Biology subjects

Chen, L.

Publications and source records attributed to Chen, L..

At least 613 records · Page 34Linked to original sources

bioRxiv2022

Development of an affinity-enhanced clinical candidate TCR targeting NY-ESO-1 with optimal potency and high specificity

The clinical success of T-cell receptor (TCR)-based immunotherapy depends on the efficacy and specificity of TCRs. Naturally occurring TCRs have limited anti-tumor potency due to their low affinity for tumor antigens. Affinity enhancement is a promising strategy to generate highly potent TCRs. However, it is concerned that affinity-enhanced TCRs are prone to lose specificity. We isolated low affinity TCRs specific for NY-ESO-1157-165/HLA-A*02:01 from peripheral blood mononuclear cells of healthy donors. An affinity-enhanced TCR candidate with optimal affinity and specificity was generated using phage display and an extensive set of in vitro and in vivo assays. Alanine scanning mutagenesis showed that the TCR candidate retained specificity by making extensive contacts to the side chains of NY-ESO-1157-165 peptide. Adoptive transfer of T cells engineered with this candidate (termed TAEST16001) significantly inhibited tumor growth in subcutaneous, metastatic, and patient-derived xenograft (PDX) mouse tumor models. This study demonstrates that sophisticated engineering and screening techniques can be utilized to generate a clinical candidate TCR with potent anti-tumor activity without losing specificity. TAEST16001 was approved by the Center for Drug Evaluation (CDE) as the first TCR-based immunotherapy clinical trial in China (ClinicalTrials.gov Identifier: NCT03159585).

immunology↗

bioRxiv2022

The Serotonergic Neural Circuit Between The Dorsal Raphe Nucleus And The Basolateral Amygdala Is Implicated In Modulating The Arousal from Sevoflurane Anesthesia

Although some advancements concerning the arousal involved in mediating the delayed emergency from general anesthesia, which will lead to the serious complications, had been made, the role by arousal in modulating in delayed emergency still remains to be unclear. In our models, based on our previous working that activation of the 5-Hydroxytryptamine (5-HT) neurons in the dorsal raphe nucleus (DRN) by optogenetics can significantly reduce the emergency time by activating arousal pathway, we further test whether the serotonergic neural circuit between the DRN and the basolateral amygdala (BLA) is implicated in modulating the arousal from the sevoflurane anesthesia and the emergency time of sevoflurane anesthesia by the pharmacological, optogenetics and fiber photometry. Our findings showed that whether the serotonergic neural circuit between the DRN and the basolateral amygdala (BLA) plays a key role in modulating the arousal from the sevoflurane anesthesia and the emergency time of sevoflurane anesthesia. Based on the serotonergic neural circuit, the 5-HT 1 A receptor is of great significance to mediate the arousal and the emergency time of the sevoflurane anesthesia. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=143 SRC="FIGDIR/small/511924v2_ufig1.gif" ALT="Figure 1"> View larger version (58K): org.highwire.dtl.DTLVardef@34621eorg.highwire.dtl.DTLVardef@115ae09org.highwire.dtl.DTLVardef@54eb12org.highwire.dtl.DTLVardef@11515ff_HPS_FORMAT_FIGEXP M_FIG O_FLOATNOFigure1C_FLOATNO C_FIG

neuroscience↗

bioRxiv2022

Disulfide stabilization reveals conserved dynamic features between SARS-CoV-1 and SARS-CoV-2 spikes.

SARS-CoV-2 spike protein (S) is structurally dynamic and has been observed by cryo-EM to adopt a variety of prefusion conformations that can be categorized as locked, closed and open. The locked conformations feature tightly packed trimers with structural elements incompatible with RBD in "up" position. For SARS-CoV-2 S, it has been shown that the locked conformations are transient under neutral pH. Probably due to their transience, locked conformations remain largely uncharacterized for SARS-CoV-1 S. Intriguingly, locked conformations were the only conformations captured for S proteins of bat and pangolin origin SARS-related coronaviruses. In this study, we introduced x1, x2, and x3 disulfides into SARS-CoV-1 S. Some of these disulfides have been shown to preserve rare locked conformations when introduced to SARS-CoV-2 S. Introduction of these disulfides allowed us to image a variety of locked and other rare conformations for SARS-CoV-1 S by cryo-EM. We identified bound cofactors and structural features that are associated with SARS-CoV-1 S locked conformations. We compare newly determined structures to other available spike structures of Sarbecoviruses to identify conserved features and discuss their possible functions.

biochemistry↗

bioRxiv2022

CRISPR-Cas phage defense systems and prophages in Candidatus Accumulibacter

Candidatus Accumulibacter is a key genus of polyphosphate-accumulating organisms (PAOs) found in laboratory- and full-scale wastewater treatment systems, mediating enhanced biological phosphorus removal (EBPR). However, little is known about their ability to resist phage infection. We conducted a systematic analysis of the occurrence and characteristics of clustered regularly interspaced short palindromic repeats and associated proteins (CRISPR-Cas) systems and prophages in diverse Ca. Accumulibacter taxa (43 in total, including 10 newly recovered genomes). Fourty complete CRISPR loci were identified in 28 genomes, falling into 6 subtypes. The occurrence and distribution of CRISPR-Cas systems did not follow a vertical evolutionary relationship. Phylogenetic analyses of the cas genes and direct repeats (DRs) suggested that the CRISPR-Cas systems were likely acquired via horizontal gene transfer, with acquisition rates higher than those of speciation, rendering different Ca. Accumulibacter distinct adaptivity to phage predations. 2448 spacers were identified, 67 of them matched to known phages. Major differences were observed among the numbers of spacers for different Ca. Accumulibacter, showing unique phages that could be resisted by different members. A comparison of the spacers in genomes having the same cas gene but from distinct geographical locations indicated that habitat isolation may have resulted in the acquisition of different spacers by different Ca. Accumulibacter. Metagenomic analysis allowed the identification of 26 viral contigs (18 are Caudovirales members) in 6 metagenomic datasets from three lab-scale enrichment reactors, matching to 73 spacers in 10 Ca. Accumulibacter genomes in these reactors, showing the specific immunity of these Ca. Accumulibacter. Metatranscriptomic analyses showed the activity of the CRISPR-Cas system under both anaerobic and aerobic conditions. Extra efforts were made to identify prophages in Ca. Accumulibacter genomes. In total, 133 prophage regions were identified. Twenty-seven of them were classified as potentially active prophages. Three prophages (all are Caudovirales members, in DS2011, SCELSE-7IIH and SCELSE-5IIH, respectively) are readily activable. Differences in the occurrence of CRISPR-Cas systems and prophages in Ca. Accumulibacter will lead to their distinct responses under phage predation. This study represents the first systematic analysis of CRISPR-Cas systems and prophages with combined experimental and bioinformatic methods in the Ca. Accumulibacter lineage, providing new perspectives on the potential impacts of phages on Ca. Accumulibacter and EBPR systems.

bioinformatics↗

bioRxiv2022

A hybrid reconstruction of the physical model with the deep-learning that improves structured illumination microscopy

In handling raw images with low signal-to-noise (SNR) ratios, conventional algorithms of structured illumination microscopy are prone to artifacts, while deep-learning-based (DL) algorithms may lead to degradation and hallucinations. We propose a hybrid that combines the physical inversion model with a Total Deep Variation regularization. In super-resolving from low SNR images such as actin filaments, our method outperforms conventional or DL methods in suppressing artifacts and hallucinations while maintaining resolutions.

bioengineering↗

bioRxiv2022

Highly synchronized cortical circuit dynamics mediate spontaneous pain

Cortical neural dynamics mediate information processing for the cerebral cortex, implicated in fundamental biological processes, such as vision and olfaction, in addition to neurological and psychiatric diseases. Spontaneous pain is a key feature of human neuropathic pain. Whether spontaneous pain pushes cortical network into an aberrant state, and if so, whether it can be brought back to a normal operating range to ameliorate pain are unknown. Using a clinically relevant mouse model of neuropathic pain with robust spontaneous pain-like behavior, we report that orofacial spontaneous pain activated a specific area within the primary somatosensory cortex (S1), displaying synchronized neural dynamics revealed by intravital two-photon calcium imaging. This synchronization was underpinned by local GABAergic interneuron hypoactivity. Pain-induced cortical synchronization could be attenuated by manipulating local S1 networks or clinically effective pain therapies. Specifically, both chemogenetic inhibition of pain-related c-Fos-expressing neurons, and selective activation of GABAergic interneurons, significantly attenuated S1 synchronization. Clinically effective pain therapies including carbamazepine and nerve root decompression could also dampen S1 synchronization. More importantly, restoring a normal range of neural dynamics, through attenuating pain-induced S1 synchronization, alleviated pain-like behavior. These results suggest spontaneous pain pushes S1 regional network into a synchronized state, whereas reversal of this synchronization alleviates pain.

neuroscience↗

bioRxiv2022

Structure of human phagocyte NADPH oxidase in the resting state

Phagocyte oxidase plays an essential role in the first line of host defense against pathogens. It oxidizes intracellular NADPH to reduce extracellular oxygen to produce superoxide anions for pathogen killing. The resting phagocyte oxidase is a heterodimeric complex formed by two transmembrane proteins NOX2 and p22. Despite the functional importance of this complex, its structure remains elusive. Here we reported the cryo-EM structure of the human NOX2-p22 complex in nanodisc in the resting state. The structure shows that p22 is formed by four transmembrane helices and interacts with NOX2 through its M1 and M4 helices. Hydrophobic residues on M3, M4, and M5 of NOX2 contribute to the complex formation. Structural analysis suggests that the cytosolic factors activate the NOX2-p22 complex by stabilizing the dehydrogenase domain (DH) in a productive docked conformation which is efficient for electron transfer between DH and the transmembrane domain.

biochemistry↗

bioRxiv2022

TIVAN-indel: A computational framework for annotating and predicting noncoding regulatory small insertion and deletion

MotivationSmall insertion and deletion (sindel) of human genome has an important implication for human disease. One important mechanism for noncoding sindel to have an impact on human diseases and phenotypes is through the regulation of gene expression. Nevertheless, current sequencing technology may lack statistical power and resolution to pinpoint the causal sindel due to lower minor allele frequency or small effect. As an alternative solution, a supervised machine learning method can identify the otherwise missing causal sindels by predicting the regulatory potential of sindels directly. However, computational methods for annotating and predicting the regulatory sindels, especially in the noncoding regions, are underdeveloped. ResultsBy leveraging recognized sindels in cis-expression quantitative trait loci (cis-eQTLs) across 44 tissues and cell types in GTEx, and a compilation of both generic functional annotations and tissue/cell typespecific multi-omics features generated by a sequence-based deep learning model, we developed TIVAN-indel, which is an XGBoost-based supervised framework for scoring noncoding sindels based their potential to regulate the nearby gene expression. As a result, we demonstrate that TIVAN-indel achieves the best prediction performance in both cross-validation with-tissue prediction and independent cross-tissue evaluation. As an independent evaluation, we train TIVAN-indel from "Whole Blood" tissue in GTEx data and test the model using 15 immune cell types from an independent study DICE. Lastly, we perform an enrichment analysis for both recognized and predicted sindels in key regulatory regions such as chromatin interactions, open chromatin and histone modification sites, and find biologically meaningful enrichment patterns. Availability and implementationhttps://github.com/lichen-lab/TIVAN-indel Contactli.chen1@ufl.edu

bioinformatics↗

bioRxiv2022

Real-Time Spatial Registration for 3D Human Atlas

AO_SCPLOWBSTRACTC_SCPLOWThe human body is made up of about 37 trillion cells (adults). Each cell has its own unique role and is affected by its neighboring cells and environment. The NIH Human BioMolecular Atlas Program (HuBMAP) aims at developing a 3D atlas of human body consisting of organs, vessels, tissues to singe cells with all 3D spatially registered in a single 3D human atlas using tissues obtained from normal individuals across a wide range of ages. A critical step of building the atlas is to register 3D tissue blocks in real-time to the right location of a human organ, which itself consists of complex 3D substructures. The complexity of the 3D organ model, e.g., 35 meshes for a typical kidney, poses a significant computational challenge for the registration. In this paper, we propose a comprehensive framework TICKET (TIssue bloCK rEgisTration) for tissue block registration for 3D human atlas, including (1) 3D mesh pre-processing, (2) spatial queries on intersection relationship and (3) intersection volume computation between organs and tissue blocks. To minimize search space and computation cost, we develop multi-level indexing on both the anatomical structure level and mesh level, and utilize OpenMP for parallel computing. Considering cuboid based shape of the tissue block, we propose an efficient voxelization-based method to estimate the intersection volume. Our experiments demonstrate that the proposed framework is practical and efficient. TICKET is now integrated into the HuBMAP CCF registration portal [1].

bioinformatics↗

bioRxiv2022

3D Printed Skull Cap and Benchtop Fabricated Microwire-based Microelectrode Array for Custom Rat Brain Recordings

Microwire microelectrode arrays (MEAs) have been a popular low-cost tool for chronic electrophysiological recordings. Multi-MEA implantations can reveal electrical dynamics crucial to brain function. However, both the fabrication and implantation procedures for multi-MEAs on a single rodent are time-consuming and highly manual skill-dependent for quality. To enable in-house design, fabrication, and implantation of custom microwire MEAs, we developed (1) a computer-aided designed and 3D printed skull cap for pre-determined implantation locations of each MEA and (2) a benchtop fabrication approach for low-cost custom microwire MEAs. A proof-of-concept design of 32-channel 4-MEA (8-wire each) recording system was prototyped and tested through Sprague Dawley rat recordings. The skull cap design based on CT-scan of single rat conforms well with multiple Sprague Dawley rats of various size, age, and weight with minimal bregma alignment error. The prototyped 32-channel system were able to record spiking activities over 5 months. In comparison with conventional stereotactic surgeries, the skull cap system simplifies the implantation location alignment for each MEA by embedding them into the pre-printed designs, thus dramatically reducing the surgical time and effort and increasing the accuracy and repeatability. Compared to commercially available custom microwire MEAs, this in-house fabrication method enables neuroscience labs to create a custom recording apparatus with lower cost and shorter lead time for design modifications. A new methodology for neuroscience labs to fabricate and insert custom microwire MEAs has been developed and it could be easily generalized to enable low-cost highly-custom multi-region recording/stimulation studies. HighlightsO_LI3D printed skull caps as implantation platform for multi-region rat brain recording C_LIO_LIComputer-aided design for custom cap geometry and predetermined implantation location C_LIO_LIBenchtop microwire fabrication approach for in-house custom microelectrode arrays C_LIO_LIDuplicable and generalizable design, fabrication, and implantation methodologies C_LI

neuroscience↗

bioRxiv2022

Reduced Thalamic Excitation to Motor Cortical Pyramidal Tract Neurons in a Mouse Model of Parkinsonism

Degeneration of midbrain dopaminergic (DA) neurons causes a reduced motor output from the primary motor cortex (M1), underlying the motor symptoms of Parkinsons disease (PD). However, cellular and circuitry mechanisms of M1 dysfunction in PD remain undefined. Using multidisciplinary approaches, we found that DA degeneration induces cell-subtype- and inputs-specific reduction of thalamic excitation to M1 pyramidal tract (PT) neurons. Physiological and anatomical analyses suggest that DA degeneration induces a loss of thalamocortical synapses to M1 PT neurons, resulting in an impaired thalamic driving of their activities. Moreover, we showed that the decreased thalamocortical connectivity are mediated by an excessive activation of NMDA receptors of M1 PT neurons. Further, the decreased thalamocortical transmission in parkinsonism can be rescued by chemogenetically suppressing basal ganglia outputs. Together, our data suggest that the reduced motor cortical outputs in parkinsonism are not only an immediate consequence of basal ganglia inhibition but also involves specific local circuitry adaptations within M1. This study reveals novel insight in the pathophysiology of parkinsonian motor deficits.

neuroscience↗

bioRxiv2022

Microbiota-stimulated Interleukin-22 regulates brain neurons and protects against stress-induced anxiety

Psychological stress and its sequelae are a major public health problem. While the immune system has been implicated in the development of stress-related disorders, how the immune signals modulate neural responses to stress is poorly understood. Contrary to our expectations, we found that the immune cytokine Interleukin (IL)-22 is the key mediator of an immune-to-brain pathway that diminishes, rather than amplifies, stress-induced anxiety. We showed that stress induced TH17 differentiation and IL-22 production in the intestine following barrier dysfunction and microbiota stimulation. IL-22 then directly signaled to septal neurons in the brain to mitigate anxiety-like behavior. Accordingly, mice treated with exogenous IL-22 showed resilience to chronic stress-induced anxiety disorders. Our study thus reveals a previously-unappreciated immune-to-brain axis that defends against psychological stress, suggesting a potential intervention strategy for stress-related mental diseases.

immunology↗

bioRxiv2022

Specimen, Biological Structure, and Spatial Ontologies in Support of a Human Reference Atlas

The Human Reference Atlas (HRA) is defined as a comprehensive, three-dimensional (3D) atlas of all the cells in the healthy human body. It is compiled by an international team of experts that develop standard terminologies linked to 3D reference objects describing anatomical structures. The third HRA release (v1.2) covers spatial reference data and ontology annotations for 26 organs. Experts access the HRA annotations via spreadsheets and view reference models in 3D editing tools. This paper introduces the Common Coordinate Framework Ontology (CCFO) v2.0.1 that interlinks specimen, biological structure, and spatial data together with the CCF API which makes the HRA programmatically accessible and interoperable with Linked Open Data (LOD). We detail how real-world user needs and experimental data guide CCFO design and implementation, present CCFO classes and properties together with examples of their usage, and report on technical validation performed. The CCFO graph database and API are used in the HuBMAP portal, Virtual Reality Organ Gallery, and other applications that support data queries across multiple, heterogeneous sources.

bioinformatics↗

bioRxiv2022

Transcriptional changes in human palate and skin healing

Most human tissue injuries lead to the formation of a fibrous scar and result in the loss of functional tissue. One adult tissue that exhibits a more regenerative response to injury with minimal scarring is the oral mucosa. We generated a microarray gene expression dataset to examine the response to injury from human palate and skin excisional biopsies spanning the first seven days after wounding. Differential expression analyses were performed in each tissue to identify genes overexpressed or underexpressed over time compared to baseline unwounded tissue. To attribute biological processes of interest to these gene expression changes, gene set enrichment analysis was used to identify core gene sets that are enriched over the time-course of the wound healing process with respect to unwounded tissue. This analysis identified gene sets uniquely enriched in either palate or skin wounds and gene sets that are enriched in both tissues in at least one time point after injury. Finally, a cell-deconvolution analysis was performed to better understand the cell type distribution in these tissues and how it changes over the time course of wound healing. This work provides a source of human wound gene expression data that includes two tissue types with distinct regenerative and scarring phenotypes.

cell biology↗

bioRxiv2022

Early Signaling Events in Renal Compensatory Hypertrophy Revealed by Multi-Omics

Loss of a kidney results in compensatory growth of the remaining kidney, a phenomenon of considerable clinical importance. However, the mechanisms involved are largely unknown. Here, we used a multi-omic approach in a mouse unilateral nephrectomy model to identify signaling processes associated with compensatory hypertrophy of the renal proximal tubule. Morphometry applied to microdissected proximal tubules showed that growth of the proximal tubule involves a marked, rapid increase in cell volume rather than cell number. Measurements of DNA accessibility (ATAC-seq), transcriptome (RNA-seq) and proteome (quantitative protein mass spectrometry) independently identified patterns of change that are indicative of activation of the lipid-regulated transcription factor, PPAR. Activation of PPAR by fenofibrate administration increased proximal tubule cell size, while genetic deletion of PPAR in mice decreased it. The results indicate that PPAR is an important determinant of proximal tubule cell size and is a likely mediator of compensatory proximal tubule hypertrophy.

systems biology↗

bioRxiv2022

Circadian Gene Expression in Mouse Renal Proximal Tubule

Circadian variability in kidney function has long been recognized but is often ignored as a potential confounding variable in in vivo physiological experiments. To provide a guide for physiological studies on the kidney proximal tubule, we have now created a data resource consisting of expression levels for all measurable mRNA transcripts in microdissected proximal tubule segments from mice as a function of the time of day. This approach employs small-sample RNA-sequencing (RNA-seq) applied to microdissected renal proximal tubules including both S1 proximal convoluted tubules (PCTs) and S2 proximal straight tubules (PSTs). The data were analyzed using JTK-Cycle to detect periodicity. The data are provided as a user-friendly web page at https://esbl.nhlbi.nih.gov/Databases/Circadian-Prox/. In PCTs, 234 transcripts were found to vary in a circadian manner (3.7 % of total quantified). In PSTs, 334 transcripts were found to vary in a circadian manner (5.3 % of total quantified). Transcripts previously known to be associated with corticosteroid action and transcripts associated with increased flow were found to be overrepresented among circadian transcripts peaking during the "dark" portion of the day (Zeitgeber 14-22), corresponding to the peak levels of corticosterone and glomerular filtration rate in mice. BlurbCircadian variation in gene expression can be an important determinant in the regulation of kidney function. The authors used RNA-seq in microdissected proximal S1 and S2 segments to identify transcripts that vary in a circadian manner. The data were used to construct a user-friendly web resource.

systems biology↗

bioRxiv2022

Automated assembly of high-density carbon fiber electrode arrays for single unit electrophysiological recordings

ObjectiveCarbon fiber (CF) is good for chronic neural recording due to the small diameter (7 {micro}m), high Youngs modulus, and low electrical resistance, but most high-density carbon fiber (HDCF) arrays are manually assembled with labor-intensive procedures and limited by the accuracy and repeatability of the operator handling. A machine to automate the assembly is desired. ApproachThe HDCF array assembly machine contains: 1) a roller-based CF extruder, 2) a motion system with three linear and one rotary stages, 3) an imaging system with two digital microscope cameras, and 4) a laser cutter. The roller-based extruder automatically feeds single CF as raw material. The motion system aligns the CF with the array backend then places it. The imaging system observes the relative position between the CF and the backend. The laser cutter cuts off the CF. Two image processing algorithms are implemented to align the CF with the support shanks and circuit connection pads. Main resultsThe machine was capable of precisely handling 6.8 m carbon fiber electrodes. Each electrode was placed into a 12 m wide trenches in a silicon support shank. Two HDCF arrays with 16 CF electrodes populated on 3 mm shanks (with 80 m pitch) were fully assembled. Impedance measurements were found to be in good agreement with manual assembled arrays. One HDCF array was implanted in the motor cortex in an anesthetized rat and was able to detect single unit activity. SignificanceThis machine can eliminate the manual labor-intensive handling, alignment and placement of single carbon fiber during assembly, providing a proof-of-concepts towards fully automated HDCF array assembly and batch production.

bioengineering↗

bioRxiv2022

Incorporating cell hierarchy to decipher the functional diversity of single cells

Cells possess functional diversity hierarchically. However, most single-cell analyses neglect the nested structures while detecting and visualizing the functional diversity. Here, we incorporate cell hierarchy to study functional diversity at subpopulation, club (i.e., sub-subpopulation), and cell layers. Accordingly, we implement a package, SEAT, to construct cell hierarchies utilizing structure entropy by minimizing the global uncertainty in cell-cell graphs. With cell hierarchies, SEAT deciphers functional diversity in 36 datasets covering scRNA, scDNA, scATAC, and scRNA-scATAC multiome. First, SEAT finds optimal cell subpopulations with high clustering accuracy. It identifies cell types or fates from omics profiles and boosts accuracy from 0.34 to 1. Second, SEAT detects insightful functional diversity among cell clubs. The hierarchy of breast cancer cells reveals that the specific tumor cell club drives AREG-EGFT signaling. We identify a dense co-accessibility network of cis-regulatory elements specified by one cell club in GM12878. Third, the cell order from the hierarchy infers periodic pseudo-time of cells, improving accuracy from 0.79 to 0.89. Moreover, we incorporate cell hierarchy layers as prior knowledge to refine nonlinear dimension reduction, enabling us to visualize hierarchical cell layouts in low-dimensional space.

bioinformatics↗