Biology subjects

Wan, M.

Publications and source records attributed to Wan, M..

At least 19 recordsLinked to original sources

bioRxiv2026

High resolution interaction surface mapping by PRISMA reveals novel ARID1A interactions

The SWI/SNF chromatin remodelling complex controls proliferation and cell fate determination by regulating chromatin accessibility at promoters and enhancers, thereby modulating programs of gene expression, and has roles in DNA damage response, replication, splicing, and translation, and cell plasticity. The cBAF-exclusive subunit ARID1A acts as scaffold for the assembly of cBAF SWI/SNF complexes through its C-terminal globular domain and is the most frequently mutated SWI/SNF subunit in cancer. More than half of the ARID1A protein sequence contains regions of intrinsic disorder which are important for protein interactions, often mediated by short linear motifs. However, these interactions are notoriously difficult to study. Whilst hundreds of ARID1A interactions have been reported in the literature, their molecular basis remains obscure, and only a few have been explored functionally or mapped at an interface level. Here, we use a PRotein Interaction Screen on a peptide MAtrix (PRISMA) combined with quantitative mass spectrometry to identify novel ARID1A interactions and map amino acid residues and motifs that mediate interactions at high sequence resolution. The ARID1A PRISMA assay recapitulates binding of BAF subunits to ARID1A and detects the previously described binding of YAP1 transcriptional coactivator to a PPXY motif. Our PRISMA data reveals binding sites for transcriptional repressor SIN3A and identifies TOX4, CDK2 and CCNA2 as novel interactors. Mutation of a cell cycle-dependent CDK2 phosphorylation site in ARID1A leads to altered gene expression of microtubule factors and defects in cell proliferation. Our work underscores the utility of PRISMA to uncover weak or low abundance interactions that are not detectable by traditional affinity purification strategies. Together, our results characterise novel interactors and a new mode of regulation of ARID1A, and provide a useful resource to further explore mechanistic aspects of ARID1A function.

biochemistry↗

bioRxiv2026

PRDM1 Drives Chemoradiotherapy-associated Enrichment of Adaptive NK Cells in Cervical Cancer

Chemoradiotherapy (CRT) induces not only direct tumor cell death but also extensive remodeling of the tumor immune microenvironment. Adaptive natural killer (aNK) cells, initially characterized in chronic viral infection, are increasingly recognized as functionally relevant immune populations in solid tumors, where they may contribute to antitumor immunity and immune memory. However, how aNK cells dynamically respond to chemoradiotherapy and the regulatory mechanisms underlying their activation in solid tumors remain poorly defined. To address this, we analyzed single-cell RNA sequencing data from cervical cancer patients collected before CRT, after the first CRT fraction, and after the second fraction. Single-cell profiling revealed a significant enrichment of aNK cells following CRT. Differential gene expression and pathway enrichment analyses demonstrated that CRT-associated aNK cells exhibit enhanced virus-defending programs with increased cytotoxicity. Among the differentially expressed genes, the transcription factor PRDM1 was consistently and robustly upregulated in aNK cells after both the first and second rounds of CRT. To investigate the functional role of PRDM1, we applied in silico perturbation analyses using scTenifoldKnk and CellOracle. Virtual knockout of PRDM1 resulted in a marked attenuation of effector programs and disruption of metabolic networks in aNK cells. Moreover, PRDM1 perturbation altered inferred cellular trajectories, reversing the progression toward the aNK cell state, suggesting a requirement for PRDM1 in maintaining aNK identity and functional maturation within the CRT-conditioned tumor microenvironment. Together, these findings identify PRDM1 as a key regulatory factor associated with the enrichment, functional activation, and trajectory stabilization of aNK cells following CRT in cervical cancer, providing insight into innate immune remodeling during CRT and highlighting PRDM1 as a potential target for enhancing radiotherapy-induced antitumor immunity.

immunology↗

bioRxiv2026

Mapping Gene Drive Dynamics onto Mendelian Models

CRISPR-based gene drives bias their own transmission and can spread even when deleterious, giving rise to evolutionary dynamics that can be substantially more complex than those governed by standard Mendelian inheritance. Identifying conditions under which gene-drive dynamics can be faithfully approximated by Mendelian models would therefore enable the extensive theoretical toolkit of classical population genetics to be applied to gene-drive systems. Here, we develop a general mapping framework that translates gene-drive models into dynamically equivalent Mendelian models, allowing their behavior to be analyzed using classical theory. By deriving both haploid and diploid effective-parameter mappings, we identify Mendelian models that closely reproduce allele-frequency trajectories of gene drives across a wide range of conversion rates, fitness costs, and dominance effects. We delineate the regions of the parameter space where a one-parameter haploid approximation provides an accurate first-order representation, and where incorporating dominance in a diploid mapping substantially improves fidelity and recovers internal equilibria and threshold behavior. Analytic approximations yield efficient mappings across most of the drive parameter space, while a trajectory-based grid search further improves accuracy near nonlinear regime boundaries. To demonstrate the utility of this framework, we apply it to predicting gene swamping in a two-deme migration-selection model and show that the mapped Mendelian system accurately forecasts transitions between fixation and loss under three relevant release scenarios: environmental variation in fitness, engineered fitness asymmetries, and environment-dependent conversion. Together, these results establish a theoretical bridge between non-Mendelian gene drives and classical population genetic models, providing an interpretable and computationally efficient foundation for predicting gene-drive outcomes and guiding the design of gene drive systems and deployment strategies.

evolutionary biology↗

bioRxiv2025

Vascular Microbleeds Without Brain Atrophy: A Microvascular Signature of Mid-Stage 5xFAD Pathology

Cerebral microbleeds are increasingly recognized as a downstream manifestation of vascular injury in Alzheimers disease (AD), arising secondary to cerebral amyloid angiopathy (CAA). Here, we examined the pathological specificity of microbleeds by comparing an amyloidosis mouse model (5xFAD) with a small-vessel disease (SVD) model characterized by vascular smooth-muscle cell loss. In vivo multimodal MRI, including gradient-echo, spin-echo, and diffusion-weighted imaging, was complemented by ex vivo high-resolution anatomical scans for validation. Both in vivo and ex vivo gradient-echo MRI consistently revealed hippocampal microbleeds in the 5xFAD model without macroscopic atrophy or ventricular enlargement, whereas no microbleeds or blood-brain barrier disruption were detected in the SVD model. Diffusion-weighted MRI further showed region-specific alterations in apparent diffusion coefficient within the midbrain of 5xFAD mice, but not in other regions or in the SVD cohort. These findings indicate that microbleeds are a pathology-specific marker of amyloid-related vascular injury. The imaging evidence underscores the potential of microbleeds as a disease-specific biomarker for detecting amyloid-driven vascular fragility and refining diagnostic and therapeutic strategies for AD.

neuroscience↗

bioRxiv2025

BudFinder: A Masked Auto-Encoder Vision Transformer Framework for Yeast Budding Detection

Yeast replicative lifespan is a crucial part of aging research, yet its quantification remains labor-intensive and time-consuming, particularly when using time-lapse imaging and microfluidics. Manual counting methods for cell division events are prone to bias and inefficiency, while existing automated approaches often require extensive annotated datasets. These limitations hinder the adaptability of such tools across different microfluidic setups. To address these challenges, we propose a versatile image analysis approach that accurately detects yeast cell division events. To reduce the burden of requiring a large cell division annotated dataset, we pretrained a Masked Auto-Encoder on large-scale segmented yeast cell images. This substantially reduced the annotated data needed to train the transformer model for detecting cellular division events. Additionally, the model is trained directly on budding event detection, circumventing reliance on arbitrary heuristics, such as changes in cell area. By leveraging self-supervised pretraining, we reduced the training data requirement to fewer than 50 mother cells ([~]1,000 divisions), representing a >5-fold reduction compared to prior methods while maintaining comparable accuracy. Author SummaryOur work addresses a longstanding challenge in in live-cell time-lapse microscopy analysis, namely, automating cellular division tracking while minimizing the amount of training data required. Traditionally, scientists identify each division event by manually inspecting thousands of time-lapse images, a process that is both tedious and prone to bias. While automated tools exist, they often require large amounts of annotated data to work effectively, limiting their use across different experimental setups. To overcome these barriers, we developed BudFinder, which can recognize and track cell divisions with far less training data. Using yeast replicative aging data as an example, we first trained a model to understand what a yeast cell "looks like", using tens of thousands of segmented yeast cell images entrapped in our custom-built microfluidic device. Then, we taught it to detect budding events directly from time-lapse movies. This approach reduces the need for manual labeling by more than five-fold compared to previous structures in place, while maintaining accuracy comparable to existing methods. By making high-throughput analysis of cellular division more accessible, our work paves the way for faster and more scalable quantification of cellular dynamics.

systems biology↗

bioRxiv2025

Single-Cell Integration of Chromatin Accessibility and Transcriptomics Reveals Regulatory Networks in Ovarian Tumor-Infiltrating Adaptive NK Cells

Natural killer (NK) cells are traditionally recognized for their rapid, non-specific responses against virus-infected or malignantly transformed cells, functioning as key effectors of innate immunity. However, a distinct subset known as adaptive NK (aNK) cells has been shown to acquire memory-like properties following viral infections, indicating their capacity for antigen-specific immune recall. Intriguingly, aNK cells have also been identified within the tumor microenvironment, where they can mediate tumor-specific recall responses. Yet, the regulatory mechanisms governing their function in tumor-infiltrating aNK cells remain largely undefined. In this study, we integrated publicly available multiomics datasets from ovarian cancer, including single-cell chromatin accessibility (scATAC-seq) and single-cell RNA sequencing (scRNA-seq), to identify chromatin-accessible regions and construct transcription factors (TF)-gene regulatory networks. To validate and extend these findings, we performed Smart-seq3 on NK cells isolated from ovarian tumors and applied SCENIC analysis to identify TF-driven gene regulation. By integrating results from both analyses, we identified PRDM1 and STAT2 as key TFs, along with their downstream targets CRCP and MTFP1, specifically enriched in tumor-infiltrating aNK cells. The expression levels of CRCP and MTFP1 positively correlated with NK cell infiltration in ovarian cancer tissues, suggesting their potential functions in supporting tumor-specific NK cell memory responses. In addition, external validation using data from the PROMIX clinical trial demonstrated that PRDM1 and STAT2 expression levels are positively associated with both overall survival and aNK cell-associated transcriptional features. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=111 SRC="FIGDIR/small/676040v1_ufig1.gif" ALT="Figure 1"> View larger version (37K): org.highwire.dtl.DTLVardef@6df026org.highwire.dtl.DTLVardef@1b16dc8org.highwire.dtl.DTLVardef@19e916borg.highwire.dtl.DTLVardef@15d431a_HPS_FORMAT_FIGEXP M_FIG C_FIG

immunology↗

bioRxiv2025

RetINaBox: A hands-on learning tool for experimental neuroscience

An exciting aspect of neuroscience is developing and testing hypotheses via experimentation. However, due to logistical and financial hurdles, the experiment and discovery component of neuroscience is generally lacking in classroom and outreach settings. To address this issue, here we introduce RetINaBox: a low-cost open-source electronic visual system simulator that provides users with a hands-on tool to discover how the visual system builds feature detectors. RetINaBox features an LED array for generating visual stimuli and a photodiode array that acts as a mosaic of model photoreceptors. Custom software on a Raspberry Pi computer reads out responses from model photoreceptors and allows users to control the polarity and delay of the signal transfer from model photoreceptors to model retinal ganglion cells. Interactive lesson plans are provided, guiding users to discover different types of visual feature detectors--including ON/OFF, center-surround, orientation selective, and direction selective receptive fields--as well as their underlying circuit computations.

neuroscience↗

bioRxiv2025

Dehydration promotes intracellular lipid synthesis and accumulation

Lipids can be considered a water reservoir used to offset dehydration stress as their oxidation by the mitochondria generates water. However, whether dehydration and the ensuing hypertonic stress directly regulate lipid synthesis is unknown. We found that hypertonic stress decreases cellular oxygen consumption, increases intracellular lipid synthesis, and favors glutamine oxidation as a carbon precursor for lipid synthesis via remodeling mitochondrial metabolism. These findings provide a mechanism whereby cellular dehydration leads to intracellular lipid accumulation, functionally linking water availability to lipid storage.

physiology↗

bioRxiv2025

The intratumoral microbiome is negatively associated with malignancy in dermatofibrosarcoma protuberans

In recent years, growing evidence demonstrated the critical roles of intratumoral microbes in tumors. However, the characteristics of intratumoral microbiome in dermatofibrosarcoma protuberans (DFSP) and their associations with tumor malignancy remains unexplored. Here, 106 whole-genome sequencing (WGS) data of the paired biopsy and blood samples from 53 DFSP patients were utilized, in which we extracted high-quality non-human reads to assign microbial taxonomic annotation by Kraken2 and PlusPF database. Meanwhile, significantly elevated non-human reads, microbial abundance, and bacterial load were detected in tumor compared to blood samples. To minimize the impact of contaminants and noise, we conducted a series of filtering procedures and treated blood samples as negative controls. Notably, we found that over 98% of intratumor microbes belonged to bacteria, and that the dominant taxa and alpha-diversity of intratumoral bacteria all exhibited significantly negative correlations with Ki67 expression in tumors. Specifically, the 84 key intratumoral species were identified to be associated with the reduced tumor malignancy, as evidenced by the diminished Ki67 expression, decreased tumor volume, and fewer COL1A1-PDGFB fusion events. These findings highlighted the tumor-suppressive potential of intratumoral microbiome, thereby providing a novel insight into DFSP pathogenesis and clinical management. IMPORTANCETo our knowledge, this is the first study to explore the intratumoral microbiome in DFSP. Microbial DNA signatures could be detected by whole-genome sequencing (WGS) and enabled a comprehensive view of the microbial landscape, thus we analyzed 106 WGS data of the paired biopsy and blood samples from 53 DFSP patients. Our analysis revealed that the dominant taxa and alpha-diversity of intratumoral microbiome all showed significantly negative correlations with Ki67 expression of tumor samples. This study indicated the tumor-suppressive role of intratumoral microbiome in DFSP and highlighted the potential clinical significance in understanding DFSP pathogenesis and guiding therapeutic management strategies.

microbiology↗

bioRxiv2025

BMP-Dependent Mobilization of Fatty Acid Metabolism Promotes Caenorhabditis elegans Survival on a Bacterial Pathogen

The Bone Morphogenetic Proteins (BMPs) are secreted peptide ligands of the Transforming Growth Factor beta (TGF-{beta}) family, initially identified for their roles in development and differentiation across animal species. They are now increasingly recognized for their roles in physiology and infectious disease. In the nematode Caenorhabditis elegans, the BMP ligand DBL-1 controls fat metabolism and immune response, in addition to its roles in body size regulation and development. DBL-1 regulates classical aspects of innate immunity, including the induction of anti-microbial peptides. We theorized that BMP-dependent regulation of fat metabolism could also promote resilience against microbial pathogens. We found that exposure to a bacterial pathogen alters total fat stores, lipid droplet dynamics, and lipid metabolism gene expression in a BMP-dependent manner. We further showed that fatty acid desaturation plays a major role in survival on a bacterial pathogen, while fatty acid {beta}-oxidation plays a more minor role. We conclude that C. elegans mobilizes fatty acid metabolism in response to pathogen exposure to promote survival. Our investigation provides a framework to study potential metabolic interventions that could support therapeutics that are complementary to antibiotic strategies.

physiology↗

bioRxiv2024

Unveiling the functional heterogeneity of endogenous tissue extracellular vesicles in skeletal muscle through multi-omics

Extracellular vesicles (EVs) have emerged as promising tools for the development of disease biomarkers and therapeutics because they can transfer various bioactive cargos between cells in vivo. A better understanding of the heterogeneous properties of EVs in vivo may provide insights into their biological roles and clinical translation potential. As a proof-of-concept, we report that different EV subpopulations from skeletal muscle tissues have distinct composition signatures and diverse biological effects on recipient cells. Multiple cell types (e.g., myoblasts and endothelial cells (ECs)) can contribute to the pool of muscle tissue-derived EVs, and large EVs (L-EVs) are enriched with proteins related to metabolic regulation, whereas small EVs (S-EVs) are enriched with original muscle cell-specific proteins related to muscle function regulation. Compared with L-EVs, S-EVs exhibited abundant surface proteins and higher cell uptake rates. Moreover, L-EVs and S-EVs can induce diverse changes in global gene expression, metabolic patterns and some cellular behaviors (e.g., proliferation and differentiation) in recipient cells. These results suggest that different EV subpopulations might control tissue hemostasis in a coordinated manner and suggest the importance of reconsidering their favorable role in future applications (e.g., S-EVs for biomarker discovery and L-EVs for metabolic intervention). This study highlights the functional heterogeneity of tissue-derived EVs in vivo, and the selection of an ideal EV subset on the basis of its specific biological properties may be a promising strategy for developing more precise biomarkers or tailored EV therapies for regenerative medicine. Graphic abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=60 SRC="FIGDIR/small/624461v1_ufig1.gif" ALT="Figure 1"> View larger version (23K): org.highwire.dtl.DTLVardef@6a9a0forg.highwire.dtl.DTLVardef@a86cbforg.highwire.dtl.DTLVardef@17b8680org.highwire.dtl.DTLVardef@bb718d_HPS_FORMAT_FIGEXP M_FIG C_FIG SignificanceExtracellular vesicles (EVs) are emerging as important tools for diagnostics, therapeutics, and prognostics in various diseases. Understanding the inherent heterogeneity of EVs is crucial, as distinct subpopulations function differently. While extensive research focuses on EVs derived from cell supernatants, endogenous tissue EVs can more accurately reflect the pathophysiological characteristics of their originating cells. Here, we propose different tissue-EV subpopulations coordinately regulate tissue homeostasis. Small EVs with tissue-specific signatures show promise for biomarker development, while large EVs with metabolic signatures are suitable for metabolic interventions. Additionally, small EVs with enhanced surface proteins are ideal for targeted delivery. This work highlights the importance of selecting tissue-EV subpopulations based on their unique properties for developing precise biomarkers and tailored therapies in regenerative medicine.

cell biology↗

bioRxiv2024

Effects of Aging on the Immune and Periosteal Response to Fracture in Mice

Aging predisposes individuals to reduced bone mass and fragility fractures, which are costly and linked to high mortality. Understanding how aging affects fracture healing is essential for developing therapies to enhance bone regeneration in older adults. During the inflammatory phase of fracture healing, immune cells are recruited to the injury site as periosteal skeletal stem/progenitor cells (pSSPCs) rapidly proliferate and differentiate into osteochondral lineages, allowing for fibrocartilaginous callus formation and complete bone healing. Irrespective of age, how periosteal mesenchymal and immune cells interact during early fracture healing is incompletely understood, limiting our ability to potentially modulate these processes. To address this, we directly analyzed, in parallel, at a single-cell level, isolated murine CD45(+) and CD45(-) periosteal cells dissected from intact and fractured bones, collected three days after injury. Through comprehensive analysis, corroborated by bulk RNA-sequencing, flow cytometry, and histology, we found aging decreases pSSPCs proliferative, marked by a reduced expression of genes required for callus formation and an increased senescence signature. We found that the chemokine Cxcl9 was highly upregulated in aged intact Prrx1+ pSSPCs, predicted to interact with other pSSPCs directly, and associated with increased recruitment of CD8+ T cells at the fracture site three days after injury. Cell-to-cell communication analysis provided insight into the complexity of interactions among the many cell types regulating fracture healing and the impact of aging on these processes. Together, these results provide insight into age-induced alterations in fracture healing, informing the development of improved therapeutic approaches for fragility fractures.

cell biology↗

bioRxiv2024

Improved Methodology for Studying Postnatal Osteogenesis via Intramembranous Ossification in a Murine Bone Marrow Injury Model

Long bone injuries heal through either endochondral or intramembranous bone formation pathways. Unlike the endochondral pathway that requires a cartilage template, the process of intramembranous ossification involves the direct conversion of skeletal stem and progenitor cells (SSPCs) into bone-forming osteoblasts. There are limited surgical methods to model this process in experimental mice. Here, we have improved upon a bone marrow injury model in mice to facilitate the study of bone repair via intramembranous ossification and to assess postnatal regulators of osteogenesis. This method is highly reproducible and user-friendly, and it allows temporal assessment of new bone formation in a short period (3-7 days post-injury) using {micro}CT and frozen section histology. Furthermore, the contributions of SSPCs and mature osteoblasts can be readily assessed using a combination of fluorescent reporter mice and this intramembranous bone marrow injury model. In clinical contexts, intramembranous bone formation is relevant for healing critical size defects, stress fractures, cortical defects, trauma from tumor resections, and joint replacements. SUMMARYThe murine bone marrow injury model is a useful tool to study postnatal osteogenesis through intramembranous ossification. This simplified surgical protocol describes the bone marrow ablation procedure for downstream assessment of new bone formation and cellular responses following injury.

cell biology↗

bioRxiv2024

Serum Amyloid P Secreted by Bone Marrow Adipocytes Drives Skeletal Amyloidosis

The accumulation of amyloid fibrils has been identified in tissues outside the brain, yet little is understood about the formation of extracerebral amyloidosis and its impact on the aging process of these organs. Here, we demonstrate that both transgenic mice modeling Alzheimers disease (AD) and naturally aging mice exhibit accumulated senescent bone marrow adipocytes (BMAds), accompanied by amyloid deposits surrounding the BMAds. Senescent BMAds acquire a secretory phenotype, resulting in a marked increase in the secretion of serum amyloid P component (SAP), also known as pentraxin 2 (PTX2). SAP/PTX2 colocalizes with amyloid deposits around senescent BMAds in vivo and is sufficient to promote the formation of insoluble amyloid deposits from soluble A{beta} peptides in in vitro and ex vivo 3D BMAd-based culture experiments. Additionally, Combined treatment with SAP/PTX2 and A{beta} peptides promotes osteoclastogenesis but inhibits osteoblastogenesis of the precursor cells. Transplantation of senescent BMAds into the bone marrow cavity of healthy young mice is sufficient to induce bone loss. Finally, pharmacological depletion of SAP/PTX2 from aged mice abolishes bone marrow amyloid deposition and effectively rescues the low bone mass phenotype. Thus, senescent BMAds, through the secretion of SAP/PTX2, contribute to the age-associated development of skeletal amyloidosis and resultant bone deficits.

cell biology↗

bioRxiv2023

Senescence of endplate osteoclasts induces sensory innervation and spinal pain

Spinal pain affects individuals of all ages and is the most common musculoskeletal problem globally. Its clinical management remains a challenge as the underlying mechanisms leading to it are still unclear. Here, we report that significantly increased numbers of senescent osteoclasts (SnOCs) are observed in mouse models of spinal hypersensitivity, like lumbar spine instability (LSI) or aging, compared to controls. The larger population of SnOCs is associated with induced sensory nerve innervation, as well as the growth of H-type vessels, in the porous endplate. We show that deletion of senescent cells by administration of the senolytic drug Navitoclax (ABT263) results in significantly less spinal hypersensitivity, spinal degeneration, porosity of the endplate, sensory nerve innervation and H-type vessel growth in the endplate. We also show that there is significantly increased SnOC-mediated secretion of Netrin-1 and NGF, two well-established sensory nerve growth factors, compared to non-senescent OCs. These findings suggest that pharmacological elimination of SnOCs may be a potent therapy to treat spinal pain.

cell biology↗

bioRxiv2023

Membrane-dependent actin polymerization mediated by the Legionella pneumophila effector protein MavH

L. pneumophila propagates in eukaryotic cells within a specialized niche, the Legionella-containing vacuole (LCV). The infection process is controlled by over 330 effector proteins delivered through the type IV secretion system. In this study, we report that the Legionella MavH effector harbors a lipid-binding domain that specifically recognizes PI(3)P (phosphatidylinositol 3-phosphate) and localizes to endosomes when ectopically expressed. We show that MavH recruits host actin capping proteins (CP) and actin to the endosome via its CP interacting (CPI) motif and WH2-like actin-binding domain, respectively. In vitro assays revealed that MavH stimulates robust actin polymerization only in the presence of PI(3)P-containing liposomes and the recruitment of CP by MavH negatively regulates F-actin density at the membrane. Furthermore, in L. pneumophila-infected cells, MavH can be detected around the LCV at the very early stage of infection. Together, our results reveal a novel mechanism of membrane-dependent actin polymerization catalyzed by MavH that may play a role at the early stage of L. pneumophila infection by regulating host actin dynamics.

molecular biology↗

bioRxiv2022

Wnt-associated adult stem cell marker Lgr6 is required for osteogenesis and fracture healing

Despite the remarkable regenerative capacity of skeletal tissues, nonunion of bone and failure of fractures to heal properly presents a significant clinical concern. Stem and progenitor cells are present in bone and become activated following injury; thus, elucidating mechanisms that promote adult stem cell-mediated healing is important. Wnt-associated adult stem marker Lgr6 is implicated in the regeneration of tissues with well-defined stem cell niches in stem cell-reliant organs. Here, we demonstrate that Lgr6 is dynamically expressed in osteoprogenitors in response to fracture injury. Using an Lgr6-null mouse model, we find that Lgr6 expression is necessary for maintaining bone volume and efficient postnatal bone regeneration in adult mice. Skeletal progenitors isolated from Lgr6-null mice have reduced colony-forming potential and reduced osteogenic differentiation capacity due to attenuated cWnt signaling. Lgr6-null mice consist of a lower proportion of self-renewing stem cells. In response to fracture injury, Lgr6-null mice have deficient proliferation of periosteal progenitors and reduced ALP activity. Further, analysis of bone regeneration phase and remodeling phase of fracture healing in Lgr6-null mice showed impaired endochondral ossification and reduced mineralization. We propose that in contrast to not being required for successful skeletal development Lgr6-positive cells have a direct role in endochondral bone repair.

cell biology↗

bioRxiv2022

Senescent Preosteoclast Secretome Promotes Metabolic Syndrome Associated Osteoarthritis through Cyclooxygenase 2

BackgroundMetabolic syndrome-associated osteoarthritis (MetS-OA) is a distinct osteoarthritis phenotype defined by the coexistence of MetS or its individual components. Despite the high prevalence of MetS-OA, its pathogenic mechanisms are unclear. The aim of this study was to determine the role of cellular senescence in the development of MetS-OA. MethodsAnalysis of the human osteoarthritis initiative (OAI) dataset was conducted to investigate the MRI subchondral bone features of MetS-human OA participants. Joint phenotype and senescent cells were evaluated in two MetS-OA mouse models: high-fat diet (HFD)-challenged mice and STR/Ort mice. In addition, the molecular mechanisms by which preosteoclasts become senescent as well as how the senescent preosteoclasts impair subchondral bone microenvironment were characterized using in vitro preosteoclast culture system. ResultsHumans and mice with MetS are more likely to develop osteoarthritis-related subchondral bone alterations than those without MetS. MetS-OA mice exhibited a rapid increase in joint subchondral bone plate and trabecular thickness before articular cartilage degeneration. Subchondral preosteoclasts undergo senescence at the pre- or early-osteoarthritis stage and acquire a unique secretome to stimulate osteoblast differentiation and inhibit osteoclast differentiation. Antagonizing preosteoclast senescence markedly mitigates pathological subchondral alterations and osteoarthritis progression in MetS-OA mice. At the molecular level, preosteoclast secretome activates COX2-PGE2, resulting in stimulated differentiation of osteoblast progenitors for subchondral bone formation. Administration of a selective COX2 inhibitor attenuated subchondral bone alteration and osteoarthritis progression in MetS-OA mice. Longitudinal analyses of the human Osteoarthritis Initiative (OAI) cohort dataset also revealed that COX2 inhibitor use, relative to non-selective nonsteroidal anti-inflammatory drug use, is associated with less progression of osteoarthritis and subchondral bone marrow lesion worsening in participants with MetS-OA. ConclusionsOur findings suggest a central role of a senescent preosteoclast secretome-COX2/PGE2 axis in the pathogenesis of MetS-OA, in which selective COX2 inhibitors may have disease-modifying potential. FundingThis work was supported by the National Institutes of Health grant R01AG068226 and R01AG072090 to M.W., R01AR079620 to S.D., and P01AG066603 to X.C.

cell biology↗