Electrochemiluminescence assay for the impact of Ganoderma lucidum polysaccharides on resisting arsenic-induced apoptosis.

Ganoderma lucidum (G. lucidum) is a traditional edible fungus with strong medicinal value. G. lucidum polysaccharides (GLP) encapsulate many of the key beneficial properties of this species, providing a valuable tool for the treatment of a range of diseases. The present study was developed to explore the protective benefits of GLP treatment in the context of arsenic poisoning. Through microscopy and flow cytometry experiments, NaAsO2 was found to induce rat tracheal epithelial (RTE) cell apoptosis, together with reductions in cell surface epidermal growth factor receptor (EGFR) expression. GLP treatment, however, was able to reduce apoptosis rates and elevate the expression of EGFR relative to NaAsO2-treated cells. GLP extracts (50, 100, 200 mg·mL-1) prepared from four types of G. lucidum were administered to RTE cells damaged with arsenic, revealing limited differences in position resistance among these varieties. This work provides reference for the pharmaceutical and medical research of G. lucidum.

Electrochemiluminescent evaluation of GLUT4 expression in rat adipocytes induced by Ganoderma lucidum polysaccharides.

Glucose transporter 4 (GLUT4) directly facilitates cellular uptake of glucose and plays a crucial role in mammalian adipose tissue glucose metabolism. In this work, we constructed a cytosensor for sensitive electrochemiluminescence (ECL) detection of GLUT4 in rat adipocytes (RA cells). A carbon nanotube sponge (CNTSP) was selected to fabricate a permeable electrode to overcome the steric hindrance of cells on the electrode. The expression of GLUT4 after treatment with Ganoderma lucidum polysaccharide (GLP) was assessed by analyzing the luminescence emitted from cell-surface ECL probes. Our preliminary results suggest that GLP promote the expression of GLUT4, thereby enhancing the uptake of the fluorescent glucose 2-NBDG. Treatment with GLP affected GLUT4 expression in RA cells in a dose-dependent manner. Additionally, the ECL cytosensor contributes to the development of ECL imaging of receptors on the cell surface for clinical drug evaluation.

Vortexing-generated high throughput single-cell droplets for facile analysis of multiplexed microRNA dynamic secretion.

Discriminating secretory phenotypes provides a direct, intact, and dynamic way to evaluate the heterogeneity in cell states and activation, which is significant for dissecting non-genetic heterogeneity for human health studies and disease diagnostics. In particular, secreted microRNAs, soluble signaling molecules released by various cells, are increasingly recognized as a critical mediator for cell-cell communication and the circulating biomarkers for disease diagnosis. However, single-cell analysis of secreted miRNAs is still lacking due to the limited available tools. Herein, we realized three-plexed miRNA secretion analysis over four time points from single cells encapsulated in picoliter droplets with extreme simplicity, coupling vortexing-generated single-cell droplets with multiplexed molecular beacons. Notably, our platform only requires pipetting and vortexing steps to finish the assay setup within 5 min with minimal training, and customized software was developed for automatic data quantification. Applying the platform to human cancer cell lines and primary cells revealed previously undifferentiated heterogeneity and paracrine signaling underlying miRNA secretion. This platform can be used to dissect secretion heterogeneity and cell-cell interactions and has the potential to become a widely used tool in biomedical research.

Acoustic-driven magnetic skyrmion motion.

Magnetic skyrmions have great potential for developing novel spintronic devices. The electrical manipulation of skyrmions has mainly relied on current-induced spin-orbit torques. Recently, it was suggested that the skyrmions could be more efficiently manipulated by surface acoustic waves (SAWs), an elastic wave that can couple with magnetic moment via the magnetoelastic effect. Here, by designing on-chip piezoelectric transducers that produce propagating SAW pulses, we experimentally demonstrate the directional motion of Néel-type skyrmions in Ta/CoFeB/MgO/Ta multilayers. We find that the shear horizontal wave effectively drives the motion of skyrmions, whereas the elastic wave with longitudinal and shear vertical displacements (Rayleigh wave) cannot produce the motion of skyrmions. A longitudinal motion along the SAW propagation direction and a transverse motion due to topological charge are simultaneously observed and further confirmed by our micromagnetic simulations. This work demonstrates that acoustic waves could be another promising approach for manipulating skyrmions, which could offer new opportunities for ultra-low power skyrmionics.

On-capillary alkylation micro-reactor: a facile strategy for proteo-metabolome profiling in the same single cells.

Single-cell multi-omics analysis can provide comprehensive insights to study cell-to-cell heterogeneity in normal and disease physiology. However, due to the lack of amplification technique, the measurement of proteome and metabolome in the same cell is challenging. Herein, a novel on-capillary alkylation micro-reactor (OCAM) was developed to achieve proteo-metabolome profiling in the same single cells, by which proteins were first covalently bound to an iodoacetic acid functionalized open-tubular capillary micro-reactor via sulfhydryl alkylation reaction, and metabolites were rapidly eluted, followed by on-column digestion of captured proteins. Compared with existing methods for low-input proteome sample preparation, OCAM exhibited improved efficiency, anti-interference ability and recovery, enabling the identification of an average of 1509 protein groups in single HeLa cells. This strategy was applied to single-cell proteo-metabolome analysis of mouse oocytes at different stages, 3457 protein groups and 171 metabolites were identified in single oocytes, which is the deepest coverage of proteome and metabolome from single mouse oocytes to date, achieving complementary characterization of metabolic patterns during oocyte maturation.

Effects of adding lignocellulose-degrading microbial agents and biochar on nitrogen metabolism and microbial community succession during pig manure composting.

This study assessed the influence of the additions of lignocellulose-degrading microbial agents and biochar on nitrogen (N) metabolism and microbial community succession during pig manure composting. Four treatments were established: CK (without additives), M (lignocellulose-degrading microbial agents), BC (biochar), and MBC (lignocellulose-degrading microbial agents and biochar). The results revealed that all treatments with additives decreased N loss compared with CK. In particular, the concentrations of total N and NO3--N were the highest in M, which were 21.87% and 188.67% higher than CK, respectively. Meanwhile, the abundance of denitrifying bacteria Flavobacterium, Enterobacter, and Devosia reduced with additives. The roles of Anseongella (nitrifying bacterium) and Nitrosomonas (ammonia-oxidizing bacterium) in NO3--N transformation were enhanced in M and BC, respectively. N metabolism pathway prediction indicated that lignocellulose-degrading microbial agents addition could enhance N retention effectively mainly by inhibiting denitrification. The addition of biochar enhanced oxidation of NH4+-N to NO2--N and N fixation, as well as inhibited denitrification. These results revealed that the addition of lignocellulose-degrading microbial agents individually was more conducive to improve N retention in pig manure compost.

Deciphering Cell-Cell Interactions with Integrative Single-Cell Secretion Profiling.

Cell-cell interactions are the fundamental behaviors to regulate cellular activities. A comprehensive evaluation of intercellular interactions requires direct profiling of various signaling behaviors simultaneously at the single-cell level, which remains lacking. Herein, an integrative single-cell secretion analysis platform is presented to profile different secreted factors (four proteins, three extracellular vesicles (EV) phenotypes), spatial distances, and migration information (distances and direction) simultaneously from high-throughput paired single cells using an antibody-barcode microchip. Applying the platform to analyze the tumor-stromal and tumor-immune interactions with the human oral squamous cell carcinoma (OSCC) cell lines and primary OSCC cells reveals that the initial distances between cells would determine their migratory distances and direction to approach stable organization. The cell-cell in close proximity enhances protein secretions while attenuating EV secretions. Migration has a more profound correlation with protein secretions than EV secretions, in which absolute migration distance affects protein secretions significantly but not the direction. These findings highlight the significance of spatial organization in regulating cell signaling behaviors and demonstrate that the integrative single-cell secretion profiling platform is well-suited for a comprehensive dissection of intercellular communication and interactions, providing new avenues for understanding cell-cell interaction biology and how different signaling behaviors coordinate within the tumor microenvironment.

Carbon dots promoted soybean photosynthesis and amino acid biosynthesis under drought stress: Reactive oxygen species scavenging and nitrogen metabolism.

With global warming and water scarcity, improving the drought tolerance and quality of crops is critical for food security and human health. Here, foliar application of carbon dots (CDs, 5 mg·L-1) could scavenge reactive oxygen species accumulation in soybean leaves under drought stress, thereby enhancing photosynthesis and carbohydrate transport. Moreover, CDs stimulated root secretion (e.g., amino acids, organic acids, and auxins) and recruited beneficial microorganisms (e.g., Actinobacteria, Ascomycota, Acidobacteria and Glomeromycota), which facilitate nitrogen (N) activation in the soil. Meanwhile, the expression of GmNRT, GmAMT, and GmAQP genes were up-regulated, indicating enhanced N and water uptake. The results demonstrated that CDs could promote nitrogen metabolism and enhance amino acid biosynthesis. Particularly, the N content in soybean shoots and roots increased significantly by 13.2 % and 30.5 %, respectively. The amino acids content in soybean shoots and roots increased by 257.5 % and 57.5 %, respectively. Consequently, soybean yields increased significantly by 21.5 %, and the protein content in soybean kernels improved by 3.7 %. Therefore, foliar application of CDs can support sustainable nano-enabled agriculture to combat climate change.

Mapping secretome-mediated interaction between paired neuron-macrophage single cells.

Neuron-immune interaction through secreted factors contributes significantly to the complex microenvironment in the central nervous system that could alter cell functionalities and fates in both physiological and pathological conditions, which remains poorly characterized at the single-cell level. Herein, using a spatially patterned antibody barcode microchip, we realized the mapping of 12 different secretomes, covering cytokines, neurotrophic factors (NFs), and neuron-derived exosomes (NDEs) from high-throughput, paired single cells (≥ 600) simultaneously under normal conditions and an Alzheimer's disease (AD) model induced with amyloid beta protein 1-42 (Aß1-42). We applied the platform to analyze the secretion profiles from paired neuron-macrophage and neuron-microglia single cells with human cell lines. We found that pairwise neuron-macrophage interaction would trigger immune responses and attenuate neuron cells' secretion, while neuron-microglia interaction generally results in opposite outcomes in secretion. When neuron cells are induced with Aß1-42 protein into the AD model, both neuron-macrophage and neuron-microglia interactions lead to increased cytokines and NDEs and decreased NFs. Further analysis of AD patients' serum showed that NDEs were significantly higher in patients' samples than in the control group, validating our observation from the interaction assay. Furthermore, we resolved previously undifferentiated heterogeneity underlying the secretions from single-neuron cells. We found that the NDE and NF secretion was less dependent on the paracrine signaling between one another and that secretions from neuron cells would attenuate after differentiation with Aß1-42. This study demonstrates the mapping of the different secretomes from paired neuron-immune single cells, providing avenues for understanding how neurons and immune cells interact through the complex secretome network.

Carbon Dots Improve Nitrogen Bioavailability to Promote the Growth and Nutritional Quality of Soybeans under Drought Stress.

The inefficient utilization of nitrogen (N) in soil and drought stress seriously threatens agricultural and food production. Herein, soil application of carbon dots (CDs, 5 mg kg-1) promoted the growth and nutritional quality of soybeans by improving N bioavailability, which was beneficial to alleviate the economic losses caused by drought stress. Soil application of CDs enhanced the N-fixing ability of nodules, regulated rhizosphere processes, and ultimately enhanced N and water uptake in soybeans under drought stress. Compared to control (drought stress), the application of CDs under drought stress enhanced soybean nitrogenase activity by 8.6% and increased N content in soybean shoots and roots by 18.5% and 14.8%, respectively. CDs in soil promoted the secretion of root exudates (e.g., organic acids, fatty acids, and polyketides) and regulated beneficial microbial communities (e.g., Proteobacteria, Acidobacteria, Gemmatimonadetes, and Actinobacteria), thus enhancing the N release from soil. Besides, compared to control, the expression of GmNRT, GmAMT, GmLB, and GmAQP genes in roots were upregulated by 1.2-, 1.8-, 2.7-, and 2.3-fold respectively, implying enhanced N transport and water uptake. Furthermore, the proteins, fatty acids, and amino acids in soybean grains were improved by 3.4%, 6.9%, and 17.3%, respectively, as a result of improved N bioavailability. Therefore, CD-enabled agriculture is promising for improving the drought tolerance and quality of soybeans, which is of significance for food security in facing the crisis of global climate change.

High-Throughput Single-Cell Extracellular Vesicle Secretion Analysis on a Desktop Scanner without Cell Counting.

Single-cell EV (extracellular vesicle) secretion analysis is emerging for a better understanding of non-genetic cellular heterogeneity regulating human health and diseases through intercellular mediators. However, the requirements of expensive and bulky instrumentations hinder its widespread use. Herein, by combining gold nanoparticle-enhanced silver staining and the Poisson distribution, we reported the use of a home-use scanner to realize high-throughput single-cell EV secretion analysis without cell counting. We applied the platform to analyze the secretions of different EV phenotypes with the human oral squamous cell carcinoma cell line and primary cells from patients, which generated single-cell results comparable with those of the immunofluorescence approach. Notably, we also realized the quantification of the number of EVs secreted from every single cell using their respective titration curves obtained from population samples, making it possible to directly compare different EV phonotypes in regard to their secretion number, secretion rate, and so forth. The technology introduced here is simple, easy to operate, and of low cost, which make it a potential, easily accessible, and affordable tool for widespread use in both basic and clinical research.

Single-Cell Secretion Analysis in the Engineered Tumor Microenvironment Reveals Differential Modulation of Macrophage Immune Responses.

It is increasingly recognized that the cellular microenvironment plays critical roles in regulating the fate and physiology of cells. Despite recent advancements in single-cell analysis technologies, engineering and integration of the microenvironment for single-cell analysis platforms remain limited. Here, we report a single-cell cytokine secretion analysis platform that integrated both the three-dimensional cell culture and the primary oral squamous cell carcinoma tumor cell co-culture to provide both physical and physiological cues for single cells to be analyzed. We apply the platform to investigate the immune responses of human macrophages stimulated with the ligand of toll-like receptor 4 lipopolysaccharide. Notably, we observe the differential modulation effect in cytokine secretions by the tumor microenvironment, in which antitumor cytokine TNF-a secretion was attenuated, and protumor cytokine IL-6 would increase. The differential modulation effect is conserved from cell line-derived macrophages to primary macrophages derived from healthy donors. Immunofluorescence staining further reveals that ∼50% of macrophage cells could be polarized from M1 to the M2 phenotype within 12 h in the engineered tumor microenvironment. This work demonstrates the significance of the cell microenvironment toward single-cell analysis, which could help to evaluate how immune cells will respond in the complex microenvironment more accurately.

Surface Roughness: A Crucial Factor To Robust Electric Double Layer Capacitors.

Electric double layer capacitors (EDLCs) usually show high rate performance and long cycling spans but inferior specific capacitance, which are mainly created by restriction of the charge storage mechanism. To improve the capacitive performance, traditional methods include enlarging surface area, optimizing porous structures, and readjusting functional groups through heteroatom doping to electrode materials. Besides that, another promising approach is suggested, which is to enhance surface roughness of the electrode materials for ion storage and transport. To prove this view, two porous carbon materials were fabricated by activation-calcination methods, which allowed the materials to have identical surface area, porous structures, and surface composition but the surface roughness. Further electrochemical measurements exhibited that the optimal sample with higher roughness has remarkable specific capacitance (up to 562 F g-1), and the increment rate is more than 50% when compared with contrast sample (367 F g-1). Therefore, optimization of the surface roughness of electrode materials is another efficient route to make robust EDLCs.

PDMS Microwell Stencil Based Multiplexed Single-Cell Secretion Analysis.

Multiplexed single-cell protein secretion analysis provides an in-depth understanding of cellular heterogeneity in intercellular communications mediated by secreted proteins in both fundamental and clinical research. However, it has been challenging to increase the proteomic parameters co-profiled from every single cell in a facile way. Herein, a simple method to improve the multiplexed proteomic parameters of PDMS microwell based single-cell secretion analysis platform by sandwiching PDMS stencil in between two antibody-coated glass slides is introduced. Two different antibody panels can be immobilized easily by static coating, without using sophisticated fluid handling or bulky equipment. 5-plexed, 3-fluorescence color single-cell secretion assay is demonstrated with this platform to investigate human monocytic U937 cells in response to lipopolysaccharide and phorbol myristate acetate stimulation, which identified the existence of functional subsets dictated by different cytokine profiles. The technology introduced here is simple, easy to operate, which holds great potential to become a powerful tool for profiling multiplexed single-cell cytokine secretion at high throughput to dissect cellular heterogeneity in secretome signatures.

Corrigendum to "Modeling Inhibitory Effect on the Growth of Uninfected T Cells Caused by Infected T Cells: Stability and Hopf Bifurcation".

[This corrects the article DOI: 10.1155/2018/3176893.].

Multiplexed profiling of single-cell extracellular vesicles secretion.

Extracellular vesicles (EVs) are important intercellular mediators regulating health and diseases. Conventional methods for EV surface marker profiling, which was based on population measurements, masked the cell-to-cell heterogeneity in the quantity and phenotypes of EV secretion. Herein, by using spatially patterned antibody barcodes, we realized multiplexed profiling of single-cell EV secretion from more than 1,000 single cells simultaneously. Applying this platform to profile human oral squamous cell carcinoma (OSCC) cell lines led to a deep understanding of previously undifferentiated single-cell heterogeneity underlying EV secretion. Notably, we observed that the decrement of certain EV phenotypes (e.g., CD63+EV) was associated with the invasive feature of both OSCC cell lines and primary OSCC cells. We also realized multiplexed detection of EV secretion and cytokines secretion simultaneously from the same single cells to investigate the multidimensional spectrum of cellular communications, from which we resolved tiered functional subgroups with distinct secretion profiles by visualized clustering and principal component analysis. In particular, we found that different cell subgroups dominated EV secretion and cytokine secretion. The technology introduced here enables a comprehensive evaluation of EV secretion heterogeneity at single-cell level, which may become an indispensable tool to complement current single-cell analysis and EV research.

Modeling Inhibitory Effect on the Growth of Uninfected T Cells Caused by Infected T Cells: Stability and Hopf Bifurcation.

We consider a class of viral infection dynamic models with inhibitory effect on the growth of uninfected T cells caused by infected T cells and logistic target cell growth. The basic reproduction number R 0 is derived. It is shown that the uninfected equilibrium is globally asymptotically stable if R 0 < 1. Sufficient conditions for the existence of Hopf bifurcation at the infected equilibrium are investigated by analyzing the distribution of eigenvalues. Furthermore, the properties of Hopf bifurcation are determined by the normal form theory and the center manifold. Numerical simulations are carried out to support the theoretical analysis.

Aberrant DNA methylation in radon and/or cigarette smoke-induced malignant transformation in BEAS-2B human lung cell line.

It is well known that cigarette smoking (CS) and/or radon (Rn) induce malignant transformation in lung cells. To investigate the mechanisms underlying lung carcinogenesis induced by CS, Rn; or Rn followed by CS using BEAS-2B cell line derived from human bronchial epithelial cells. BEAS-2B cells were exposed to either Rn (20,000 Bq/m3) for 30 min or CS (20%) for 10 min or Rn followed by CS for 40 min. Global and gene-specific DNA methylation modifications were measured by microarray and methylation-specific polymerase chain reaction. Cell cycle and apoptosis were determined by flow cytometry, while soft agar colony formation was conducted to assess the characteristics of malignant transformation. Data demonstrated global hypomethylation as well as gene-specific DNA methylation alterations in all treatment groups compared to unexposed control cells. In addition, Rn and CS produced DNA hypermethylation of protein tyrosine phosphatase receptor type M and ectodysplasin A2 receptor, two genes related to malignant transformation. In all treatment conditions, cell proliferation and survival of malignant cells was increased, while apoptosis was initially first passage elevated but decreased at passages 5-15. Our results indicate that aberrant DNA methylation plays an important role in Rn- and/or CS-induced malignant transformation. In addition, BEAS-2B cell line may be used as an in vitro model to investigate mechanisms underlying malignant transformation induced by ambient environmental contaminants.