneering conifer species used for afforestation, timber production, and oleoresin extraction. The genome size was determined to be 23.41 Gb, with a contig N50 value of 1.15 Mb and a scaffold N50 value of 2.02 Gb. A total of 23.16 Gb of reads (98.93% of the assembled genome) were anchored and oriented onto 12 pseudochromosomes, with chromosome lengths from 1.47 to 2.29 Gb. A total of 32,144 protein-coding genes were predicted, of which 27,173 (84.54%) were functionally annotated. The P. massoniana Minlin WY36 genome consists of 79.41% (18.57 Gb) repetitive sequences, among which long terminal repeat sequences (LTR) were the most abundant. Through comparative genomic analysis, we found that the unique and significantly expanded genes in P. massoniana Minlin WY36 were enriched in the metabolism related to resin synthesis. This study offers valuable insights into conifer evolution and contributes resources for further investigations of conifer adaptation and developmental processes.

genomics↗

bioRxiv2025

ERM proteins regulate the shape and number of Endoplasmic Reticulum-Plasma Membrane Junctions in neurons

Endoplasmic Reticulum (ER) - Plasma Membrane (PM) Junctions (EPJs) are specialized contact sites between ER membrane and the inner leaflet of PM. These junctions are critical for lipid exchange and Ca2+ signaling. In excitable cells like neurons and muscle, EPJs further modulate membrane excitability by regulating Ca2+ homeostasis. The mechanisms controlling EPJ abundance and morphology remain poorly understood. Using in vivo fluorescence imaging and electron microscopy of C. elegans neurons, we showed that EPJs form discrete, patch-like structures distributed across the soma. Through a forward genetics screen, we identified two conserved ERM (Ezrin-Radixin-Moesin) proteins, FRM-4 and FRM-1, as key regulators of EPJ shape and abundance. Both proteins localize to EPJs and exhibit liquid-liquid phase separation properties (LLPS). in vitro, purified FRM-4 binds to FRM-1, and together bundle filamentous actin. However, their presence in LLPS condensates and actin-bundling activity are mutually exclusive. In vivo, F-actin cables surround--but do not penetrate--EPJs, where FRM proteins are enriched as phase-separated condensates. Loss of FRM-4, FRM-1, or disruption of F-actin led to increased mobility of EPJs that fused into fewer but enlarged junctions. Together, our findings demonstrate that FRM-4 and FRM-1 control EPJ morphology by organizing peri-junctional F-actin networks, thereby restricting EPJ mobility and fusion.

cell biology↗

bioRxiv2025

Super-giga and tiny orchid genomes illuminate evolution of Orchidaceae

Orchidaceae (orchids) is commonly known as one of the largest families of seed plants, and grow in an extensive range of habitats worldwide. In the present study, we generated chromosome-level reference genomes for two orchids using a combination of PacBio, Illumina, and Hi-C sequencing, Cypripedium singchii has the largest genome and chromosomes among the sequenced species so far, with a genome size of 43.19 Gb (1C) with ten chromosomes, and Apostasia fujianica has the smallest known genome and chromosomes in Orchidaceae, with a genome size of 340.90 Mb (1C) with 35 chromosomes. We predicted a total of 32,412 and 21,724 protein-coding genes for C. singchii and A. fujianica, respectively. The overall BUSCO score was 85.01% for C. singchii and 91.80% in A. fujianica. Based on protein-coding sequences from 55 conserved single-copy families across 21 plant species, we constructed a high-confidence phylogenetic tree and estimated the divergence times. The high-quality genomes of super-giga and tiny orchids offer key insight for future evolutionary researches.

genomics↗

bioRxiv2025

Transient AMPK activation by nutrient stress of high fat diet preserves cardiac electrophysiological stability and protects against arrhythmias

Sudden cardiac death (SCD) is a major complication of obesity, yet it remains unclear whether early metabolic stress, prior to the onset of overt obesity or structural remodeling, can independently promote arrhythmias. In vitro studies suggest that fatty acids can allosterically stimulate AMP-activated protein kinase (AMPK), a key metabolic sensor known to preserve myocardial viability and mitochondrial function following ischemia-reperfusion (I/R) injury. We hypothesized that AMPK signaling critically modulates the electrophysiological (EP) response to high-fat diet (HFD)-induced metabolic stress. MethodsTo test this, wild-type (WT) and AMPK kinase-dead (AMPK-KD) mice were subjected to an 8-week HFD regimen beginning at 4 weeks of age. Controls remained on normal diet (ND) for the same duration. Arrhythmia susceptibility was assessed ex vivo using rapid pacing and I/R challenge protocols. Changes in the EP substrate were defined by high-resolution optical action potential mapping. Underlying mechanisms were probed using western blotting, confocal and transmission electron microscopy. ResultsHFD-fed wild-type (WT) hearts did not display increased arrhythmia susceptibility in response to either burst pacing or I/R challenge. On the contrary, they exhibited a paradoxical enhancement in post-ischemic EP recovery compared to ND-fed controls. This improvement was associated with increased phosphorylation of canonical AMPK targets, including acetyl-CoA carboxylase (ACC) and raptor, consistent with the activation of a cardioprotective metabolic program. In sharp contrast, AMPK-deficient (AMPK-KD) hearts demonstrated heightened vulnerability to inducible ventricular tachycardia (VT), irrespective of diet. Conduction slowing emerged as an early EP abnormality in these hearts and served as the initial substrate (or first hit) that promoted their increased incidence of non-sustained VT. Notably, this conduction impairment arose in conjunction with an increase (rather than decrease) in Cx43 and Nav1.5 protein expression. Mechanistically, defective conduction in AMPK-KD hearts was linked to impaired autophagic degradation of intercalated disc proteins resulting from reduced phosphorylation of ULK1, a downstream effector of AMPK. Consequently, unphosphorylated Cx43 accumulated at the intercalated disc, likely replacing phosphorylated isoforms (p-Cx43). In addition, AMPK-KD hearts exhibited swollen, fragmented mitochondria and reduced levels of mitochondrial fusion proteins. Upon HFD challenge, this vulnerable mitochondrial substrate generated excessive reactive oxygen species (ROS) coinciding with accelerated repolarization. Together, impaired conduction and action potential shortening promoted VT sustenance in HFD-fed AMPK-deficient hearts. ConclusionsOur findings identify AMPK as a key metabolic regulator that integrates redox balance, mitochondrial integrity, and protein homeostasis to preserve cardiac excitability during early nutrient overload. Loss of AMPK signaling, as occurs with aging and advanced metabolic disease, may therefore represent a pivotal mechanism linking HFD to increased SCD risk.

physiology↗

bioRxiv2025

Cancer systems immunology reveals myeloid - T cell interactions and B cell activation mediate response to checkpoint inhibition in metastatic breast cancer

Sensitization of the immune-suppressed tumor microenvironment (TME) of breast cancer by histone deacetylase inhibition shows promise, but the mechanisms of sensitization are unknown. We investigated the TME of breast-to-lung metastases by combining experimental and clinical data with theory. Knowledge-guided subclustering of single-cell RNA-sequencing data and cell circuits analysis identified 39 cell states and salient interactions, of which myeloid, T cell and B cell subpopulations were most affected by treatment. Using functional immunologic assays, we verified that inhibition of the ICAM pathway partially recapitulated treatment effects. Mathematical modeling of tumor-immune dynamics confirmed that tumor reduction required simultaneous modulation of multiple TME interactions. We found evidence that treatment affected anti-tumor antibody production. Analysis of patient biopsies via spatial proteomics corroborated preclinical findings: in responders we observed increased B cell activation, mature tertiary lymphoid structures, and increased CD8+ T cell--macrophage distances with treatment. Overall, this study provides a framework for the discovery of cell-cell interactions that govern responses in complex TMEs. Statement of significanceThis study provides a framework for the discovery of cell-cell interactions that control responses in complex TMEs. We not only identify impactful tumor immunologic interactions that facilitate sensitization of the metastatic TME but also demonstrate how interdisciplinary data integration fuels cancer systems immunology to accelerate discovery of mechanisms of successful immunotherapeutic response in breast cancers and other previously unresponsive solid tumor types.

cancer biology↗