Biology subjects

Chen, B.

Publications and source records attributed to Chen, B..

At least 73 records · Page 4Linked to original sources

bioRxiv2025

Antibody feedback establishes an affinity brake in the germinal center

Recent advances in mRNA vaccine technology have opened the door to novel types of antigen display, but little is yet known as to how B cells recognize and respond to these formats. By delivering an mRNA-LNP encoded membrane-bound immunogen displaying three conserved HIV-1 Envelope (Env) epitopes to knock-in mouse models with B cell receptors (BCRs) of defined affinities, we investigated how epitope-specific competition shapes germinal center (GC) responses. Co-activation of B cells targeting different epitopes did not alter GC kinetics observed in individual activations, but a striking inverse correlation was observed between BCR affinity and GC residence time in either scenario: high-affinity B cells exhibited shorter persistence in GCs, while those with lower affinity to the antigen were maintained. Furthermore, B cells were able to engage in GC reactions at equivalent rates in the presence or absence of clonal lineages binding the same epitope with similar affinities, while higher-affinity clones suppressed lower-affinity counterparts targeting the same epitope. Spatial transcriptomics revealed plasma-like cells within and adjacent to the GC which, together with the detection of early IgG in draining lymph nodes, suggests that local antibody production from these cells may contribute to feedback-driven kinetics. These findings indicate that a self-modulated local antibody feedback loop may act as a "brake" on epitope-specific recognition--dampening further affinity enhancement for high-affinity B cells and facilitating epitope spreading by redirecting the response toward alternative epitopes.

immunology↗

bioRxiv2025

Leveraging Disease Association Degree for High-Accuracy MicroRNA Target Prediction

AO_SCPLOWBSTRACTC_SCPLOWMicroRNAs (miRNAs) regulate gene expression by binding to mRNAs, inhibiting translation, or promoting mRNA degradation. Accurate identification of functional miRNA-target interactions (MTIs), typically validated by methods like western blot or reporter assay, remains challenging due to the scarcity of experimental data compared to the vast number of sequence-based predictions. This study pioneers a novel approach focusing solely on the disease association degree between miRNAs and their target genes. We propose that this single feature is sufficient for distinguishing experimentally validated functional MTIs from sequence-based predicted MTIs in a binary classification task. To quantify miRNA-gene disease association, we fine-tuned Sentence-BERT to generate disease description embeddings and compute their semantic similarity. Remarkably, using only disease association features, miRTarDS achieved an F1 score of 0.88 on the task of distinguishing functional from predicted MTIs in the external validation set. The approach also exhibits generalizability across different gene-disease association databases. This study demonstrates disease association as a powerful, independent dimension for prioritizing high-confidence functional MTIs.

bioinformatics↗

bioRxiv2025

A Study on the Pharmacokinetic/Pharmacodynamic Profiles of the Novel PPAR Pan Agonist Chiglitazar Sodium in Rats with Hypoalbuminemia

ObjectiveTo compare the changes in the pharmacokinetic/pharmacodynamic (PK/PD) profiles of Chiglitazar Sodium between healthy rats and those with hypoalbuminemia, and to evaluate the medication safety of Chiglitazar in rats with hypoalbuminemia. MethodsLiquid Chromatography-Tandem Mass Spectrometry (LC-MS/MS) method was employed to determine the plasma concentration ofChiglitazar Sodium in both healthy rats and rats with hypoalbuminemia after administration. Pharmacokinetics were calculated based on the measured data. Blood glucose levels were monitored and recorded in both groups to observe pharmacodynamic changes. ResultsPharmacokinetic analysis revealed that, compared to the healthy group, only the Tmax in the hypoalbuminemia group was significantly earlier by 34.3% (P<0.001), while other pharmacokinetic parameters showed no significant differences. Pharmacodynamic studies indicated that blood glucose levels remained stable in both healthy rats and rats with hypoalbuminemia during the 0.25-10 hour monitoring period. ConclusionExcept for Tmax, there were no significant differences in PK/PD parameters between rats with hypoalbuminemia and healthy rats. Blood glucose levels remained stable in both groups after oral administration of Chiglitazar Sodium. The animal experimental results suggest that, under normal conditions, hypoalbuminemia does not affect the pharmacokinetic and pharmacodynamic profiles of Chiglitazar Sodium.

pharmacology and toxicology↗

bioRxiv2025

Iron addicted colorectal cancers exploit Heme-Complex II axis to resist oxidative cell death

Colorectal cancer (CRC) cells are addicted to iron, which fuels nucleotide synthesis, mitochondrial respiration, and rapid proliferation. Yet paradoxically, high intracellular iron is cytotoxic to most other cells, raising the question of how CRC cells tolerate and exploit iron-rich environments. One pathway thought to mediate iron toxicity is ferroptosis, an iron-dependent form of cell death. However, most ferroptosis regulators were identified through synthetic chemical screens or small molecule activators, and it remains unclear whether these canonical pathways explain how iron itself triggers cell death, particularly in vivo. Here, using multi-omics profiling, CRISPR screening, and in vivo models, we uncover a heme-succinate dehydrogenase (SDH)-Coenzyme Q (CoQ) axis that enables CRC cells to buffer iron-induced oxidative stress. Heme-dependent SDH reduces CoQ, which redistributes to mitochondrial and plasma membranes to detoxify lipid ROS as a radical trapping antioxidant. This pathway functions alongside, and in some contexts independently of, canonical ferroptosis regulators. These findings reveal that CRCs co-opt metabolic cofactors not only for growth but also for survival under physiologically toxic iron levels, uncovering new vulnerabilities for therapy.

cancer biology↗

bioRxiv2025

Standardized pipeline for establishing, expanding, and differentiating airway and alveolar organoids from human BAL fluid

Lung organoids are versatile experimental models, but their broader use in studying human disease is limited by the scarcity of starting material and the complexity of current methods. To align organoid technology with common clinical practice, we developed airway and alveolar organoids using cells obtained from patients bronchoalveolar lavage (BAL) fluid. Building on existing techniques, we showed that BAL is a reliable, accessible source of primary human epithelial cells, yielding airway and alveolar organoids within 10 days. Organoids can then be expanded over many passages for downstream analysis. Our streamlined methods do not require cell sorting or other complex procedures, all cells are derived from a single patient, and media are based on serum-free, chemically-defined formulations. Here, we present detailed protocols for organoid establishment, standardized passaging and phenotyping, and differentiation of both airway and alveolar models. We provide a time course of BAL-derived airway organoid differentiation at air-liquid interface, and we demonstrate proof of principle for differentiation of BAL-derived alveolar organoids in 3D culture. These methods can be readily adapted to generate and characterize organoids from lung tissue, tracheobronchial specimens, or other primary cells from humans or mice, expanding the potential to use lung organoids for disease modeling. NEW AND NOTEWORTHYWe provide streamlined protocols to generate both airway and alveolar epithelial organoids from a single, clinical BAL specimen. From standardized specimen collection to organoid plating to passaging and differentiation, we show that rare, primary epithelial cells in BAL can give rise to all the major airway and alveolar cell types. Our serum-free, feeder-free, sorting-free methods offer a simplified starting point for using patient-derived organoids to model lung disease. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=97 SRC="FIGDIR/small/682184v1_ufig1.gif" ALT="Figure 1"> View larger version (24K): org.highwire.dtl.DTLVardef@e0abe2org.highwire.dtl.DTLVardef@349f9aorg.highwire.dtl.DTLVardef@da4b0eorg.highwire.dtl.DTLVardef@175a97f_HPS_FORMAT_FIGEXP M_FIG Graphical Abstract Standardized pipeline for BAL organoids. Created in BioRender. C_FIG

cell biology↗

bioRxiv2025

Multi-immersion Oblique Plane Microscope (miOPM): A reconfigurable platform for high-resolution Light-Sheet Fluorescence Microscopy.

Light-Sheet Fluorescence Microscopy (LSFM) enables gentle, rapid, and efficient volumetric imaging of biological specimens. Despite its potential, LSFM has fragmented into multiple variants, each narrowly optimized for specific samples or imaging regimes, limiting its broad applicability. To overcome this barrier, we introduce the multi-immersion oblique plane microscope (miOPM), an adaptable LSFM platform for high-resolution imaging ranging from subcellular dynamics to whole organisms and cleared tissues. miOPM uniquely supports seamless interchangeability of oil, water, and air objectives, and maintains diffraction limited performance across a refractive index range of 1.33 - 1.51. It is compatible with standard sample mounting, current clearing protocols and high-throughput 3D imaging. We leverage miOPM to image diverse biological specimens at multiple spatial scales. By greatly expanding the adaptability and ease of use of LSFM, miOPM is poised to democratize advanced three-dimensional imaging.

bioengineering↗

bioRxiv2025

A Single Structure-Derived Computational Metric Predicts High-Affinity Antibody Selection Against a Malaria Antigen

There is an increasing need for improved malaria antibodies that can be used in passive immunization strategies to reduce the burden of malaria in endemic regions. Despite considerable progress, the identification or development of variants that meet stringent performance requirements remains a challenge. A key strategy has been the improvement of prototypic antibodies targeting the repeat antigens on Plasmodium falciparum circumsporozoite protein (PfCSP). In this work, we derive a computational metric from predicted protein structures that efficiently captures affinity information of antibody variants of the PfCSP-targeting antibody, CIS43. We then use this metric to rapidly explore sequence space as large as >3x10^47 variants using principles of the germinal center, deriving new high-affinity CIS43 variants from the method. We further extend this framework to generate high-affinity variants of an unrelated PfCSP-targeting antibody, L9, by maturing both homotypic and antigen-binding interactions, which demonstrates substantial flexibility of the approach. Taken together, we show that coupling micro-evolutionarily selected mutations to in silico screening permits the selection of high-affinity malaria antibodies.

immunology↗

bioRxiv2025

Engineered Promoter System Enables High-efficiency Transgenic CRISPR Editing in Malaria Transmitting Mosquito Anopheles sinensis

The CRISPR/Cas9 system deployed through crosses of transgenic lines expressing Cas9 and gRNA facilitates efficient mutagenesis. However, its application in non-model insects remains limited, primarily due to a lack of well-characterized promoters capable of driving robust and stable expression of Cas9 and gRNA. In the malaria mosquito Anopheles sinensis, we evaluated several ovary-biased promoters--Asvasa2, Aszpg, and Asnanos--for driving Cas9 expression. Notably, the Asvasa2 promoter mediated mutagenesis in nearly 60% of G0 individuals following microinjection of gRNAAswhite. Among four RNA polymerase III promoters derived from AsU6 genes, AsU6-1 yielded the highest gRNA transcriptional output, enabling 62% editing efficiency in G0 offspring. In addition, hybrid crosses between established transgenic lines demonstrated that the Asvasa2-Cas9 and AsU6-1-gRNA combination enabled complete germline editing penetrance, where all F2 progeny inherited the intended mutations. This work provides a essential genetic toolkit for synthetic biology applications in Anopheles mosquitoes and a scalable framework for engineering other non-model insects.

genetics↗

bioRxiv2025

Breaking Free from the Acquisition Dogma for Volume Electron Microscopy

Volume electron microscopy (VEM) enables nanometer-resolution three-dimensional (3D) visualization of biological specimens via serial sectioning and imaging. Owing to precautions against additional challenges in downstream analysis, VEM datasets are often acquired unnecessarily at slow speeds and high resolutions, thereby limiting achievable imaging throughput. By systematically searching for optimal VEM acquisition conditions, we find that sufficient spatial resolution effectively counteracts high image noise in preserving 3D structural information. To further verify that denoising is more effective in restoring volumetric datasets than axial interpolation, machine learning-based methods, including a newly developed 3D context-based denoising model, were compared through various tasks on VEM datasets acquired simultaneously. Our volumetric approach not only outperforms other baseline methods in faithful feature recovery but also facilitates robust serial block-face cutting down to 20 nm by allowing fast imaging. This work provides both an optimized acquisition strategy and volumetric denoising methods as actionable guidelines for maximizing VEM throughput.

bioinformatics↗

bioRxiv2025

Affinity Maturation and Light-Chain-Mediated Paratope Diversification Anticipate Viral Evolution

A key goal of vaccinology is to train the immune system to combat current pathogens while simultaneously preparing it for future evolved variants. Understanding factors contributing to anticipatory breadth, wherein affinity maturation against an ancestral strain yields neutralization capacity against evolved variants, is therefore of great importance. Here, we investigated the mechanism of anticipatory breadth development in a public antibody family targeting the functionally restricted ACE2 binding site on SARS-CoV-2. IGHV3-53/66 antibodies isolated from memory B cells of infection-naive individuals vaccinated with the ancestral Wuhan-strain mRNA vaccine frequently neutralized evolved Omicron variants and contained several hallmark mutations previously shown to enhance neutralization breadth. Comparative analyses with antibodies from Omicron breakthrough infections revealed that breadth-associated patterns of somatic hypermutation emerged independently of variant exposure. However, Omicron infection had a marked impact on light chain pairing frequencies, suggestive of variant-imposed selection of favorable light chains. Analysis of available IGHV3-53/66 antibody structures complexed with SARS-CoV-2 receptor binding domain (RBD) clarified these findings; convergent somatic mutations on the heavy chain largely refined contacts with invariant RBD residues, while light chain pairings shifted epitopes to avoid steric challenges posed by Omicron mutations. These findings support a model of anticipatory breadth with three key elements: (1) targeting of a functionally restricted epitope, (2) affinity maturation to establish an affinity buffer, and (3) variable chain pairing to generate paratope diversity. These elements each serve to compensate for a distinct consequence of viral mutagenesis, offering a mechanistic framework for anticipating viral evolution. Graphical Summary O_FIG O_LINKSMALLFIG WIDTH=157 HEIGHT=200 SRC="FIGDIR/small/672735v1_ufig1.gif" ALT="Figure 1"> View larger version (51K): org.highwire.dtl.DTLVardef@1e4e893org.highwire.dtl.DTLVardef@1610bddorg.highwire.dtl.DTLVardef@520909org.highwire.dtl.DTLVardef@1cf4edc_HPS_FORMAT_FIGEXP M_FIG C_FIG

immunology↗

bioRxiv2025

MAIT cell enrichment in Lynch syndrome is associated with immune surveillance and colorectal cancer risk

Tissue microenvironment characteristics associated with elevated risk of colorectal cancer (CRC) in Lynch syndrome (LS) are poorly characterized. We applied the multimodal single cell sequencing platform ExCITE-seq to define the colonic cellular composition and transcriptome of LS carriers with and without a history of CRC compared with general population controls. Our analysis revealed widespread remodeling in LS that included striking expansion of epithelial stem and progenitor cells, and loss of fibroblast populations. Although clonally expanded and terminally exhausted CD8 T cells were more prominent in individuals with a history of CRC, LS carriers without CRC displayed enrichment of cytotoxic mucosal-associated invariant T (MAIT) cells associated with CCL20 expression in epithelial progenitors, validated by orthogonal techniques including demonstration of a protective function in a murine model of CRC. These findings highlight cellular features that distinguish LS carriers and suggest a protective role of MAIT cells in human CRC surveillance.

immunology↗

bioRxiv2025

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

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

pharmacology and toxicology↗

bioRxiv2025

Suppression of interferon signaling via small molecule modulation of TFAM

The mitochondrial transcription factor A (TFAM) is essential for mitochondrial genome maintenance. It binds to mitochondrial DNA (mtDNA) and determines the abundance, packaging and stability of the mitochondrial genome. Because its function is tightly associated with mtDNA, TFAM has a protective role in mitochondrial diseases and supportive studies demonstrate reversal of disease phenotypes by TFAM overexpression. In addition, TFAM deficiency has been shown to cause release of mtDNA into the cytosol and activation of the cGAS/STING innate immune response pathway. As such, TFAM presents as a unique target for therapeutic intervention, but limited efforts for activators have been reported. Herein we disclose novel TFAM small molecule modulators with sub-micromolar activity. Our results demonstrate that these compounds result in increase of TFAM protein levels and mtDNA copy number. This results in inhibition of a mtDNA stress-mediated inflammatory response by preventing mtDNA escape into the cytosol. Furthermore, we see beneficial effects in cellular disease models in which boosting TFAM activity has been advanced as a disease modifying strategy including improved energetics in MELAS cybrid cells and a decrease of fibrotic markers in Systemic Sclerosis (SSc) fibroblasts. These results highlight the therapeutic potential of using small molecule TFAM activators in indications characterized by mitochondrial dysfunction.

cell biology↗

bioRxiv2025

GFFx: A Rust-based suite of utilities for ultra-fast genomic feature extraction

Genome annotations are becoming increasingly comprehensive due to the discovery of diverse regulatory elements and transcript variants. However, this improvement in annotation resolution poses major challenges for efficient querying, especially across large genomes and pangenomes. Existing tools often exhibit performance bottlenecks when handling large-scale genome annotation files, particularly for region-based queries and hierarchical model extraction. Here, we present GFFx, a Rust-based toolkit for ultra-fast and scalable genome annotation access. GFFx introduces a compact, model-aware indexing system inspired by binning strategies and leverages Rusts strengths in execution speed, memory safety, and multithreading. It supports both feature- and region-based extraction with significant improvements in runtime and scalability over existing tools. Distributed via Cargo, GFFx provides a cross-platform command-line interface and a reusable library with a clean API, enabling seamless integration into custom pipelines. Benchmark results demonstrate that GFFx offers substantial speedups and makes a practical, extensible solution for genome annotation workflows.

bioinformatics↗

bioRxiv2025

in vivo Reprogramming of NG2 Glia Improves Bladder Function After Spinal Cord Injury

Neurogenic bladder is a debilitating consequence of spinal cord injury (SCI), with few treatment options that restore voluntary voiding. Here, we show that SOX2-mediated in vivo reprogramming of NG2 glia improves bladder function in a clinically relevant mouse model of contusive SCI. NG2 glia reprogramming induces adult neurogenesis, reduces glial scarring, and significantly improves urinary performance, as measured by voiding assays and conscious cystometry. Functional recovery correlates positively with neurogenesis and negatively with glial scarring. These findings demonstrate that SOX2-mediated glial reprogramming promotes autonomic repair and offers a regenerative strategy for neurogenic bladder after SCI.

neuroscience↗

bioRxiv2025

The Rab32-LRMDA-Retriever Complex is a Key Regulator of Intestinal Immune Homeostasis

Maintaining intestinal homeostasis relies on the intricate interplay among the mucosal epithelium, immune system, and host microbiome. A key question is how innate immune cells sense and process microbes in the gut lumen, eliciting appropriate protective responses without causing tissue injury. Clearance of invading microbes and initiation of downstream inflammatory responses are central to this process and require proper function of the endolysosomal system. Dysfunction of this system can predispose the host to chronic inflammatory disorders and acute infections. Here, through forward genetic screening of N-ethyl-N-nitrosourea (ENU)-mutagenized mice and CRISPR/Cas9 validation, we identify Lrmda, encoding leucine-rich melanocyte differentiation-associated protein (LRMDA), as a key regulator of intestinal homeostasis. Using hematopoietic chimera and conditional knockouts, we show that LRMDA functions primarily in CD11c+ cells, including mucosal dendritic cells (DCs) and macrophages, but not in non-hematopoietic cells. Proteomic, cellular, and biochemical analyses reveal that LRMDA directly and cooperatively interacts with the endolysosome-specific small GTPase Rab32 and the endosomal recycling complex Retriever. Loss of LRMDA or Retriever function increases susceptibility to dextran sodium sulfate (DSS)-induced colitis and impairs clearance of Listeria monocytogenes. Together, our findings establish the Rab32-LRMDA-Retriever complex as a critical regulator of endolysosomal trafficking in innate immune cells, essential for maintaining intestinal immune homeostasis.

immunology↗

bioRxiv2025

Mitochondrial DNA Mutations Determine Favourable Molecular Responses to Targeted Kinase Inhibitor Therapy and Impair Oxidative Phosphorylation

Somatic mutations in mitochondrial DNA (mtDNA) are not typically considered key oncogenic drivers of cancer, primarily because of a high synonymous to non-synonymous variant ratio. Here, we surveyed 248 matched diagnosis and remission samples from patients with chronic myeloid leukemia (CML) and found a 75% had mitochondrial mutations with a median number of 2 mutations per patient. mtDNA mutations were predominantly non-synonymous, enriched in the D-loop control region, and likely originated from replication and transcriptional errors. Functionally, mtDNA mutations were associated with reduced oxidative phosphorylation (OXPHOS), as measured by Seahorse analyser. This metabolic vulnerability could be phenocopied by treatment with the complex I inhibitor IACS-10759 in combination with the targeted tyrosine kinase inhibitor (TKI) imatinib, which significantly reduced the colony-forming potential of TKI resistant leukemic stem/progenitor cells (LSPCs). Strikingly, we show that mtDNA mutations were associated with increased sensitivity to imatinib therapy in the clinic. Patients with [≥]3 mutations and patients with mutations in the D-loop showed significantly higher cumulative incidence of major molecular response at 24 months (90% vs. 68%, p = 0.004, and 89% vs 68%, p = 0.004 respectively). Single-cell RNA sequencing further revealed enrichment in non-synonymous mtDNA variants in LSPCs from TKI-sensitive patients, while TKI-resistant cells exhibited upregulated gene signatures related to glycerolipid and phospholipid metabolism and mitochondrial biogenesis. Together, our findings demonstrate that mtDNA mutations are key determinants of sensitivity to targeted therapy, rather than oncogenic drivers of leukemogenesis. Mechanistically, non-synonymous mtDNA mutations appear to restrict mitochondrial metabolic plasticity, with widespread implications for precision oncology.

cancer biology↗

bioRxiv2025

Construction of recombinant Bacillus subtilis spores surface-displaying PRRSV ORF1b, GP5, M, and N proteins in tandem and evaluation of their immunogenicity in mice

Porcine reproductive and respiratory syndrome virus (PRRSV), a single-stranded RNA virus, is a highly contagious pathogen that causes severe reproductive and respiratory disorders in pigs, leading to significant economic losses in the swine industry worldwide. However, current commercial vaccines provide only limited protection against circulating PRRSV strains, highlighting the urgent need for novel vaccine strategies. B. Bacillus subtilis, a well-characterized probiotic, has emerged as a promising platform for mucosal vaccine delivery due to its safety profile and ability to induce robust immune responses. In this study, we screened B cell and T cell linear epitopes from PRRSV-ORF1b, GP5, M, and N proteins to construct a recombinant tandem antigen (ER), which was displayed on the surface of B. subtilis 168 spores. The resulting recombinant strain, designated B. subtilis BE, was evaluated for immunogenicity in mice via oral administration. The results showed that Oral immunization with B. subtilis BE significantly elevated antigen-specific secretory IgA levels in intestinal contents and IgG levels in serum, indicating potent mucosal and systemic humoral responses. Moreover, B. subtilis BE induced measurable neutralizing antibody titers and enhanced cellular immunity, as evidenced by increased frequencies of CD3+ and CD8+ T cells in mesenteric lymph nodes and upregulation of IFN-, TNF-, and IFN-{gamma} expression in intestinal tissues. Collectively, these findings demonstrate that B. subtilis BE elicits both humoral and cellular immune responses and may serve as a novel oral vaccine candidate against PRRSV, offering a promising alternative for controlling the ongoing epidemic.

immunology↗