Biology subjects

Ma, H.

Publications and source records attributed to Ma, H..

At least 217 records · Page 12Linked to original sources

bioRxiv2020

Tocilizumab treatment in severe COVID-19 patients attenuates the inflammatory storm incited by monocyte centric immune interactions revealed by single-cell analysis

ABSTRACTDespite the current devastation of the COVID-19 pandemic, several recent studies have suggested that the immunosuppressive drug Tocilizumab can powerfully treating inflammatory responses that occur in this disease. Here, by employing single-cell analysis of the immune cell composition of severe-stage COVID-19 patients and these same patients in post Tocilizumab-treatment remission, we have identified a monocyte subpopulation specific to severe disease that contributes to inflammatory storms in COVID-19 patients. Although Tocilizumab treatment attenuated the strong inflammatory immune response, we found that immune cells including plasma B cells and CD8+ T cells still exhibited an intense humoral and cell-mediated anti-virus immune response in COVID-19 patients after Tocilizumab treatment. Thus, in addition to providing a rich, very high-resolution data resource about the immune cell distribution at multiple stages of the COVID-19 disease, our work both helps explain Tocilizumab’s powerful therapeutic effects and defines a large number of potential new drug targets related to inflammatory storms.Competing Interest StatementJingwen Fang is the executive officer of HanGen BiotechView Full Text

genomics↗

bioRxiv2020

Selective Cell Propagation via Micropatterning of Thermal-activated Hydrogel

The ability to selectively propagate specific cells is fundamentally important to the development of clonal cell populations. Current methods rely on techniques such as limiting dilution, colony picking, and flow cytometry to transfer single cells into single wells, resulting in workflows that are low-throughput, slowed by propagation kinetics, and susceptible to contamination. Here, we developed a method, called selective laser gelation (SLG), to micropattern hydrogels in cell culture media in order to encapsulate specific cells to selectively arrest their growth. This process relies on the inverse gelation of methylcellulose, which forms a hydrogel when heated rather than cooled. Local heating using an infrared laser enables hydrogel micropatterning, while phase transition hysteresis retains the hydrogel after laser excitation. As a demonstration, we used this approach to selectively propagate transgenic CHO cells with increased antibody productivity. More generally, hydrogel micropatterning provides a simple and non-contact method to selective propagation of cells based on features identified by imaging. One Sentence SummaryInverse gelation of methylcellulose enables hydrogel micropatterning to selectively propagate cells identified by microscopy.

bioengineering↗

bioRxiv2020

Chromosome-Scale Genome Assembly Provides Insights into Speciation of Allotetraploid and Massive Biomass Accumulation of Elephant Grass (Pennisetum purpureum Schum.)

Elephant grass (Pennisetum purpureum Schum., AABB, 2n=4x=28), which is characterized as robust growth and high biomass, and widely distributed in tropical and subtropical areas globally, is an important forage, biofuels and industrial plant. We sequenced its allopolyploid genome and assembled 2.07 Gb (96.88%) into A and B sub-genomes of 14 chromosomes with scaffold N50 of 8.47 Mb. A total of 38,453 and 36,981 genes were annotated in A and B sub-genomes, respectively. A phylogenetic analysis with species in Pennisetum identified that the speciation of the allotetraploid occurred approximately 15 MYA after the divergence between S.italica and P. glaucum. Double whole-genome duplication (WGD) and polyploidization events resulted in large scale gene expansion, especially in the key steps of growth and biomass accumulation. Integrated transcriptome profiling revealed the functional differentiation between sub-genomes; A sub-genome contributed more to plant growth, development and photosynthesis whereas B sub-genome primarily offered functions of effective transportation and resistance to stimulation. The results uncovered enhanced cellulose and lignin biosynthesis pathways with 645 and 666 genes expanded in A and B sub-genomes, respectively. Our findings provided deep insights into the speciation and genetic basis of fast growth and high biomass accumulation in the species. The genetic, genomic, and transcriptomic resources generated in this study will pave the way for further domestication and selection of these economical species and making them more adaptive to industrial utilization.

genomics↗

bioRxiv2020

Lossless Immunocytochemistry using Photo-polymerized Hydrogel Thin-films

Immunocytochemistry (ICC), or immunofluorescence microscopy, is an essential biological technique for phenotyping cells in both research and diagnostic applications. Standard ICC methods often do not work well when the cell sample contains a small number of cells (<10,000) because of the significant cell loss that occurs during washing, staining, and centrifugation steps. Cell loss is particularly relevant when working with rare cells, such as circulating tumor cells, where such losses could significantly bias experimental outcomes. In order to eliminate cell loss in ICC protocols, we present a method to encapsulate the cell sample in a photo-polymerized hydrogel thin-film. The hydrogel thin-film is permeable to antibodies and other ICC reagents, thereby allowing the use of standard ICC protocols without modification. The cell sample is physically constrained by the hydrogel at the bottom surface of a standard (unmodified) imaging microtiter plate, thereby enabling the acquisition of high-quality micrographs regardless of the properties of the cell sample or staining reagents. Furthermore, while standard ICC requires several centrifugation steps during staining and washing, our hydrogel encapsulation method requires only a single centrifugation step. This property greatly reduces the time required to perform ICC protocols and is more compatible with robotic platforms. In this study, we show that standard ICC and Cytospin protocols are extremely lossy (>70% loss) when the sample contains less than 10,000 cells, while encapsulating the cells using a permeable hydrogel thin-film results in a lossless ICC process.

bioengineering↗

bioRxiv2020

CRISPR-based Live Imaging of Epigenetic Modification-Mediated Genome Reorganization

Epigenetic modifications play an essential role in chromatin architecture and dynamics. The role of epigenetic modification in chromatin organization has been studied by Hi-C from population cells, but imaging techniques to study their correlation and regulation in single living cells are lacking. Here we develop a CRISPR-based EpiGo (Epigenetic perturbation induced Genome organization) system to track epigenetic modification-mediated relocation, interaction or reorganization of genomic regions in living cells. EpiGo-KRAB is sufficient to induce the relocation of genomic loci to HP1 condensates and trigger genomic interactions. EpiGo-KRAB also triggers the induction of H3K9me3 at large genomic regions, which decorate on the surface of HP1 condensates possibly driven by phase separation.

molecular biology↗

bioRxiv2020

Recombinant human MG53 protein preserves mitochondria integrity in cardiomyocytes during ischemia reperfusion-induced oxidative stress

Ischemic injury to the heart causes a loss of mitochondria function due to an increase in oxidative stress. MG53, also known as TRIM72, is highly expressed in striated muscle and is essential to repair damage to plasma membrane. We have shown that mg53-/- mice are more susceptible to ischemia-reperfusion injury, whereas treatment with exogenous recombinant human MG53 (rhMG53) reduces both infarct damage and restores cardiac function. This study assesses whether MG53 protects and repairs mitochondria injury after oxidative stress associated with myocardial infarction. We hypothesize that in addition to known cell membrane repair function, MG53 acts as a myokine to protect cardiomyocytes by maintaining mitochondrial function. A combination of in vivo and in vitro ischemia/reperfusion models were used to assess MG53s effect on mitochondria using biochemical assays and confocal microscopic imaging. Treatment with rhMG53 allowed cells to maintain a healthy mitochondrial membrane potential, reduced release of mitochondrial reactive oxygen species, and mitigated mitophagy. Mitochondrial localization of rhMG53 is mediated by exposure of and interaction with cardiolipin on the mitochondrial membrane. Our data demonstrates that rhMG53 protein preserves mitochondria integrity in cardiomyocytes during ischemia reperfusion-induced oxidative stress.

physiology↗

bioRxiv2020

Characterization of a novel NDM-5-harboring plasmid from a carbapenem-resistant Escherichia coli isolate from China

A carbapenem-resistant Escherichia coli (sequence type 5415) strain was isolated from a male patient. S1-PFGE and Southern blotting showed that the blaNDM-5 gene was located on a novel 66-kb IncFII [F2:A-:B-] plasmid. Conjugation assays revealed that the blaNDM-5-bearing plasmid was self-transferrable. Genomic sequencing and comparative analysis suggested that plasmid p2947-NDM5 likely originated from a combination of an IncFII-type backbone and the blaNDM-5 flanking genetic elements. This study highlights the genetic complexity of an NDM-5 carbapenemase and the urgent need for continuous active monitoring. IMPORTANCECarbapenem resistance due to the NDM enzyme has become a serious clinical problem in medical care and public health. We here identified a blaNDM-5-positive E. coli strain ST5414. The blaNDM-5 gene was located on a novel self-transferrable IncFII type plasmid p2947-NDM5. Genomic sequencing and comparative analysis suggested that plasmid p2947-NDM5 likely originated from a combination of an IncFII-type backbone and the blaNDM-5 flanking genetic elements. The genetic context of NDM-5 is highly similar to that of animal origins, and we speculate that the blaNDM-harboring IncFII plasmid has a potential risk of being transmitted from animals to humans and should be paid more attention. Our findings highlight the threat of NDM-5 carbapenemase circulation and the urgency of stringent surveillance and control measures.

genomics↗

bioRxiv2020

Alterations of human lung and gut microbiome in non-small cell lung carcinomas and distant metastasis

BackgroundNon-small cell lung cancer (NSCLC) is the leading cause of cancer-related deaths worldwide. Although dysbiosis of lung and gut microbiota have been associated with NSCLC, their relative contributions are unclear; in addition, their roles in distant metastasis (DM) are still illusive. ResultsWe surveyed the fecal and sputum (as a proxy for lung) microbiota in healthy controls and NSCLC patients of various stages, and found significant perturbations of gut- and sputum-microbiota in patients with NSCLC and DM. Machine-learning models combining both microbiota (mixed models) performed better than either dataset in patient stratification, with the highest area under the curve (AUC) value of 0.842. Sputum-microbiota contributed more than the gut in the mixed models; in addition, sputum-only models performed similarly to the mixed models in most cases. Several microbial-biomarkers were shared by both microbiota, indicating their similar roles at distinct body sites. Microbial-biomarkers of distinct disease stages were mostly shared, suggesting biomarkers for distant metastasis could be acquired early. Furthermore, Pseudomonas aeruginosa, a species previously associated with wound infections, was significantly more abundant in brain metastasis, indicating distinct types of DMs could have different microbial-biomarkers. ConclusionOur results indicate that alterations of sputum-microbiota have stronger relationships with NSCLC and distant metastasis than the gut, and strongly support the feasibility of metagenome-based non-invasive disease diagnosis and risk evaluation.

microbiology↗

bioRxiv2019

Visual Analytics for Deep Embeddings of Large Scale Molecular Dynamics Simulations

Molecular Dynamics (MD) simulation have been emerging as an excellent candidate for understanding complex atomic and molecular scale mechanism of bio-molecules that control essential bio-physical phenomenon in a living organism. But this MD technique produces large-size and long-timescale data that are inherently high-dimensional and occupies many terabytes of data. Processing this immense amount of data in a meaningful way is becoming increasingly difficult. Therefore, specific dimensionality reduction algorithm using deep learning technique has been employed here to embed the high-dimensional data in a lower-dimension latent space that still preserves the inherent molecular characteristics i.e. retains biologically meaningful information. Subsequently, the results of the embedding models are visualized for model evaluation and analysis of the extracted underlying features. However, most of the existing visualizations for embeddings have limitations in evaluating the embedding models and understanding the complex simulation data. We propose an interactive visual analytics system for embeddings of MD simulations to not only evaluate and explain an embedding model but also analyze various characteristics of the simulations. Our system enables exploration and discovery of meaningful and semantic embedding results and supports the understanding and evaluation of results by the quantitatively described features of the MD simulations (even without specific labels).

bioinformatics↗

bioRxiv2019

Deformability based sorting of stored red blood cells reveals donor-dependent aging curves

A fundamental challenge in the transfusion of red blood cells (RBCs) is that a subset of donated RBC units may not provide optimal benefit to transfusion recipients. This variability stems from the inherent ability of donor RBCs to withstand the physical and chemical insults of cold storage, which ultimately dictate their survival in circulation. The loss of RBC deformability during cold storage is well-established and has been identified as a potential biomarker for the quality of donated RBCs. While RBC deformability has traditionally been indirectly inferred from rheological characteristics of the bulk suspension, there has been considerable interest in directly measuring the deformation of RBCs. Microfluidic technologies have enabled single cell measurement of RBC deformation but have not been able to consistently distinguish differences between RBCs between healthy donors. Using the microfluidic ratchet mechanism, we developed a method to sensitively and consistently analyze RBC deformability. We found that the aging curve of RBC deformability varies significantly across donors, but is consistent for each donor over multiple donations. Specifically, certain donors seem capable of providing RBCs that maintain their deformability during two weeks of cold storage in standard test tubes. The ability to distinguish between RBC units with different storage potential could provide a valuable opportunity to identify donors capable of providing RBCs that maintain their integrity, in order to reserve these units for sensitive transfusion recipients.

bioengineering↗

bioRxiv2019

Image-based Cell Phenotyping Using Deep Learning

The ability to phenotype cells is fundamentally important in biological research and medicine. Current methods rely primarily on fluorescence labeling of specific markers. However, there are many situations where this approach is unavailable or undesirable. Machine learning has been used for image cytometry but has been limited by cell agglomeration and it is unclear if this approach can reliably phenotype cells indistinguishable to the human eye. Here, we show disaggregated single cells can be phenotyped with a high degree of accuracy using low-resolution bright-field and non-specific fluorescence images of the nucleus, cytoplasm, and cytoskeleton. Specifically, we trained a convolutional neural network using automatically segmented images of cells from eight standard cancer cell-lines. These cells could be identified with an average classification accuracy of 94.6%, tested using separately acquired images. Our results demonstrate the potential to develop an \"electronic eye\" to phenotype cells directly from microscopy images indistinguishable to the human eye.

bioinformatics↗

bioRxiv2019

Direct, sensitive and specific detection of individual single- or double-strand DNA breaks by fluorescence microscopy

We here describe a technique termed STRIDE (SensiTive Recognition of Individual DNA Ends), which enables highly sensitive, specific, direct in situ detection of single- or double-strand DNA breaks (sSTRIDE or dSTRIDE), in nuclei of single cells, using fluorescence microscopy. Sensitivity of STRIDE was tested using specially developed CRISPR/Cas9 DNA damage induction system, capable of inducing small clusters or individual single- or double-strand breaks. STRIDE exhibits significantly higher sensitivity and specificity of detection of DNA breaks than the commonly used TUNEL assay or methods based on monitoring of recruitment of repair proteins or histone modifications at the damage site (e.g. {gamma}H2AX). Even individual genome site-specific DNA double-strand cuts induced by CRISPR/Cas9, as well as individual single-strand DNA scissions induced by the nickase version of Cas9, can be detected by STRIDE and precisely localized within the cell nucleus. We further show that STRIDE can detect low-level spontaneous DNA damage, including age-related DNA lesions, DNA breaks induced by several agents (bleomycin, doxorubicin, topotecan, hydrogen peroxide, UV, photosensitized reactions), and fragmentation of DNA in human spermatozoa. STRIDE methods are potentially useful in studies of mechanisms of DNA damage induction and repair in cell lines and primary cultures, including cells with impaired repair mechanisms.

cell biology↗

bioRxiv2019

Epitope containing short peptides capture distinct IgG serodynamics that enable DIVA for live-attenuated vaccines

Differentiating infected from vaccinated animals (DIVA) strategies have been central enabling techniques in several successful viral disease elimination programs. However, owing to their long and uncertain development process, no DIVA-compatible vaccines are available for many important diseases. We report herein a new DIVA strategy based on hybrid protein-peptide microarrays which can theoretically work with any vaccine. Leading from our findings from Peste des petits ruminants (PPR), we found 4 epitope containing short peptides (ECSPs) which have distinct IgG serodynamics: anti-ECSP IgGs only exist for 10-60 days post vaccination (dpv), while anti-protein IgGs remained at high levels for >1000 dpv. These data enabled design of a DIVA diagnostic microarray containing 4 ECSPs and 3 proteins, which unlike cELISA and VNT, enables ongoing monitoring of serological differences between vaccinated individuals and individuals exposed to the pathogen. For 50 samples after 60 dpv, 20 animals were detected with positive anti-ECSP IgGs, indicating recent infections in vaccinated goat/sheep herds. These DIVA diagnostic microarrays will almost certainly facilitate eradication programs for (re-)emerging pathogens and zoonoses.

immunology↗

bioRxiv2019

Binding to m6A RNA promotes YTHDF2-mediated phase separation

As the most abundant modification on mRNA in mammal, N6-Methyladenosine (m6A) has been demonstrated to play important roles in various biological processes including mRNA splicing, translation and degradation. m6A reader proteins have been shown to play central roles in these processes. One of the m6A readers, YTHDF2 is localized to the P granules, which are liquid-like droplets where RNA degradation occurs. How YTHDF2 is localized to P granules is unknown. Here we provide evidence that YTHDF2 forms liquid droplets and phase separate, mediated by its low complexity (LC) domains. Interestingly, the ability to phase separate is robustly stimulated by m6A RNAs in vitro. In vivo, YTHDF2 phase separation may in fact be dependent on m6A RNA and YTHDF2 binding to m6A RNA, since a YTHDF2 m6A-binding defective mutant or a wildtype YTHDF2 assayed in cells lacking m6A RNAs, both fail to phase separate. The ability of phase separate is not limited to YTHDF2; we find other members of the YTH-domain m6A readers can also undergo phase separation. Our findings suggest that m6A RNA induced phase separation of m6A readers may play an important role in their distributions to different phase-separated compartments in cells.

molecular biology↗

bioRxiv2019

Use of extracellular polymer substances as additives for improving biogas yield and digestion performance

To understand how extracellular polymer substances (EPS) as additives promotes methanogenesis, batch tests of methane production potential in anaerobic reactors with the addition of EPS or not were conducted. Research showed that EPS increased remarkably methane production during anaerobic digestion (36.5% increase compared with the control). EPS enriched functional microorganisms such as Firmicutes, Actinobacteria, Synergistetes, and Chloroflexi. Among them, 8.86% OTUs from the important hydrolysis and acidification phyla, which may be an important reason for the enhanced methanogenic capacity of anaerobic granular sludge. Additionally, EPS also improved the abundance of cytochrome c (c-Cyts), accelerating the direct interspecies electron transfer (DIET) between syntrophic bacteria and methanogens, thus enhancing the methane production. Interestingly, the average particle size, volatile suspended solids/total suspended solids (VSS/TSS) and EPS content of anaerobic granular sludge (AnGS) in the EPS reactor were approximately equal to that of the control reactor during the anaerobic digestion, illustrating that EPS could not affect the physicochemical properties of AnGS. Therefore, these results suggested that EPS mainly played a role in the form of conductive materials in the anaerobic digestion process. Compared with conductive materials, EPS as biomass conductive materials was not only environmentally friendly and economical but also no secondary pollution.\n\nImportanceCompared with the reported conductive materials, EPS has the potential of biodegradation, electron transfer and no significant secondary pollution. Besides, there are few studies on the utilization of EPS resources, especially the effect of EPS as an additive on anaerobic digestion performance. To clarify whether EPS as conductive materials or carbon source promotes methanogenesis. Therefore, in this study, we investigated the influence of EPS as an additive on the methanogenic capacity, physical-chemical properties, microbial community structure and metabolic function of anaerobic granular sludge (AnGS), and preliminarily expatiate the influence mechanism of EPS as an additive on methanogenesis. At the meantime, the research is expected to provide new solutions for the improvement of anaerobic digestion performance and disposal of waste mud.

biochemistry↗

bioRxiv2019

Mutant ACTB mRNA 3′UTR Promotes Hepatocellular Carcinoma Development by Regulating miR-1 and miR-29a

In recent years, mounting studies have shown that ACTB is closely related to various tumors. Although ACTB is dysregulated in numerous cancer types, limited data are available on the potential function and mechanism of ACTB in hepatocellular carcinoma (HCC). This study evaluated the expression and biological roles of mutant ACTB mRNA 3'UTR in HCC. Transcriptome sequence and qRT-PCR analysis determined that mutant ACTB mRNA 3'UTR was high expression in HCC tissues. Luciferase reporter assay showed that the ACTB mRNA 3'UTR mutations made it easier to interact with miR-1 and miR-29a. Moreover, mutant ACTB mRNA 3'UTR regulated miR-1 and miR-29a degradation via AGO2. Furthermore, mutant ACTB mRNA 3'UTR promoted hepatocellular carcinoma cells migration and invasion in vitro and in vivo by up-regulating miR-1 target gene MET and miR-29a target gene MCL1. In a word, our study demonstrates that 3'UTR of ACTB plays a key role in the tumor growth of hepatocellular carcinoma (HCC) and highlights the molecular mechanisms of ACTB-involved cancer growth and development.

cancer biology↗

bioRxiv2019

Super-resolution imaging reveals the evolution of higher-order chromatin folding in early carcinogenesis

Aberrant chromatin structure is a hallmark in cancer cells and has long been used for clinical diagnosis of cancer. However, underlying higher-order chromatin folding during malignant transformation remains elusive, due to the lack of molecular scale resolution. Using optimized stochastic optical reconstruction microscopy (STORM) for pathological tissue (PathSTORM), we uncovered a gradual decompaction and fragmented higher-order chromatin folding throughout all stages of carcinogenesis in multiple tumor types, even prior to the tumor formation. Our integrated imaging, genomic, and transcriptomic analyses reveal the functional consequences in enhanced formation of transcription factories, spatial juxtaposition with relaxed nanosized chromatin domains and impaired genomic stability. We also demonstrate the potential of imaging higher-order chromatin decompaction to detect high-risk precursors that cannot be distinguished by conventional pathology. Taken together, our findings reveal the gradual decompaction and fragmentation of higher-order chromatin structure as an enabling characteristic in early carcinogenesis to facilitate malignant transformation, which may improve cancer diagnosis, risk stratification, and prevention.\n\nSIGNIFICANCEGenomic DNA is folded into a higher-order structure that regulates transcription and maintains genomic stability. Although much progress has been made on understanding biochemical characteristics of epigenetic modifications in cancer, the higher-order folding of chromatin structure remains largely unknown. Using optimized super-resolution microscopy, we uncover de-compacted and fragmented chromatin folding in tumor initiation and stepwise progression in multiple tumor types, even prior to the presence of tumor cells. This study underlines the significance of unfolding higher-order chromatin structure as an enabling characteristic to promote tumorigenesis, which may facilitate the development and evaluation of new preventive strategies. The potential of imaging higher-order chromatin folding to improve cancer detection and risk stratification is demonstrated by detecting high-risk precursors that cannot be distinguished by conventional pathology.

cancer biology↗

bioRxiv2019

VGLUT2/ Cdk5/p25 Signaling Pathway Contributed to Inflammatory Pain By CFA

Vesicular glutamate transporter type 2 (VGLUT2) is known to play an important role in mediating the heat hyperalgesia induced by inflammation. However, the underlying mechanism for this activity is poorly understood. Cyclin-dependent kinase 5 (Cdk5), serving as a key regulator in mediating release of glutamate, contributed to the inflammatory heat. It remains unknown whether there is a bridge between Cdk5 and VGLUT2 for mediating inflammatory pain. Therefore, we designed the experiment to determine whether VGLUT2 signaling pathway is involved in Inflammatory pain mediated by Cdk5 and the heat hyperalgesia induced by complete Freunds adjuvant (CFA) can be reversed by roscovitine, a selective inhibitor for Cdk5 through inhibition of VGLUT2 expression. Immunohistochemistry results suggest that when compared with rats in a control group, rats in an experimental group showed significant coexpression of Cdk5/VGLUT2 in small and medium-sized neuronal cells of the dorsal root ganglion (DRG) and spinal cord between days 1 and 3 following subcutaneous injection of CFA. Moreover, our study revealed that the expression of VGLUT2 protein in DRG and spinal cord was remarkably increased between days 1 and 3 following CFA injection. Additionally, p25 but not p35, a activator of Cdk5, protein was significantly increased and reduced by roscovitine. The increased expressions of VGLUT2 protein was significantly reduced by roscovitine as well. Our study showed that VGLUT2/Cdk5 signaling pathway contributed to the inflammatory pain medicated by Cdk5/p25.

neuroscience↗