Biology subjects

Han, X.

Publications and source records attributed to Han, X..

At least 181 records · Page 10Linked to original sources

bioRxiv2023

Reprogramming a high robust Geobacillus thermoglucosidasius for efficient synthesis of polymer-grade lactic acid under extremely high temperature (60℃)

Optically pure lactic acid is an important precursor for the synthesis of biodegradable plastic polylactic acid. Geobacillus thermoglucosidasius is a thermophilic bacterial chassis with industrial potential. In this study, we reprogrammed this chassis to efficiently produce polymer-grade lactic acid by combining rational and semi-rational design strategies. We obtained optically pure L-lactic acid- and D-lactic acid-producing strains, GTD17 and GTD7, respectively, via rational strategies: constructing a lactic acid synthesis module, deleting by-product synthesis, and enhancing lactic acid production. At an extremely high temperature (60{degrees}C), the engineered strain GTD17 produced 94.2 g L-1 of L-lactic acid with an overall yield and productivity of 91.5% and 2.0 g L-1 h-1, respectively, and the optical purity of L-lactic acid was 99.5%. We then performed semi-rational adaptive evolution on the engineered strain GTD7 to obtain an optically pure D-lactic acid-producing strain GTD7-144; at the extremely high temperature (60{degrees}C), it produced 153.1 g L-1 of D-lactic acid with an overall yield and productivity of 93.0% and 3.2 g L-1 h-1, respectively. The optical purity of the D-lactic acid was 99.6%. Genome resequencing and analysis of the strain GTD7-144 revealed potential genetic targets that may improve production performance. This study represents an advancement in the high-temperature production of polymer-grade lactic acid by G. thermoglucosidasius.

bioengineering↗

bioRxiv2023

High-level production of nervonic acid in the oleaginous yeast Yarrowia lipolytica by systematic metabolic engineering

Brain and neurological diseases are influencing more than one billion worlds people. Nervonic acid (cis-15-tetracosenoic acid, C24:1 {Delta}15) benefits the treatment of neurological diseases and the health of brain. Currently, the sources of nervonic acid are limited to the seeds of a couple of plants. In this study, we employed the oleaginous yeast Yarrowia lipolytica to overproduce nervonic acid oil by systematic metabolic engineering. First, engineering the fatty acid elongation (FAE) pathway by expressing a heterologous {beta}-ketoacyl-CoA synthase gene CgKCS enabled the production of nervonic acid in Y. lipolytica. Second, modulation of endogenous pathways by expressing a C16:0-acyl-CoA preferred fatty acid elongase gELOVL6 together with a C18:0-acyl-CoA preferred fatty acid desaturase MaOLE2 increased the content of nervonic acid in total fatty acids (TFA). Third, iterative expression of CgKCS, gELOVL6 and MaOLE2 at the genomic loci of rDNA, FAD2, TGL4, GSY1 and SNF1 dramatically improved the production of nervonic acid. Fourth, the biosynthesis of both nervonic acid and lipids were further enhanced by expression of the MaOLE2-CgKCS fusion protein and glycerol-3-phosphate acyltransferases (GPAT) and diacylglycerol acyltransferases (DGAT) from Malania oleifera in the endoplasmic reticulum (ER) membrane. Fifth, an ER structure regulator YlINO2 was identified in Y. lipolytica and the overexpression of YlINO2 led to a 39.3% increase in lipid production. Next, pilot-scale fermentation in 50-L reactor using the strain YLNA9 exhibited a lipid titer of 96.7 g/L and a nervonic acid titer of 17.3 g/L, the highest reported titer to date for de novo nervonic acid production. We also found that disruption of the AMP-activated S/T protein kinase SNF1 increased the ratio of nervonic acid (C24:1) to lignoceric acid (C24:0) by 61.6% and a ratio of 3.5:1 (nervonic acid to lignoceric acid) was achieved in the strain YLNA10. Finally, a proof-of-concept purification and separation of nervonic acid were performed and the purity of it reached 98.7%. This study suggested that oleaginous yeasts are attractive hosts for the cost-efficient production of nervonic acid and possibly other very long-chain fatty acids (VLCFAs).

bioengineering↗

bioRxiv2023

Development of membrane-like pre-stem cells after released from tube-shaped niches

We previously identified three distinct pre-stem cell lineages that further develop into blood CD34-positive stem cells, and mesenchymal stem cells. The pre-CD34-positive stem cells are spore-like, while the pre-mesenchymal stem cells are fusiform-shaped. All of them originate or released from tube-shaped tissue structures, or niches. In the current study, we present two membrane-like pre-stem progenitors. One of them had red thin membrane structures that were from disconnection of bamboo-like tissues and developed into thin membrane-like multi-nucleated cells. The other membrane-like structure had geometric-shape and was the place of developing numerous c-kit-positive stem cell progenitors. One geometric-shaped membrane could produce dozens to hundreds of progenitors in a synchronized pattern. Our findings provide more evidence that postnatal stem cells are unipotent and self-renewed in sealed tube-shaped structures. The tube-shaped structures released their inclusions that have distinct morphological differences among the cellular lineages. Thus, the self-renewal of each lineage-distinct stem cell has its unique pattern. Further, our data suggests that postnatal stem cells are renewed via a recurrent pattern. Our findings again challenged the old concepts of stem cell niche components, the origins of stem cell lineages and the self-renewal of stem cells.

cell biology↗

bioRxiv2023

Discovery and characterization of cross-reactive intrahepatic antibodies in severe alcoholic hepatitis

The pathogenesis of antibodies in severe alcoholic hepatitis (SAH) remains unknown. We sought to determine if there was antibody deposition in SAH livers and whether antibodies extracted from SAH livers were cross-reactive against both bacterial antigens and human proteins. We analyzed immunoglobulins (Ig) in explanted livers from SAH patients (n=45) undergoing liver transplantation and tissue from corresponding healthy donors (HD, n=10) and found massive deposition of IgG and IgA isotype antibodies associated with complement fragment C3d and C4d staining in ballooned hepatocytes in SAH livers. Ig extracted from SAH livers, but not patient serum exhibited hepatocyte killing efficacy in an antibody-dependent cell-mediated cytotoxicity (ADCC) assay. Employing human proteome arrays, we profiled the antibodies extracted from explanted SAH, alcoholic cirrhosis (AC), nonalcoholic steatohepatitis (NASH), primary biliary cholangitis (PBC), autoimmune hepatitis (AIH), hepatitis B virus (HBV), hepatitis C virus (HCV) and HD livers and found that antibodies of IgG and IgA isotypes were highly accumulated in SAH and recognized a unique set of human proteins as autoantigens. The use of an E. coli K12 proteome array revealed the presence of unique anti-E. coli antibodies in SAH, AC or PBC livers. Further, both Ig and E. coli captured Ig from SAH livers recognized common autoantigens enriched in several cellular components including cytosol and cytoplasm (IgG and IgA), nucleus, mitochondrion and focal adhesion (IgG). Except IgM from PBC livers, no common autoantigen was recognized by Ig and E. coli captured Ig from AC, HBV, HCV, NASH or AIH suggesting no cross-reacting anti-E. coli autoantibodies. The presence of cross-reacting anti-bacterial IgG and IgA autoantibodies in the liver may participate in the pathogenesis of SAH.

pathology↗

bioRxiv2023

SPA-STOCSY: An Automated Tool for Identification of Annotated and Non-Annotated Metabolites in High-Throughput NMR Spectra

Nuclear Magnetic Resonance (NMR) spectroscopy is widely used to analyze metabolites in biological samples, but the analysis can be cumbersome and inaccurate. Here, we present a powerful automated tool, SPA-STOCSY (Spatial Clustering Algorithm - Statistical Total Correlation Spectroscopy), which overcomes the challenges by identifying metabolites in each sample with high accuracy. As a data-driven method, SPA-STOCSY estimates all parameters from the input dataset, first investigating the covariance pattern and then calculating the optimal threshold with which to cluster data points belonging to the same structural unit, i.e. metabolite. The generated clusters are then automatically linked to a compound library to identify candidates. To assess SPA-STOCSYs efficiency and accuracy, we applied it to synthesized and real NMR data obtained from Drosophila melanogaster brains and human embryonic stem cells. In the synthesized spectra, SPA outperforms Statistical Recoupling of Variables, an existing method for clustering spectral peaks, by capturing a higher percentage of the signal regions and the close-to-zero noise regions. In the real spectra, SPA-STOCSY performs comparably to operator-based Chenomx analysis but avoids operator bias and performs the analyses in less than seven minutes of total computation time. Overall, SPA-STOCSY is a fast, accurate, and unbiased tool for untargeted analysis of metabolites in the NMR spectra. As such, it might accelerate the utilization of NMR for scientific discoveries, medical diagnostics, and patient-specific decision making.

bioinformatics↗

bioRxiv2023

Arousal state transitions occlude sensory-evoked neurovascular coupling in neonatal mice

In the adult sensory cortex, increases in neural activity elicited by sensory stimulation usually drives vasodilation mediated by neurovascular coupling. However, whether neurovascular coupling is the same in neonatal animals as adults is controversial, as both canonical and inverted responses have been observed. We investigated the nature of neurovascular coupling in unanesthetized neonatal mice using optical imaging, electrophysiology, and BOLD fMRI. We find in neonatal (postnatal day 15, P15) mice, sensory stimulation induces a small increase in blood volume/BOLD signal, often followed by a large decrease in blood volume. An examination of arousal state of the mice revealed that neonatal mice were asleep a substantial fraction of the time, and that stimulation caused the animal to awaken. As cortical blood volume is much higher during REM and NREM sleep than the awake state, awakening occludes any sensory-evoked neurovascular coupling. When neonatal mice are stimulated during an awake period, they showed relatively normal (but slowed) neurovascular coupling, showing that that the typically observed constriction is due to arousal state changes. These result show that sleep-related vascular changes dominate over any sensory-evoked changes, and hemodynamic measures need to be considered in the context of arousal state changes. Significance StatementIn the adult brain, increases in neural activity are often followed by vasodilation, allowing activity to be monitored using optical or magnetic resonance imaging. However, in neonates, sensory stimulation can drive vasoconstriction, whose origin was not understood. We used optical and magnetic resonance imaging approaches to investigate hemodynamics in neonatal mice. We found that sensory-induced vasoconstriction occurred when the mice were asleep, as sleep is associated with dilation of the vasculature of the brain relative to the awake state. The stimulus awakens the mice, causing a constriction due to the arousal state change. Our study shows the importance of monitoring arousal state, particularly when investigating subjects that may sleep, and the dominance arousal effects on brain hemodynamics.

neuroscience↗

bioRxiv2023

Distinct regulatory networks control toxin gene expression in elapid and viperid snakes

Venom systems are ideal models to study genetic regulatory mechanisms that underpin evolutionary novelty. Snake venom glands are thought to share a common origin, but there are major distinctions between venom toxins from the medically significant snake families Elapidae and Viperidae, and toxin gene regulation in elapids is largely unexplored. Here, we used high-throughput RNA-sequencing to profile gene expression and microRNAs between active (milked) and resting (unmilked) venom glands in an elapid (Eastern Brown Snake, Pseudonaja textilis), in addition to comparative genomics, to identify cis- and trans- acting regulation of venom production in an elapid in comparison to viperids (Crotalus viridis and C. tigris). Although there is conservation in high-level mechanistic pathways regulating venom production, there are histone methylation, transcription factor, and microRNA regulatory differences between these two snake families. Histone methyltransferases (KMT2A, KMT2C and KMT2D) and transcription factor (TF) specificity protein 1 (Sp1) were highly upregulated in the milked elapid venom gland, whereas nuclear factor I (NFI) TFs were upregulated after viperid venom milking. Sp1 and NFI cis-regulatory elements were common to toxin gene promoter regions, but many unique elements were also present between elapid and viperid toxins. microRNA profiles were distinctive between milked and unmilked venom glands for both snake families, and microRNAs were predicted to target different toxin transcripts. Our comparative transcriptomic and genomic analyses between toxin genes and isoforms in elapid and viperid snakes suggests independent toxin evolution between these two snake families, demonstrating multiple toxin genes and regulatory mechanisms converged to underpin a highly venomous phenotype.

evolutionary biology↗

bioRxiv2023

The self-renewal procedures of mesenchymal stem cells in the blood

BackgroundAlthough mesenchymal stem cells (MSCs) are most commonly used in cell therapy and stem cell research, the mechanism and the locations of their self-renewal are still unknown. MethodMouse blood was collected, and examined under microscopy. The results were compared with the data of human umbilical cord blood (hUCB) collected 10 years ago. ResultsWe found that the procedure of self-renewal for the mesenchymal stem cells in mouse blood and hUCB needs at least 5 steps. First, tube-shaped stem cell niches release long segmented materials composed of sand-like particles and semitransparent granules. Second, the sand-like particles and semitransparent granules separate from the segmented materials. Third, each of the individual semitransparent granules releases groups of fusiform-shaped structures that do not stain to H&E. The sizes of the fusiform-shaped structures range from 1 to 100 m in length in mouse blood, but can be 200 m in hUCB. Fourth, the large-sized fusiform structures can directly transform into lineage-restricted cellular structures; the medium-sized fusiform structures fuse or engulf each other to form cellular structures. The cellular structures further acquire membranes from the adjacent nucleated cells. Fifth, the nucleolus appears in the new cellular structures before the nucleus. During all the procedures, the adjacent nucleated mesenchymal cells are must needed. Thus, these newly formed cellular structures will further differentiate into nucleated mesenchymal stem cells. ConclusionOur findings again provide new evidence that, in physiological conditions, mesenchymal stem cell self-renewal needs several steps to complete, which, however, does not occur by mitotic division. The tube-shaped structures are the niches of the stem cells.

cell biology↗

bioRxiv2023

Deep Learning-based Identification of Intraocular Pressure-Associated Genes Influencing Trabecular Meshwork Cell and Organelle Morphology

PURPOSEThe exact pathogenesis of primary open-angle glaucoma (POAG) is poorly understood. Genome-wide association studies (GWAS) have recently uncovered many loci associated with variation in intraocular pressure (IOP); a crucial risk factor for POAG. Artificial intelligence (AI) can be used to interrogate the effect of specific genetic knockouts on the morphology of trabecular meshwork cells (TMCs), the regulatory cells of IOP. METHODSSixty-two genes at fifty-five loci associated with IOP variation were knocked out in primary TMC lines. All cells underwent high-throughput microscopy imaging after being stained with a five-channel fluorescent cell staining protocol. A convolutional neural network (CNN) was trained to distinguish between gene knockout and normal control cell images. The area under the receiver operator curve (AUC) metric was used to quantify morphological variation in gene knockouts to identify potential pathological perturbations. RESULTSCells where RALGPS1 had been perturbed demonstrated the greatest morphological variation from normal TMCs (AUC 0.851, SD 0.030), followed by LTBP2 (AUC 0.846, SD 0.029) and BCAS3 (AUC 0.845, SD 0.020). Of seven multi-gene loci, five had statistically significant differences in AUC (p<0.05) between genes, allowing for pathological gene prioritisation. The mitochondrial channel most frequently showed the greatest degree of morphological variation (33.9% of cell lines). CONCLUSIONSWe demonstrate a robust method for functionally interrogating genome-wide association signals using high-throughput microscopy and AI. Genetic variations inducing marked morphological variation can be readily identified, allowing for the gene-based dissection of loci associated with complex traits.

genetics↗

bioRxiv2023

Multi-omics Comparison among Populations of Three Plant Sources of Amomi Fructus

Amomi Fructus (Sharen, AF) is a traditional Chinese medicine (TCM) from three source species (or subspecies) including Wurfbainia villosa var. villosa (WVV), W. villosa var. xanthioides (WVX) or W. longiligularis (WL). Among them, WVV has been transplanted from its top-geoherb region Guangdong to its current main production area Yunnan for more than 50 years in China. However, the genetic and transcriptomic differentiation among multiple AF source (sub)species and between the origin and transplanted populations of WVV is unknown. In our study, the observed overall higher expression of terpenoid biosynthesis genes in WVV than that of WVX supplied possible evidence for the better pharmacological effect of WVV. We also screened ten candidate borneol dehydrogenase (BDH) genes that potentially catalyzed borneol into camphor in WVV. The BDH genes may experience independent evolution after acquiring the ancestral copies and the followed tandem duplications might account for the abundant camphor content in WVV. Furthermore, four populations of WVV, WVX and WL are genetically differentiated and the gene flow from WVX to WVV in Yunnan contributed to the increased genetic diversity in the introduced population (WVV-JH) compared to its top-geoherb region (WVV-YC), which showed the lowest genetic diversity and might undergo genetic degradation. In addition, TPS and BDH genes were selected among populations of multiple AF source (sub)species and between the top-geoherb and non-top-geoherb regions, which might explain the metabolite difference of these populations. Our findings provide important guidance for the conservation, genetic improvement, industrial development of the three source (sub)species, and identifying top-geoherbalism with molecular markers and proper clinical application of AF.

evolutionary biology↗

bioRxiv2023

Wingless signaling promotes lipid mobilization through signal-induced transcriptional repression

Conserved Wnt/Wingless signaling plays pivotal roles in regulating normal development and energy metabolism in metazoans, and aberrant activation of Wnt signaling drives the pathogenesis of many diseases including cancer. However, the role of Wnt signaling in regulating cellular lipid homeostasis, particularly lipid mobilization, remains poorly understood. Here we show that canonical Wg signaling inhibits lipid accumulation in Drosophila larval adipocytes by stimulating lipid catabolism while simultaneously inhibiting lipogenesis. Using a combination of RNA-sequencing and CUT&RUN assays, we identified a battery of Wg target genes encoding key factors required for lipogenesis (such as FASN1 and AcCoS), lipolysis (such as lipid droplet-associated proteins Lsd-1 and Lsd-2), and fatty acid {beta}-oxidation in the mitochondria and peroxisome (e.g., CPT1 and CRAT), most of which are directly repressed by active Wg signaling. Furthermore, lipid accumulation defects caused by active Wg signaling are rescued by either ectopically expressing Lsd-1 and Lsd-2 or depleting the transcriptional repressor Aef1, whose binding motif was identified in 52% of Wg signaling-repressed genes. These findings suggest that active Wg signaling reduces intracellular lipid accumulation by inhibiting lipogenesis and fatty acid {beta}-oxidation and by promoting lipolysis and lipid mobilization, and Wg signaling-induced transcriptional repression play a prominent role in these converging mechanisms.

cell biology↗

bioRxiv2022

Disrupting central carbon metabolism increases antibiotic susceptibility in Vibrio cholerae

Antibiotic tolerance, the ability of bacteria to sustain viability in the presence of typically bactericidal antibiotics for extended time periods, is an understudied contributor to treatment failure. The Gram-negative pathogen Vibrio cholerae, the causative agent of cholera disease, becomes highly tolerant to {beta}-lactam antibiotics (penicillin and related compounds) in a process requiring the two-component system VxrAB. VxrAB is induced by exposure to cell wall damage conditions, which results in the differential regulation of >100 genes. While the effectors of VxrAB are relatively well-known, VxrAB environment-sensing and activation mechanisms remain a mystery. Here, we used transposon mutagenesis to screen for mutants that spontaneously upregulate VxrAB signaling. This screen was answered by genes known to be required for proper cell envelope homeostasis, validating the approach. Unexpectedly, we also uncovered a new connection between central carbon metabolism and antibiotic tolerance. Inactivation of pgi (vc0374, coding for Glucose-6-phosphate isomerase) resulted in an intracellular accumulation of glucose-6-phosphate and fructose-6-phosphate, concomitant with a marked cell envelope defect, resulting in VxrAB induction. Deletion of pgi also increased sensitivity to {beta}-lactams and conferred a growth defect on salt-free LB; phenotypes that could be suppressed by deleting sugar uptake systems and by supplementing cell wall precursors in the growth medium. Our data suggest an important connection between central metabolism and cell envelope integrity and highlight a potential new target for developing novel antimicrobial agents. ImportanceAntibiotic tolerance (the ability to survive exposure to antibiotics) is a stepping-stone towards antibiotic resistance (the ability to grow in the presence of antibiotics), an increasingly common cause of antibiotic treatment failure. The mechanisms promoting tolerance are poorly understood. Herein, we discovered central carbon metabolism as a key contributor to antibiotic tolerance and resistance. A mutant in a sugar utilization pathway accumulates metabolites that likely shut down the synthesis of cell wall precursors, which weakens the cell wall and thus increases susceptibility to cell wall-active drugs. Our results illuminate the connection between central carbon metabolism and cell wall homeostasis in V. cholerae and suggest that interfering with metabolism may be a fruitful future strategy for development of antibiotic adjuvants.

microbiology↗

bioRxiv2022

Integrated proteomics reveals the landscape of autophagy degradation in human neurons and autophagy receptors regulating neuronal activity

Autophagy is a catabolic and self-degradative process crucial for maintaining cellular homeostasis. Malfunctional autophagy is implicated in neurodevelopmental and neurodegenerative diseases. However, the exact role and targets of autophagy in human neurons remain elusive. Here we reported a systematic investigation of neuronal autophagy targets through integrated proteomics. Deep proteomic profiling of multiple autophagy-deficient lines of human induced neurons, mouse brains, and LC3-interactome uncovers a role of neuronal autophagy in targeting primarily endoplasmic reticulum (ER), mitochondria, endosome, Golgi apparatus, synaptic vesicle (SV) proteins, and cAMP-PKA pathway, for degradation. Tubular ER and specific SV proteins are significant autophagy cargos in the axons. Functional validation identified calumenin as an ER resident autophagy receptor and illuminated a role of autophagy in regulating PKA and neuronal activity through AKAP11-mediated degradation. Our study thus reveals the landscape of autophagy degradation in human neurons and offers molecular insight into mechanisms of neurological disorders linked to autophagy deficiency. HighlightO_LIIntegrated proteomics reveals the landscape of autophagy degradation in human neurons C_LIO_LIAutophagy clears tubular ER and selective ER and synaptic vesicle proteins in neurons C_LIO_LICalumenin is an ER resident autophagy receptor C_LIO_LIAutophagy controls PKA pathway and neuronal activity through autophagy receptor AKAP11 C_LI

neuroscience↗

bioRxiv2022

Membrane Voltage Dynamics of Parvalbumin Interneurons Orchestrate Hippocampal Theta Rhythmicity

Hippocampal network activity at theta frequencies (5-10Hz) is important for behavior. However, it remains unclear how behaviorally-relevant network theta rhythms arise and interact with cellular dynamics to dictate spike timing. We performed membrane voltage (Vm) imaging of individual CA1 pyramidal cells and parvalbumin interneurons with simultaneous local field potential (LFP) recordings in mice during locomotion. We found that Vm theta rhythms organize spike timing in both cell types regardless of behavioral conditions, but the Vm of parvalbumin interneurons is better synchronized with LFP. The temporal relationships between spikes and LFP theta reliably reflect the Vm-LFP relationships in parvalbumin cells, but not in pyramidal cells. Thus, cellular theta rhythms broadly organize spike timing in CA1 neurons, and parvalbumin interneurons are critical in coordinating network theta rhythms. One-Sentence SummaryCellular membrane voltage of parvalbumin interneurons organizes spiking and network dynamics in the hippocampus.

neuroscience↗

bioRxiv2022

An in vitro microfluidic culture device for peripheral neurotoxicity prediction at low concentration based on deep learning

In this study, a microfluidic culture device was developed including related evaluation methods using deep learning for the purpose of constructing a rapid assessment platform for peripheral neuropathy caused by compounds. Primary rodent dorsal root ganglion was cultured in the microfluidic culture device that could separate the cell body and neurites, and the neurites morphological changes were analyzed by immunofluorescence images. Separated neurites successful culture in the microfluidic device for more than 1 month indicated that a series of test processes from culture to drug stimulation and fluorescence observation is possible. Additionally, cultured samples were treated with several anticancer drugs known to cause peripheral neurotoxicity (i.e., vincristine, oxaliplatin, and paclitaxel) and analyzed the neurites morphological changes by deep learning for image analysis. After training, artificial intelligence (AI) could identify neurite morphological changes caused by each compound and precisely predicted the toxicity positivity, even at low concentrations. For testing compounds, AI could also precisely detect toxicity negative and positive based on neurite images, even at low concentrations. Therefore, this microfluidic culture system is supposed to be useful for in vitro toxicity assessment.

pharmacology and toxicology↗

bioRxiv2022

Large-area electrical imaging having single neuron resolution using 236,880 electrodes CMOS-MEA technology

The electrophysiological technology having a high spatio-temporal resolution at the single-cell level, and noninvasive measurements of large areas provides insights on underlying neuronal function. Here, we used a complementary metal-oxide semiconductor (CMOS)-microelectrode array (MEA) that uses 236,880 electrodes each with an electrode size of 11.22 x 11.22 {micro}m and 236,880 covering a wide area of 5.5 x 5.7 mm in presenting a detailed and single-cell-level neural activity analysis platform for brain slices, human iPS cell-derived cortical networks, peripheral neurons, and human brain organoids. Propagation pattern characteristics between brain regions changes the synaptic strength into compounds based on single-cell time-series patterns, classification based on single DRG neuron firing patterns and compound responses, axonal conduction characteristics and changes to anticancer drugs, and network activities and transition to compounds in brain organoids were extracted. This detailed analysis of neural activity at the single-cell level using our CMOS-MEA provides a new understanding the basic mechanisms of brain circuits in vitro and ex vivo, on human neurological diseases for drug discovery, and compound toxicity assessment.

neuroscience↗

bioRxiv2022

Bio-friendly long-term subcellular dynamic recording by self-supervised image enhancement microscopy

Fluorescence microscopy has become an indispensable tool for revealing the dynamic regulations of cells and organelles in high resolution noninvasively. However, stochastic noise inherently restricts the upper bonds of optical interrogation quality and exacerbates the observation fidelity in encountering joint demand of high frame rate, long-term, and low photobleaching and phototoxicity. Here, we propose DeepSeMi, a self-supervised-learning-based denoising framework capable of increasing SNR by over 12 dB across various conditions. With the introduction of newly designed eccentric blind-spot convolution filters, DeepSeMi accomplished efficacious denoising requiring no clean data as references and no compromise of spatiotemporal resolution on diverse imaging systems. The computationally 15-fold multiplied photon budget in a standard confocal microscope by DeepSeMi allows for recording organelle interactions in four colors and high-frame-rate across tens of thousands of frames, monitoring migrasomes and retractosomes over a half day, and imaging ultra-phototoxicity-sensitive Dictyostelium cells over thousands of frames, all faithfully and sample-friendly. Through comprehensive validations across various cells and species over various instruments, we prove DeepSeMi is a versatile tool for reliably and bio-friendly breaking the shot-noise limit, facilitating automated analysis of massive data about cell migrations and organelle interactions.

bioengineering↗

bioRxiv2022

Whole Human-Brain Mapping of Single Cortical Neurons for Profiling Morphological Diversity and Stereotypy

Quantification of individual cells morphology and their distribution at the whole brain scale is essential to understand the structure and diversity of cell types. Despite recent technological advances, especially single cell labeling and whole brain imaging, for many prevailing animal models, it is exceedingly challenging to reuse similar technologies to study human brains. Here we propose Adaptive Cell Tomography (ACTomography), a low-cost, high-throughput, high-efficacy tomography approach, based on adaptive targeting of individual cells suitable for human-brain scale modeling of single neurons to characterize their 3-D structures, statistical distributions, and extensible for other cellular features. Specifically, we established a platform to inject dyes into cortical neurons in surgical tissues of 18 patients with brain tumors or other conditions and 1 donated fresh postmortem brain. We collected 3-D images of 1746 cortical neurons, of which 852 neurons were subsequentially reconstructed to quantify their local dendritic morphology, and mapped to standard atlases both computationally and semantically. In our data, human neurons are more diverse across brain regions than by subject age or gender. The strong stereotypy within cohorts of brain regions allows generating a statistical tensor-field of neuron morphology to characterize 3-D anatomical modularity of a human brain.

neuroscience↗