Biology subjects

Fan, Y.

Publications and source records attributed to Fan, Y..

At least 145 records · Page 8Linked to original sources

bioRxiv2024

Ovarian disrupting effects and mechanisms of long- and short-chain per- and polyfluoroalkyl substances in mice

BackgroundThe extensive use of per- and polyfluoroalkyl substances (PFAS) has led to environmental contamination and bioaccumulation. Previous research linked PFAS exposure to female reproductive disorders, but the mechanism remains elusive. Further, most studies focused on legacy long-chain PFOA and PFOS, yet the reproductive impacts of other long-chain PFAS and short-chain alternatives are rarely explored. ObjectivesWe investigated the effects and mechanisms of long- and short-chain PFAS on the ovary and associated ovarian functions. MethodsA 3D in vitro ovarian follicle culture system and an in vivo mouse model, together with approaches of reverse transcription-quantitative polymerase chain reaction, enzyme-linked immunosorbent assay, RNA-sequencing, pharmacological treatment, in situ zymography, histology, in situ hybridization, analytical chemistry, and benchmark dose modeling (BMD), were used to test environmentally relevant exposure levels of six long- and short-chain PFAS on follicle maturation, hormone secretion, and ovulation. ResultsIn vitro exposure revealed that long-but not short-chain PFAS interfered with gonadotropin-dependent follicle maturation, ovulation, and hormone secretion. Mechanistically, long-chain perfluorononanoic acid (PFNA) acted as a peroxisome proliferator-activated receptor gamma (PPAR{gamma}) agonist in granulosa cells to disrupt follicle-stimulating hormone (FSH)-dependent follicle maturation, luteinizing hormone (LH)-stimulated ovulation, and associated gene regulatory pathways. In vivo mouse exposure confirmed the ovarian accumulation of PFNA and the mechanism of PPAR{gamma}-mediated ovarian toxicities of PFNA observed in vitro. The BMD analysis of in vitro and in vivo results suggested human relevant exposure levels of long-chain PFAS in our study pose an extra risk of ovarian defects, with follicular rupture as the most sensitive endpoint. DiscussionUsing in vitro follicle culture and in vivo mouse models, we discovered that long-chain PFAS interfere with gonadotropin-dependent follicle maturation, hormone secretion, and ovulation, posing a non-negligible risk to womens reproductive health including anovulation, irregular menstrual cycles, and sub- or infertility.

pharmacology and toxicology↗

bioRxiv2024

2-D Neural Geometry Underpins Hierarchical Organization of Sequence in Human Working Memory

Working memory (WM) is constructive in nature. Instead of passively retaining information, WM reorganizes complex sequences into hierarchically embedded chunks to overcome capacity limits and facilitate flexible behavior. To investigate the neural mechanisms underlying hierarchical reorganization in WM, we performed two electroencephalography (EEG) and one magnetoencephalography (MEG) experiments, wherein humans retain in WM a temporal sequence of items, i.e., syllables, which are organized into chunks, i.e., multisyllabic words. We demonstrate that the 1-D sequence is represented by 2-D neural representational geometry in WM arising from parietal-frontal regions, with separate dimensions encoding item position within a chunk and chunk position in the sequence. Critically, this 2-D geometry is observed consistently in different experimental settings, even during tasks discouraging hierarchical reorganization in WM and correlates with WM behavior. Overall, these findings strongly support that complex sequences are reorganized into factorized multi-dimensional neural representational geometry in WM, which also speaks to general structure-based organizational principles given WMs involvement in many cognitive functions.

neuroscience↗

bioRxiv2024

Generalizable Neural Models of Emotional Engagement and Disengagement

Emotional experiences are never static but continuously evolve in response to internal and external contexts. Little is known about how neural patterns change as a function of these experiences, particularly in response to complex, real-world stimuli. This study aimed to identify generalizable neural patterns as individuals collectively engage and disengage from emotions dynamically. To do so, we analyzed functional magnetic resonance imaging (fMRI) along with subjective emotional annotations from two independent studies as individuals watched negative and neutral movie clips. We used predictive modeling to test if a model trained to predict a group emotional signature response in one study generalizes to the other study and vice versa. Disengagement patterns generalized specifically across intense clips. They were supported by connections within and between the sensorimotor and salience networks, maybe reflecting the processing of feeling states as individuals regulate their emotions. Prediction success for the engagement signature was mixed, but primarily linked to connections within the visual and between the visual and dorsal attention networks, maybe supporting visual attention orienting as emotions intensify. This work offers potential pathways for identifying generalizable neural patterns contributing to future affective research and clinical applications aiming to better understand dynamic emotional responses to naturalistic stimuli.

neuroscience↗

bioRxiv2023

Timp2 loss-of-function mutation and TIMP2 treatment in murine model of NSCLC: modulation of immunosuppression and oncogenic signaling

Mounting evidence suggests that the tissue inhibitor of metalloproteinases-2 (TIMP2) can reduce tumor burden and metastasis. However, the demonstration of such anti-tumor activity and associated mechanisms using in vivo tumor models is lacking. The effects of a Timp2 functional mutation and administration of recombinant TIMP2 were examined in both orthotopic and heterotopic murine models of lung cancer using C57Bl/6 syngeneic Lewis Lung 2-luciferase 2 cells (LL2-luc2) cells. Mice harboring a functional mutation of TIMP2 (mT2) display markedly increased primary lung tumor growth, increased mortality, enriched vasculature, and enhanced infiltration of pro-tumorigenic, immunosuppressive myeloid cells. Treatment with recombinant TIMP2 reduced primary tumor growth in both mutant and wild-type (wt) mice. Comparison of transcriptional profiles of lung tissues from tumor-free, wt versus mT2 mice reveals only minor changes. However, lung tumor-bearing mice of both genotypes demonstrate significant genotype-dependent changes in gene expression following treatment with TIMP. In tumor-bearing wt mice, TIMP2 treatment reduced the expression of upstream oncogenic mediators, whereas treatment of mT2 mice resulted in an immunomodulatory phenotype. A heterotopic subcutaneous model generating metastatic pulmonary tumors demonstrated that daily administration of recombinant TIMP2 significantly downregulates the expression of heat shock proteins, suggesting a reduction of cell-stress responses. In summary, we describe how TIMP2 exerts novel, anti-tumor effects in a murine model of lung cancer and that rTIMP2 treatment supports a normalizing effect on the tumor microenvironment. Our findings show that TIMP2 treatment demonstrates significant potential as an adjuvant in the treatment of NSCLC. One Sentence SummaryTIMP2 treatment compensates for TIMP2 mutation to reveal immunomodulatory and oncogene regulation that inhibits tumor growth and metastatic niche formation.

cancer biology↗

bioRxiv2023

DNA sensor ABCF1 phase separates with cccDNA to inhibit HBV replication

Objectives: Hepatitis B virus (HBV) covalently closed circular DNA (cccDNA) contributes to viral persistence and recurrence. However, it is still less known how the host immune system responds to cccDNA and suppresses HBV replication. Therefore, it is essential to explore host proteins that interact with cccDNA and efficiently suppress HBV replication. Design: The interaction of cccDNA and ABCF1 was assessed by ChIP and pull-down assay. HBV replication was assessed in different cell models, including cccDNA surrogate-transfected and HBV-infected hepatocytes models. Phase separation of recombinant ABCF1 fusion proteins with cccDNA was observed by fluorescence microscopy. Results: Our data found that ABCF1 interacts with cccDNA to form phase-separated condensates via the poly-glutamine (PolyQ) of N-terminal intrinsically disordered low-complexity domain (LCD). Subsequently, ABCF1-cccDNA phase separation not only activates the type I/III interferon (IFN-I/III) pathway but also prevents Pol II accumulation on cccDNA to inhibit HBV transcription. Conclusions: Taken together, our study showed that ABCF1 acts as an antiviral restriction factor of HBV cccDNA by phase-separation-driven innate immune signaling and transcription inhibition. These findings shed new light on the understanding of host defense against cccDNA and provide a novel promising therapeutic strategy for HBV infection.

microbiology↗

bioRxiv2023

scPharm: identifying pharmacological subpopulations of single cells for precision medicine in cancers

Intratumour heterogeneity is a major challenge that limits the effectiveness of anticancer therapies, thus compromising treatment outcomes. Single-cell RNA sequencing (scRNA-seq) technology offers a means to capture gene expression profiles at a single-cell resolution, while drug perturbation experiments yield valuable pharmacological data at the bulk cell level. Here, we introduce "scPharm", a computational framework to integrate large-scale pharmacogenomics profiles with scRNA-seq data, for identifying pharmacological subpopulations within a tumour and prioritizing tailored drugs. scPharm assesses the distribution of the identity genes of single cell (Cell-ID) within drug response-determined gene list, which is accomplished using the normalized enrichment score (NES) obtained from Gene Set Enrichment Analysis (GSEA) as the statistic. One key strength of scPharm is rooted in the robust positive correlation between NES statistics and drug response values at single-cell resolution. scPharm successfully identifies sensitive subpopulations in ER-positive and HER2-positive human breast cancer tissues, discovers dynamic changes in resistant subpopulation of human PC9 cells treated with Erlotinib, and expands its prediction capabilities to a mouse model. By a thoroughly comparative evaluation with other single-cell prediction tools, scPharm presents the superior predictive performance and computational efficiency. Furthermore, scPharm offers a unique feature by predicting combination strategies, gauging compensation effects or booster effects between two drugs through the Set covering method, as well as evaluating drug toxicity on healthy cells within the tumour microenvironment. Together, scPharm provides a novel approach to uncover therapeutic heterogeneity within tumours at single-cell resolution and facilitates precision medicine in cancers.

bioinformatics↗

bioRxiv2023

uniLIVER: a Human Liver Cell Atlas for Data-Driven Cellular State Mapping

The liver performs several vital functions such as metabolism, toxin removal and glucose storage through the coordination of various cell types. The cell type compositions and cellular states undergo significant changes in abnormal conditions such as fatty liver, cirrhosis and liver cancer. As the recent breakthrough of the single-cell/single-nucleus RNA-seq (sc/snRNA-seq) techniques, there is a great opportunity to establish a reference cell map of liver at single cell resolution with transcriptome-wise features. In this study, we build a unified liver cell atlas uniLIVER by integrative analyzing a large-scale sc/snRNA-seq data collection of normal human liver with 331,125 cells and 79 samples from 6 datasets. Besides the hierarchical cell type annotations, uniLIVER also proposed a novel data-driven strategy to map any query dataset to the normal reference map by developing a machine learning based framework named LiverCT. Applying LiverCT on the datasets from multiple abnormal conditions (1,867,641 cells and 439 samples from 12 datasets), the alterations of cell type compositions and cellular states were systematically investigated in liver cancer.

bioinformatics↗

bioRxiv2023

Multifrequency control of Faraday wave bioassembly for constructing multiscale hPSC-derived neuronal networks

Bioassembly is recently regarded as a critical alternative biofabrication technical route to bioprinting since it can directly manipulate millions of live cells to form multicellular structures with close intercellular proximity, improving contact-dependent cell communication and promoting the emergence of tissue-specific functions. However, acoustic bioassembly techniques are currently limited to generating cytoarchitecture with a single characteristic length which cannot faithfully mimic the multiscale cellular structures in native tissues. To overcome this challenge, herein we report a novel acoustic bioassembly technique that employs multifrequency control of Faraday waves to form multiscale cellular structures. By superimposing multiple sine wave signals with proper amplitude ratios, Faraday waves containing multiple wavelengths can be induced and enabled to generate multiscale structures in few seconds. Using this technique, we construct functional neuronal networks with multiscale connectivity that display spontaneous neuroelectrical activities. We anticipate this technique will find wide applications in tissue engineering and regenerative medicine.

bioengineering↗

bioRxiv2023

GFETM: Genome Foundation-based Embedded Topic Model for scATAC-seq Modeling

Single-cell Assay for Transposase-Accessible Chromatin with sequencing (scATAC-seq) has emerged as a powerful technique for investigating open chromatin landscapes at single-cell resolution. However, analyzing scATAC-seq data remain challenging due to its sparsity and noise. Genome Foundation Models (GFMs), pre-trained on massive DNA sequences, have proven effective at genome analysis. Given that open chromatin regions (OCRs) harbour salient sequence features, we hypothesize that leveraging GFMs sequence embeddings can improve the accuracy and generalizability of scATAC-seq modeling. Here, we introduce the Genome Foundation Embedded Topic Model (GFETM), an interpretable deep learning framework that combines GFMs with the Embedded Topic Model (ETM) for scATAC-seq data analysis. By integrating the DNA sequence embeddings extracted by a GFM from OCRs, GFETM demonstrates superior accuracy and generalizability and captures cell-state specific TF activity both with zero-shot inference and attention mechanism analysis. Finally, the topic mixtures inferred by GFETM reveal biologically meaningful epigenomic signatures of kidney diabetes.

bioinformatics↗

bioRxiv2023

Engineered immunomodulatory extracellular vesicles derived from epithelial cells acquire capacity for positive and negative T cell co-stimulation in cancer and autoimmunity

Extracellular vesicles (EVs) are generated by all cells and systemic administration of allogenic EVs derived from epithelial and mesenchymal cells have been shown to be safe, despite carrying an array of functional molecules, including thousands of proteins. To address whether epithelial cells derived EVs can be modified to acquire the capacity to induce immune response, we engineered 293T EVs to harbor the immunomodulatory CD80, OX40L and PD-L1 molecules. We demonstrated abundant levels of these proteins on the engineered cells and EVs. Functionally, the engineered EVs efficiently elicit positive and negative co-stimulation in human and murine T cells. In the setting of cancer and auto-immune hepatitis, the engineered EVs modulate T cell functions and alter disease progression. Moreover, OX40L EVs provide additional benefit to anti-CTLA-4 treatment in melanoma-bearing mice. Our work provides evidence that epithelial cell derived EVs can be engineered to induce immune responses with translational potential to modulate T cell functions in distinct pathological settings.

cancer biology↗

bioRxiv2023

Polygonum Cillinerve polysaccharide inhibits Transmissible gastroenteritis virus by regulating microRNA-181

Transmissible gastroenteritis virus (TGEV) is an important pathogen that can cause changes in the expression profile of cell miRNA. In this study, PCP could treat piglets infected with TGEV by in vivo experiment. High-throughput sequencing technology was used to detect that 9 miRNAs were up-regulated and 17 miRNAs were down-regulated during PCP inhibition of TGEV infection in PK15 cells. Meanwhile, miR-181 was related to the target genes of key proteins of apoptosis pathway. PK15 was treated with different PCP after transfection of miR-181 mimic or inhibitor. Real-time PCR was used to detect the effect of TGEV replication, electron microscopy, TEM and Hoechst fluorescence staining were used to detect the biological function of the treated cells, and western blot (WB) was used to detect the expression of key signaling factors cyt C, caspase 9 and P53 in the apoptotic signaling pathway. The results showed that compared with the control group, 250 g/mL PCP significantly inhibited the replication of TGEV gRNA and gene N (P < 0.01). PK15 cells in PCP groups (250 and 125 g/mL) had uniform cell morphology and a small number of floating cells by microscopic, no typical virus structure was observed in 250 g/mL PCP group under TEM, and apoptosis staining showed that 250 g/mL PCP significantly reduced the number of apoptotic cells. PCP may inhibit TGEV-induced apoptosis through Caspase-dependent mitochondrial pathway after transfection of miR-181. The above results provided a theoretical basis for further research on the mechanism of PCP anti-TGEV.

pharmacology and toxicology↗

bioRxiv2023

Precise Therapeutic Targeting of Distinct NRXN1+/- Mutations

Given the large number of genes significantly associated with risk for neuropsychiatric disorders, a critical unanswered question is the extent to which diverse mutations --sometimes impacting the same gene--will require tailored therapeutic strategies. Here we consider this in the context of rare neuropsychiatric disorder-associated copy number variants (2p16.3) resulting in heterozygous deletions in NRXN1, a pre-synaptic cell adhesion protein that serves as a critical synaptic organizer in the brain. Complex patterns of NRXN1 alternative splicing are fundamental to establishing diverse neurocircuitry, vary between the cell types of the brain, and are differentially impacted by unique (non-recurrent) deletions. We contrast the cell-type-specific impact of patient-specific mutations in NRXN1 using human induced pluripotent stem cells, finding that perturbations in NRXN1 splicing result in divergent cell-type-specific synaptic outcomes. Via distinct loss-of-function (LOF) and gain-of-function (GOF) mechanisms, NRXN1+/- deletions cause decreased synaptic activity in glutamatergic neurons, yet increased synaptic activity in GABAergic neurons. Reciprocal isogenic manipulations causally demonstrate that aberrant splicing drives these changes in synaptic activity. For NRXN1 deletions, and perhaps more broadly, precision medicine will require stratifying patients based on whether their gene mutations act through LOF or GOF mechanisms, in order to achieve individualized restoration of NRXN1 isoform repertoires by increasing wildtype, or ablating mutant isoforms. Given the increasing number of mutations predicted to engender both LOF and GOF mechanisms in brain disorders, our findings add nuance to future considerations of precision medicine.

neuroscience↗

bioRxiv2023

Analysis of uveal melanoma scRNA sequencing data identifies neoplastic-immune hybrid cells that exhibit metastatic potential

Uveal melanoma (UM) is the most common non-cutaneous melanoma and is an intraocular malignancy that affects nearly 7,000 individuals per year worldwide. Of these, nearly 50% will progress to metastatic disease for which there are currently no effective therapies. Despite advances in the molecular profiling and metastatic stratification of class 1 and 2 UM tumors, little is known regarding the underlying biology of UM metastasis. Our group has identified a disseminated tumor cell population characterized by co-expression of immune and melanoma proteins, (circulating hybrid cells (CHCs), in patients with UM. Compared to circulating tumor cells, CHCs are detected at an increased prevalence in peripheral blood and can be used as a non-invasive biomarker to predict metastatic progression. To identify mechanisms underlying enhanced hybrid cell dissemination we sought to identify hybrid cells within a primary UM single cell RNA-seq dataset. Using rigorous doublet discrimination approaches, we identified UM hybrids and evaluated their gene expression, predicted ligand-receptor status, and cell-cell communication state in relation to other melanoma and immune cells within the primary tumor. We identified several genes and pathways upregulated in hybrid cells, including those involved in enhancing cell motility and cytoskeleton rearrangement, evading immune detection, and altering cellular metabolism. In addition, we identified that hybrid cells express ligand-receptor signaling pathways implicated in promoting cancer metastasis including IGF1-IGFR1, GAS6-AXL, LGALS9-P4HB, APP-CD74 and CXCL12-CXCR4. These results contribute to our understanding of tumor progression and interactions between tumor cells and immune cells in the UM microenvironment that may promote metastasis.

cancer biology↗

bioRxiv2023

Distinct eLPB ChAT projections for methamphetamine anxiety and relapse

Choline acetyltransferase-positive neurons in the external lateral parabrachial nucleus (eLPBChAT) send projections to PKC{delta}-positive (PKC{delta}+) neurons in lateral portion of central nucleus of amygdala (lCeAPKC{delta}) and oval portion of bed nucleus of the stria terminalis (ovBNSTPKC{delta}), forming eLPBChAT-lCeAPKC{delta} and eLPBChAT-ovBNSTPKC{delta} pathways. At least in part, the eLPBChAT neurons positively innervate lCeAPKC{delta} and ovBNSTPKC{delta} through regulating synaptic elements of presynaptic acetylcholine (Ach) release and postsynaptic nicotinic acetylcholine receptors (nAChRs). Methamphetamine (METH) withdrawal anxiety and METH-primed reinstatement of conditioned place preference (CPP) recruit eLPBChAT-lCeAPKC{delta} pathway and eLPBChAT-ovBNSTPKC{delta} pathway in male METH-exposed mice, respectively.

neuroscience↗

bioRxiv2023

Single-cell bisulfite-free 5mC and 5hmC sequencing with high sensitivity and scalability

Existing single cell bisulfite-based DNA methylation analysis is limited by low DNA recovery, and the measurement of 5hmC at single-base resolution remains challenging. Here we present a bisulfite-free single-cell whole-genome 5mC and 5hmC profiling technique, named Cabernet, which can characterize 5mC and 5hmC at single-base resolution with high genomic coverage. Cabernet utilizes Tn5 transposome for DNA fragmentation, which enables the discrimination between different alleles for measuring hemi-methylation status. Using Cabernet, we revealed the 5mC, hemi-5mC and 5hmC dynamics during early mouse embryo development, uncovering genomic regions exclusively governed by active or passive demethylation. We show that hemi-methylation status can be used to distinguish between pre- and post-replication cells, enabling more efficient cell grouping when integrated with 5mC profile. The property of Tn5 naturally enables Cabernet to achieve high-throughput single-cell methylome profiling, where we probed mouse cortical neurons and embryonic day 7.5 (E7.5) embryos, and constructed the library for thousands of single cells at high efficiency, demonstrating its potential for analyzing complex tissues at substantially low cost. Together, we present a new way of high-throughput methylome and hydroxymethylome detection at single-cell resolution, enabling efficient analysis of epigenetic status of biological systems with complicated nature such as neurons and cancer cells. Significance StatementMost of current methylation profiling techniques rely on bisulfite treatment, which suffers low DNA recovery. The technique proposed in this study, named Cabernet, can be used to measure 5mC and 5hmC at single-base resolution with high genomic coverage. By using Tn5 transposome, hemi-methylation status can be measured and high-throughput methylome profiling can be achieved. Together, it provides an efficient way to analyze the epigenetic landscape of complicated biological systems.

genomics↗

bioRxiv2023

BBQ methods: Streamlined workflows for Bacterial Burden Quantification in infected cells by confocal microscopy

Accurate quantification of bacterial burden within macrophages, termed Bacterial Burden Quantification (BBQ), is crucial for understanding host-pathogen interactions. Various methods have been employed, each with strengths and weaknesses. This article addresses limitations in existing techniques and introduces two novel automated methods for BBQ within macrophages based on confocal microscopy data analysis. The first method refines total fluorescence quantification by incorporating filtering steps to exclude uninfected cells, while the second method calculates total bacterial volume per cell to mitigate potential biases in fluorescence-based readouts. These workflows utilize PyImageJ and Cellpose software, providing reliable, unbiased, and rapid quantification of bacterial load. The proposed workflows were validated using Salmonella enterica serovar Typhimurium and Mycobacterium tuberculosis models, demonstrating their effectiveness in accurately assessing bacterial burden. These automated workflows offer valuable tools for studying bacterial interactions within host cells and provide insights for various research applications.

cell biology↗

bioRxiv2023

Powerful and accurate detection of temporal gene expression patterns from multi-sample multi-stage single-cell transcriptomics data

We present a non-parametric statistical method called TDEseq that takes full advantage of smoothing splines basis functions to account for the dependence of multiple time points, and uses hierarchical structure linear additive mixed models to model the correlated cells within an individual. As a result, TDEseq demonstrates powerful performance in identifying four potential temporal expression patterns within a specific cell type. Extensive simulation studies and the analysis of four published scRNA-seq datasets show that TDEseq can produce well-calibrated p-values and up to 20% power gain over the existing methods for detecting temporal gene expression patterns.

genomics↗

bioRxiv2023

Pseudorabies virus causes splenic injury via inducing the oxidative stress and apoptpsos related factors in mice

Pseudorabies virus (PRV) is an immunosuppressive disease that causes significant damage to the pig industry. This study aimed to detect the effects of PRV on oxidative stress related factors and cell apoptosis in the spleen, providing a basis for the research on the pathogenesis of PRV in mice model. Pathological observation was performed by hematoxylin and eosin Y staining. Biochemical and Flow cytometry method were performed to determine the reactive oxygen species profile of the spleen post-infection and apoptosis detection. In addition, q-PCR and Western blot were adopted to measure the apoptotic conditions of the spleen infected with PRV. The results indicated that the ROS level in the PRV infection group was remarkedly increased (p<0 01) at a time-dependent pattern. Furthermore, the Malondialdehyde levels in the spleen of mice in the infection group increased significantly (p<0.01) in a time-dependent mode. However, the Catalase, Superoxide dismutase, and Glutathione activity and expression levels in the infection group were significantly decreased with the control group (p<0 01) in a time-dependent manner. Furthermore, the ratio of splenocyte apoptosis in the infection group significantly increased (p<0 05, p<0 01) in a time-dependent manner. In conclusion, PRV infection causes apoptosis of the spleen via oxidative stress in mice.

microbiology↗