Seed osmopriming with polyethylene glycol (PEG) enhances seed germination and seedling physiological traits of Coronilla varia L. under water stress.

Water stress can adversely affect seed germination and plant growth. Seed osmopriming is a pre-sowing treatment in which seeds are soaked in osmotic solutions to undergo the first stage of germination prior to radicle protrusion. Seed osmopriming enhances germination performance under stressful environmental conditions, making it an effective method to improve plant resistance and yield. This study analyzed the effect of seed osmopriming with polyethylene glycol (PEG) on seed germination and physiological parameters of Coronilla varia L. Priming treatments using 10% to 30% PEG enhanced germination percentage, germination vigor, germination index, vitality index, and seedling mass and reduced the time to reach 50% germination (T50). The PEG concentration that led to better results was 10%. The content of soluble proteins (SP), proline (Pro), soluble sugars (SS), and malondialdehyde (MDA) in Coronilla varia L. seedlings increased with the severity of water stress. In addition, under water stress, electrolyte leakage rose, and peroxidase (POD) and superoxide dismutase (SOD) activities intensified, while catalase (CAT) activity increased at mild-to-moderate water stress but declined with more severe deficiency. The 10% PEG priming significantly improved germination percentage, germination vigor, germination index, vitality index, and time to 50% germination (T50) under water stress. Across the water stress gradient here tested (8 to 12% PEG), seed priming enhanced SP content, Pro content, and SOD activity in Coronilla varia L. seedlings compared to the unprimed treatments. Under 10% PEG-induced water stress, primed seedlings displayed a significantly lower MDA content and electrolyte leakage than their unprimed counterparts and exhibited significantly higher CAT and POD activities. However, under 12% PEG-induced water stress, differences in electrolyte leakage, CAT activity, and POD activity between primed and unprimed treatments were not significant. These findings suggest that PEG priming enhances the osmotic regulation and antioxidant capacity of Coronilla varia seedlings, facilitating seed germination and seedling growth and alleviating drought stress damage, albeit with reduced efficacy under severe water deficiency.

Magnetic Field Enhanced Oxygen Reduction Reaction via Oxygen Diffusion Speedup.

The mass-transfer of oxygen in liquid phases (including in the bulk electrolyte and near the electrode surface) is a critical step to deliver oxygen to catalyst sites (especially immersed catalyst sites) and use the full capacity of oxygen reduction reaction (ORR). Despite the extensive efforts of optimizing the complex three-phase reaction interfaces to enhance the gaseous oxygen transfer, strong limitations remain due to oxygen's poor solubility and slow diffusion in electrolytes. Herein, a magnetic method for boosting the directional hydrodynamic pumping of oxygen toward immersed catalyst sites is demonstrated which allows the ORR to reach otherwise inaccessible catalytic regions where high currents normally would have depleted oxygen. For Pt foil electrodes without forced oxygen saturation in KOH electrolytes, the mass-transfer-limited current densities can be improved by 60% under an external magnetic field of 435 mT due to the synergistic effect between bulk- and surface-magnetohydrodynamic (MHD) flows induced by Lorentz forces. The residual magnetic fields are further used at the surface of magnetic materials (such as CoPt alloys and Pt/FeCo heterostructures) to enhance the surface-MHD effect, which helps to retain part of the ORR enhancement permanently without applying external magnetic fields.

Gas chromatography-mass spectrometry pilot study to identify volatile organic compound biomarkers of childhood obesity with dyslipidemia in exhaled breath.

Objectives: Childhood obesity affects multiple organs in the body and is associated with both significant morbidity and ultimately premature mortality. Childhood obesity, especially dyslipidemia, can lead to early atherosclerosis and premature cardiovascular disease (CVD) in adulthood. The detection of exhaled volatile organic compounds (VOCs) in the breath offers the opportunity for the discovery of novel disease-specific biomarkers. This study aimed to identify VOCs that correlate with childhood obesity accompanied by dyslipidemia. Methods: A total of 82 overweight or obese children between the ages of 8 and 12 years were recruited from the exercise on obesity adolescents in Peking (EXCITING) study (NCT04984005). The breath VOCs of the participants were measured by gas chromatography-mass spectrometry (GC-MS). The classification was performed using principal component analysis (PCA) of the relative abundance of VOCs. The difference between the obese and overweight groups with or without dyslipidemia was analyzed. Results: Among the 82 children, 25 were overweight, of whom 10 had dyslipidemia. The other 57 children were obese, and 17 of them had dyslipidemia. Obese children with dyslipidemia had higher triglycerides and elevated non-high-density lipoprotein-cholesterol compared to overweight children without dyslipidemia. We confirmed 13 compounds based on database well matches (average score > 80) for mass spectra and refractive index. These 13 VOCs were grouped into three chemical functional groups: saturated hydrocarbons, aromatic hydrocarbons and unsaturated aldehydes. For obese children with dyslipidemia, the PCA scatter plot of the three chemical groups was obviously separated from the other groups. Some of the candidates, including heptadecane, naphthalene, and cis-6-nonnenol, were significantly higher in obese children with dyslipidemia than in overweight groups with or without dyslipidemia. Conclusion: A suite of VOCs from three chemical function groups, saturated hydrocarbons, aromatic hydrocarbons, and unsaturated aldehydes, were separated in the obese children with dyslipidemia. Heptadecane, naphthalene, and cis-6-nonenol were significantly elevated in obese children with dyslipidemia. Our findings underscore the potential value of the candidate VOCs for future risk categorization.

The heterogeneity of oxidized lipids in individual tumor cells reveals NK cell-mediated cytotoxicity by label-free mass cytometry.

NK cell-mediated immunotherapy has received increasing attention in the past decade due to its efficacy and bio-safety. The composition and content of lipids in individual cells are closely related to NK cell-mediated cytotoxicity, especially polyunsaturated fatty acids (PUFA) which are oxidized during NK cell-mediated apoptosis. Here we investigated the changes of lipids in single HepG2 cells by label-free mass cytometry and obtained information on 53 lipids and 13 oxidized lipids after the interaction with NK92 MI cells. We found that the contents of lipids and oxidized lipids of HepG2 cells changed obviously during the NK cell-mediated apoptosis. The HepG2 cells could be classified into two phenotypes after co-culturing with NK92 MI cells based on the ratio of PC(38:6-2OH)/PC(38:6) in individual cells, which may serve as a feature to evaluate NK cell-mediated cytotoxicity. The present work used the lipids and oxidized lipids of individual cells to reveal the heterogeneity in NK cell-mediated apoptosis which would be a powerful method for evaluating the cytotoxicity of NK cells at the single-cell level.

Dynamic metabolic change of cancer cells induced by natural killer cells at the single-cell level studied by label-free mass cytometry.

Natural killer cells (NK cells) are important immune cells which have attracted increasing attention in cancer immunotherapy. Due to the heterogeneity of cells, individual cancer cells show different resistance to NK cytotoxicity, which has been revealed by flow cytometry. Here we used label-free mass cytometry (CyESI-MS) as a new tool to analyze the metabolites in Human Hepatocellular Carcinoma (HepG2) cells at the single-cell level after the interaction with different numbers of NK92 MI cells. A large amount of chemical information from individual HepG2 cells was obtained showing the process of cell apoptosis induced by NK cells. Nineteen metabolites which consecutively change during cell apoptosis were revealed by calculating their average relative intensity. Four metabolic pathways were impacted during cell apoptosis which hit 4 metabolites including glutathione (GSH), creatine, glutamic acid and taurine. We found that the HepG2 cells could be divided into two phenotypes after co-culturing with NK cells according to the bimodal distribution of concentration of these 4 metabolites. The correlation between metabolites and different apoptotic pathways in the early apoptosis cell group was established by the 4 metabolites at the single-cell level. This is a new idea of using single-cell specific metabolites to reveal the metabolic heterogeneity in cell apoptosis which would be a powerful means for evaluating the cytotoxicity of NK cells.

Functional Analysis of Genes GlaDFR1 and GlaDFR2 Encoding Dihydroflavonol 4-Reductase (DFR) in Gentiana lutea L. Var. Aurantiaca (M. Laínz) M. Laínz.

Anthocyanins are important pigments for flower color, determining the ornamental and economic values of horticultural plants. As a key enzyme in the biosynthesis of anthocyanidins, dihydroflavonol 4-reductase (DFR) catalyzes the reduction of dihydroflavonols to generate the precursors for anthocyanidins (i.e., leucoanthocyanidins) and anthocyanins. To investigate the functions of DFRs in plants, we cloned the GlaDFR1 and GlaDFR2 genes from the petals of Gentiana lutea var. aurantiaca and transformed both genes into Nicotiana tabacum by Agrobacterium-mediated leaf disc method. We further investigated the molecular and phenotypic characteristics of T1 generation transgenic tobacco plants selected based on the hygromycin resistance and verified by both PCR and semiquantitative real-time PCR analyses. The phenotypic segregation was observed in the flower color of the transgenic tobacco plants, showing petals darker than those in the wild-type (WT) plants. Results of high-performance liquid chromatography (HPLC) analysis showed that the contents of gentiocyanin derivatives were decreased in the petals of transgenic plants in comparison to those of WT plants. Ours results revealed the molecular functions of GlaDFR1 and GlaDFR2 in the formation of coloration, providing solid theoretical foundation and candidate genes for further genetic improvement in flower color of plants.

Artificial Neural Network Assisted Error Correction for MLC NAND Flash Memory.

The multilevel per cell technology and continued scaling down process technology significantly improves the storage density of NAND flash memory but also brings about a challenge in that data reliability degrades due to the serious noise. To ensure the data reliability, many noise mitigation technologies have been proposed. However, they only mitigate one of the noises of the NAND flash memory channel. In this paper, we consider all the main noises and present a novel neural network-assisted error correction (ANNAEC) scheme to increase the reliability of multi-level cell (MLC) NAND flash memory. To avoid using retention time as an input parameter of the neural network, we propose a relative log-likelihood ratio (LLR) to estimate the actual LLR. Then, we transform the bit detection into a clustering problem and propose to employ a neural network to learn the error characteristics of the NAND flash memory channel. Therefore, the trained neural network has optimized performances of bit error detection. Simulation results show that our proposed scheme can significantly improve the performance of the bit error detection and increase the endurance of NAND flash memory.

Subcellular localization of biomolecules and drug distribution by high-definition ion beam imaging.

Simultaneous visualization of the relationship between multiple biomolecules and their ligands or small molecules at the nanometer scale in cells will enable greater understanding of how biological processes operate. We present here high-definition multiplex ion beam imaging (HD-MIBI), a secondary ion mass spectrometry approach capable of high-parameter imaging in 3D of targeted biological entities and exogenously added structurally-unmodified small molecules. With this technology, the atomic constituents of the biomolecules themselves can be used in our system as the "tag" and we demonstrate measurements down to ~30 nm lateral resolution. We correlated the subcellular localization of the chemotherapy drug cisplatin simultaneously with five subnuclear structures. Cisplatin was preferentially enriched in nuclear speckles and excluded from closed-chromatin regions, indicative of a role for cisplatin in active regions of chromatin. Unexpectedly, cells surviving multi-drug treatment with cisplatin and the BET inhibitor JQ1 demonstrated near total cisplatin exclusion from the nucleus, suggesting that selective subcellular drug relocalization may modulate resistance to this important chemotherapeutic treatment. Multiplexed high-resolution imaging techniques, such as HD-MIBI, will enable studies of biomolecules and drug distributions in biologically relevant subcellular microenvironments by visualizing the processes themselves in concert, rather than inferring mechanism through surrogate analyses.

Nanoscopic subcellular imaging enabled by ion beam tomography.

Multiplexed ion beam imaging (MIBI) has been previously used to profile multiple parameters in two dimensions in single cells within tissue slices. Here, a mathematical and technical framework for three-dimensional (3D) subcellular MIBI is presented. Ion-beam tomography (IBT) compiles ion beam images that are acquired iteratively across successive, multiple scans, and later assembled into a 3D format without loss of depth resolution. Algorithmic deconvolution, tailored for ion beams, is then applied to the transformed ion image series, yielding 4-fold enhanced ion beam data cubes. To further generate 3D sub-ion-beam-width precision visuals, isolated ion molecules are localized in the raw ion beam images, creating an approach coined as SILM, secondary ion beam localization microscopy, providing sub-25 nm accuracy in original ion images. Using deep learning, a parameter-free reconstruction method for ion beam tomograms with high accuracy is developed for low-density targets. In cultured cancer cells and tissues, IBT enables accessible visualization of 3D volumetric distributions of genomic regions, RNA transcripts, and protein factors with 5 nm axial resolution using isotope-enrichments and label-free elemental analyses. Multiparameter imaging of subcellular features at near macromolecular resolution is implemented by the IBT tools as a general biocomputation pipeline for imaging mass spectrometry.

Metal-isotope-tagged monoclonal antibodies for high-dimensional mass cytometry.

Advances in single-cell mass cytometry have increasingly improved highly multidimensional characterization of immune cell heterogeneity. The immunoassay multiplexing capacity relies on monoclonal antibodies labeled with stable heavy-metal isotopes. To date, a variety of rare-earth elements and noble and post-transition metal isotopes have been used in mass cytometry; nevertheless, the methods used for antibody conjugation differ because of the individual metal coordination chemistries and distinct stabilities of various metal cations. Herein, we provide three optimized protocols for conjugating monoclonal IgG antibodies with 48 high-purity heavy-metal isotopes: (i) 38 isotopes of lanthanides, 2 isotopes of indium, and 1 isotope of yttrium; (ii) 6 isotopes of palladium; and (iii) 1 isotope of bismuth. Bifunctional chelating agents containing coordinative ligands of monomeric DOTA (1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid) or polymeric pentetic acid (DTPA) were used to stably sequester isotopic cations in aqueous solutions and were subsequently coupled to IgG antibodies using site-specific biorthogonal reactions. Furthermore, quantification methods based on antibody inherent absorption at 280 nm and on extrinsic absorption at 562 nm after staining with bicinchoninic acid (BCA) are reported to determine metal-isotope-tagged antibodies. In addition, a freeze-drying procedure to prepare palladium isotopic mass tags is described. To demonstrate the utility, experiments using six palladium-tagged CD45 antibodies for barcoding assays of live immune cells in cytometry by time-of-flight (CyTOF) are described. Conjugation of pure isotopes of lanthanides, indium, or yttrium takes ~3.5 h. Conjugation of bismuth takes ~4 h. Preparation of palladium mass tags takes ~8 h. Conjugation of pure isotopes of palladium takes ~2.5 h. Antibody titration takes ~4 h.

Commonly Occurring Cell Subsets in High-Grade Serous Ovarian Tumors Identified by Single-Cell Mass Cytometry.

We have performed an in-depth single-cell phenotypic characterization of high-grade serous ovarian cancer (HGSOC) by multiparametric mass cytometry (CyTOF). Using a CyTOF antibody panel to interrogate features of HGSOC biology, combined with unsupervised computational analysis, we identified noteworthy cell types co-occurring across the tumors. In addition to a dominant cell subset, each tumor harbored rarer cell phenotypes. One such group co-expressed E-cadherin and vimentin (EV), suggesting their potential role in epithelial mesenchymal transition, which was substantiated by pairwise correlation analyses. Furthermore, tumors from patients with poorer outcome had an increased frequency of another rare cell type that co-expressed vimentin, HE4, and cMyc. These poorer-outcome tumors also populated more cell phenotypes, as quantified by Simpson's diversity index. Thus, despite the recognized genomic complexity of the disease, the specific cell phenotypes uncovered here offer a focus for therapeutic intervention and disease monitoring.

Atomic mass tag of bismuth-209 for increasing the immunoassay multiplexing capacity of mass cytometry.

Mass cytometry (or CyTOF) is an atomic mass spectrometry-based single-cell immunoassay technology, which has provided an increasingly systematic and sophisticated view in basic biological and clinical studies. Using elemental reporters composed of stable heavy metal isotopes, more than 50 cellular parameters are measured simultaneously. However, this current multiplexing does not meet the theoretical capability of CyTOF instrumentation with 135 detectable channels, primarily due to the limitation of available chemistries for conjugating elemental mass tags to affinity reagents. To address this issue, we develop herein additional metallic mass tag based on bismuth-209 (209 Bi) for efficient conjugation to monoclonal antibody. This enables the use of an addtional channel m/z = 209 of CyTOF for single-cell immunoassays. Bismuth has nearly the same charge-to-radius ratio as lanthanide elements; thus, bismuth(III) cations (209 Bi3+ ) could coordinate with DTPA chelators in the same geometry of O- and N-donor groups as that of lanthanide. In this report, the coordination chemistry of 209 Bi3+ with DTPA chelators and Maxpar® X8 polymers were investigated in details. Accordingly, the protocols of conjugating antibody with bismuth mass tag were provided. A method based on UV-Vis absorbance at 280 nm of 209 Bi3+ -labeling DTPA complexes was developed to evaluate the stoichiometric ratio of 209 Bi3+ cations to the conjugated antibody. Side-by-side single-cell analysis experiments with bismuth- and lanthanide-tagged antibodies were carried out to compare the analytical sensitivities. The measurement accuracy of bismuth-tagged antibody was validated within in vitro assay using primary human natural killer cells. Furthermore, bismuth-tagged antibodies were successfully employed in cell cycle measurements and high-dimensional phenotyping immunoassays. © 2017 International Society for Advancement of Cytometry.

Low temperature hydrogen plasma assisted chemical vapor generation for Atomic Fluorescence Spectrometry.

Chemical vapor generation techniques have long been considered as important ways of sample introduction for analytical atomic spectrometry. In this paper, a low temperature plasma assisted chemical vapor generation method which avoids the massive use of consumptive chemical agents was proposed by using atmospheric pressure dielectric barrier discharge. The plasma was generated by hydrogen doped argon gas flow through a quartz tube, serving as a dielectric barrier, which had a copper wire inner electrode and a copper foil outer electrode. An alternative high voltage was applied to electrodes to ignite and sustain the plasma. Sample solutions were converted to aerosol by a nebulizer and then mixed with the plasma to generate hydrides. To confirm the utility of this method, four hydride forming elements, As, Te, Sb and Se, were determined by coupling the low temperature plasma assisted chemical vapor generation system with an atomic fluorescence spectrometer. Responses of As, Te, Sb and Se were linear in the range of 0.5-20µg mL(-1). The RSDs of As, Te, Sb and Se in the present method were less than 4.1% and the absolute detection limits for As, Te, Sb and Se were 0.6ng, 1.0ng, 1.4ng and 1.2ng, respectively. Furthermore, four arsenic species were determined after HPLC separation. The method is green and simple compared with hydride generation with tetrahydroborate and the most attractive characteristic is micro-sampling. In principle, the method offers potential advantages of miniaturization, less consumption and ease of automation.

Multiplex DNA assay based on nanoparticle probes by single particle inductively coupled plasma mass spectrometry.

A multiplex DNA assay based on nanoparticle (NP) tags detection utilizing single particle mode inductively coupled plasma mass spectrometry (SP-ICP-MS) as ultrasensitive readout has been demonstrated in the article. Three DNA targets associated with clinical diseases (HIV, HAV, and HBV) down to 1 pM were detected by DNA probes labeled with AuNPs, AgNPs, and PtNPs via DNA sandwich assay. Single nucleotide polymorphisms in genes can also be effectively discriminated. Since our method is unaffected by the sample matrix, it is well-suited for diagnostic applications. Moreover, with the high sensitivity of SP-ICP-MS and the variety of NPs detectable by SP-ICP-MS, high-throughput DNA assay could be achieved without signal amplification or chain reaction amplification.

MicroRNA expression profiles in supraglottic carcinoma.

MicroRNAs (miRNAs) are single-stranded RNA molecules which regulate gene expression at the post-transcriptional level and several miRNAs have been found to be associated with some types of cancer. We sought to identify the expression and involvement of miRNAs in supraglottic carcinoma tissues compared with normal tissues and to determine whether miRNA expression is predictive of disease. Unsupervised clustering shows that miRNA profiles can distinguish tumor from normal tissues. Analysis of miRNA contents in supraglottic carcinoma highlighted nineteen differentially expressed miRNAs, three upregulated miRNAs (miR-21, miR-19a, miR-33a) and two downregulated miRNAs (miR-206, miR-375). The microarray results of supraglottic carcinoma and related computer analysis may be beneficial for further analysis of cancer diagnosis and therapy.

[Responses of tomato leaf photosynthesis to rapid water stress].

By using polyethylene glycol (PEG-6000) solution to regulate the water potential of tomato (Lycopersicon esculentum) rhizosphere to simulate water stress, this paper studied the dynamic changes of net photosynthetic rate, dark respiratory rate and CO2 compensatory concentration of detached tomato leaves in the process of photosynthetic induction. Under 1000 micromol m-2 s-1 of light induction, the time required to reach the maximum net photosynthetic rate of water-stressed tomato leaves was shortened by 1/3, while the stomatal conductance was increased by 1.5 times, as compared to the non-stress control. Also, the light saturation point (LSP) of water-stressed tomato leaves was lowered by 65% to 85%, and the light compensation point (LCP) was increased by 75% to 100%, suggesting that the effective range of light utilized by tomato leaves was reduced. Furthermore, water stress decreased the maximum photosynthetic capacity of tomato leaves by 40%, but increased the dark respiration rate by about 45% . It was suggested that rapid water stress made the stomata of tomato leaves quickly opened, without initial photosynthetic induction stage. In conclusion, water stress could induce the decrease of plant light-energy use efficiency and potential, being the main reason for the decrease of plant productivity, and stomatal regulation could be the main physiological mechanism of tomato plants to adapt to rapid water stress.

Inhibitory action of chamaejasmin A against human HEP-2 epithelial cells: effect on tubulin protein.

In this work, the anticancer activity of chamaejasmin A was studied by evaluating its in vitro cytotoxicity against several cell lines (CAL-27, UMSCC-1, UMSCCG19, HEP-2 and Vero cells) using the 3-(4,5)-dimethylthiazoly1)-3,5-diphenytetrazolium bromide assay. Results indicated chamaejasmin A shows more notable anticancer activity against HEP-2 cells, with IC(50) values of 3.48 µM. Furthermore, western blot analysis showed that chamaejasmin A is able to increase the expression of ß-tubulin (TB), but not α-TB. In vivo, chamaejasmin A intake through gavage resulted in ß-TB depolymerization inhibition in HEP-2 tumors. In silico simulations indicated that chamaejasmin A specifically interacts with the binding site which is located at the top of ß-TB, thanks to the presence of strong hydrophobic effects between the core templates and the hydrophobic surface of the TB protein active site, associated with two strong H-bonds. The binding energy (E (inter)) was calculated to be -129.40 kcal mol(-1). Results above suggest that chamaejasmin A possesses anti-cancer properties relating to ß-TB depolymerization inhibition.