Mass Spectrometric Analysis Research Articles

FT-ICR-MS reveals the molecular imprints of the brewing process.

The study of fermentation and brewing has a long history of pioneering discoveries that continue to influence modern industrial food production. Since then, numerous research endeavors have yielded conventional criteria that guide contemporary brewing practices. However, the intricate open challenges faced today necessitate a more exhaustive understanding of the process at the molecular scale. We have developed an ultra-high-resolution mass spectrometric analysis (FT-ICR-MS) of the brewing process that can rapidly and comprehensively resolve thousands of molecules. This approach allows us to track molecular fluctuation during brewing at the level of chemical compositions. Employing biological triplicates, our investigation of two brewing lines that are otherwise identical except for the malt used revealed over 8,000 molecular descriptors of the brewing process. Metabolite imprints of both the similarities and differences arising from deviating malting temperatures were visualized. Additionally, we translated traditional brewing attributes such as the EBC-value, free amino nitrogen, pH-value, and concentration curves of specific molecules, into highly correlative molecular patterns consisting of hundreds of metabolites. These in-depth molecular imprints provide a better understanding of the molecular circumstances leading to various changes throughout the brewing process. Such chemical maps go beyond the observation of traditional brewing attributes and are of great significance in the investigation strategies of current open challenges in brewing research. The molecular base of knowledge, along with advancements in technological and data integration schemes, can facilitate the efficient monitoring of brewing and other productions processes.

Secondary metabolites from Pseudallescheria boydii and their anti-melanoma activity

A new asperitaconic acid (1) and an undescribe cyclotetrapeptide (2), together with nine known compounds (3–11), were isolated from the endophytic fungus Pseudallescheria boydii XL-0256 obtained from the leaves of Dimocarpus longan Lour. The structures of the new compounds have been successfully assigned based on the NMR spectroscopic data and the mass spectrometric analysis. Compound 3 was confirmed to significantly reduce B16–F10 cell viability in a dose- and time-dependent manner, as well as promoting cell apoptosis and inhibiting cell migration in B16–F10 cell. In addition, compound 3 was hypothesized to have an anti-melanoma effect through target proteins (ADAM17 and ADAMTS5) based on network pharmacological target prediction and molecular docking studies.

Heterogeneity of otolith chemical composition from two-dimensional mapping: Relationship with biomineralization mechanisms and implications for microchemistry analyses.

Although otoliths are widely used as archives to infer life-history traits and habitat use in fishes, their biomineralization process remains poorly understood. This lack of knowledge is problematic as it can lead to misinterpretation of the different types of signals (e.g., optical or chemical) that provide basic data for research in fish ecology, fisheries management, and species conservation. Otolith calcification relies on a complex system involving a pericrystalline fluid, the endolymph, whose organic and inorganic compositions are spatially heterogeneous for some constituents. This property stems from the particular structure of the calcifying saccular epithelium. In this study, we explored the spatial heterogeneity of elemental incorporation in otoliths for two species of high economic interest, European hake Merluccius merluccius (L. 1758) and European sea bass Dicentrarchus labrax (L. 1758). Two-dimensional mappings of chemical elements were obtained using UV high-repetition-rate femtosecond laser ablation (fs-LA) system coupled to a high-resolution inductively coupled plasma sector field mass spectrometer analyses on transverse sections of sagittae. Results highlighted a clear asymmetry between proximal (sulcus) and distal (antisulcus) concentrations for elements such as magnesium (Mg), phosphorus (P), manganese (Mn), and potassium (K) with concentration gradient directions that varied depending on the element. Strontium (Sr) and barium (Ba) did not show a proximo-distal gradient. These results are discussed in light of current knowledge on the endolymph composition and the mechanisms that lead to its compartmentalization, highlighting the need for further research on otolith biomineralization. Operational implications for studies based on otolith chemical composition are also discussed with emphasis on advice for sampling strategies to avoid analytical biases and the need for in-depth analyses of analytical settings before comparing otolith signatures between species or geographical areas.

Heat-shock protein 90α protects NME1 against degradation and suppresses metastasis of breast cancer.

NME1 has been exploited as a potential translational target for decades. Substantial efforts have been made to upregulate the expression of NME1 and restore its anti-metastasis function in metastatic cancer. Cycloheximide (CHX) chase assay was used to measure the steady-state protein stability of NME1 and HSP90α. The NME1-associating proteins were identified by immunoprecipitation combined with mass spectrometric analysis. Gene knockdown and overexpression were employed to examine the impact of HSP90AA1 on intracellular NME1 degradation. The motility and invasiveness of breast cancer cells were examined in vitro using wound healing and transwell invasion assays. The orthotopic spontaneous metastasis and intra-venous experimental metastasis assays were used to test the formation of metastasis in vivo, respectively. HSP90α interacts with NME1 and increases NME1 lifetime by impeding its ubiquitin-proteasome-mediated degradation. HSP90α overexpression significantly inhibits the metastatic potential of breast cancer cells in vitro and in vivo. A novel cell-permeable peptide, OPT22 successfully mimics the HSP90α function and prolongs the life span of endogenous NME1, resulting in reduced metastasis of breast cancer. These results not only reveal a new mechanism of NME1 degradation but also pave the way for the development of new and effective approaches to metastatic cancer therapy.

Hybrid insulin peptide isomers spontaneously form in pancreatic beta-cells from an aspartic anhydride intermediate

Hybrid insulin peptides (HIPs) form in beta-cells when insulin fragments link to other peptides through a peptide bond. HIPs contain nongenomic amino acid sequences and have been identified as targets for autoreactive T cells in type 1 diabetes. A subgroup of HIPs, in which N-terminal amine groups of various peptides are linked to aspartic acid residues of insulin C-peptide, was detected through mass spectrometry in pancreatic islets. Here, we investigate a novel mechanism that leads to the formation of these HIPs in human and murine islets. Our research herein shows that these HIPs form spontaneously in beta-cells through a mechanism involving an aspartic anhydride intermediate. This mechanism leads to the formation of a regular HIP containing a standard peptide bond as well as a HIP-isomer containing an isopeptide bond by linkage to the carboxylic acid side chain of the aspartic acid residue. We used mass spectrometric analyses to confirm the presence of both HIP isomers in islets, thereby validating the occurrence of this novel reaction mechanism in beta-cells. The spontaneous formation of new peptide bonds within cells may lead to the development of neoepitopes that contribute to the pathogenesis of type 1 diabetes as well as other autoimmune diseases.

Elucidating sulfate radical-induced oxidizing mechanisms of solid-phase pharmaceuticals: Comparison with liquid-phase reactions

As a class of organic micropollutants of global concern, pharmaceuticals have prevalent distributions in the aqueous environment (e.g., groundwater and surface water) and solid matrices (e.g., soil, sediments, and dried sludge). Their contamination levels have been further aggravated by the annually increased production of expired drugs as emerging harmful wastes worldwide. Sulfate radicals (SO4•−)-based oxidation has attracted increasing attention for abating pharmaceuticals in the environment, whereas the transformation mechanisms of solid-phase pharmaceuticals remain unknown thus far. This investigation presented for the first time that SO4•−, individually produced by mechanical force-activated and heat-activated persulfate treatments, could effectively oxidize three model pharmaceuticals (i.e., methotrexate, sitagliptin, and salbutamol) in both solid and liquid phases. The high-resolution mass spectrometric analysis suggested their distinct transformation products formed by different phases of SO4•− oxidation. Accordingly, the SO4•−-mediated mechanistic differences between the solid-phase and liquid-phase pharmaceuticals were proposed. It is noteworthy that the products from both systems were predicted with the remaining persistence, bioaccumulation, and multi-endpoint toxicity. Therefore, some post-treatment strategies need to be considered during practical applications of SO4•−-based technologies in remediating different phases of micropollutants. This work has environmental implications for understanding the comparative transformation mechanisms of pharmaceuticals by SO4•− oxidation in remediating the contaminated solid and aqueous matrices.

DIFFERENTIAL CELL SURFACE PROTEOMICS OF FLOTILLIN-1, HMGB1 AND AMINOPEPTIDASE IN H3 MUTATED PAEDIATRIC HIGH-GRADE GLIOMA

Abstract AIMS High-grade gliomas (HGG) are the deadliest central nervous system tumour in children. Despite this, treatment options remain elusive, leaving children with a dismal prognosis. Histone H3.3 mutations, K27M and G34V/R have been identified as the two most common recurrent somatic mutations in paediatric HGG (pHGG). These mutations are present in 50% of pHGG compared to only 1% in adults. Our previous research on cell membrane protein analyses have identified different signature of unique and common cell membrane proteins in H3-G34R/V mutated pHGG, in comparison to H3 wild type tumour. Among these we have identified flotillin-1, HMGB1 and aminopeptidase N. Flotillin-1 promotes numerous malignancies, High mobility group box 1(HMGB1) functions as an oncogene and aminopeptidase N is a cell surface metallo- protease involved in tumour progression. We have now verified the expression and localization of these cell surface proteins in H3 mutated and wildtype tumour, in order to target them for developing novel therapeutics. METHOD Cell membrane proteomics were further verified though mass-spectrometric data analyses. Expression and localization of flotillin-1, HMGB1 and Aminopeptidase in H3.3 mutated pHGG cell lines in comparison to H3 wildtype glioma were analysed, using different protein expression assays including immunofluorescence microscopy and western blot assays. RESULTS Experimental analyses showed differential protein expression and localization of Flotillin-1, HMGB1 and Aminopeptidase and verified the mass-spectrometric proteomic data in H3-G34V pHGG, in comparison to wild- type pHGG. CONCLUSIONS This new knowledge will help us utilise Flotillin-1, HMGB1 and Aminopeptidase N, as potential therapeutic targets for these unique H3 mutated children’s brain tumours.

DETERMINATION OF THE CHEMICAL COMPOSITION OF COUSINIA MINDSCHELKENSIS B. FEDTSCH. BY GAS CHROMATOGRAPHY-MASS SPECTROMETRY METHOD

Vegetation is important component and factor of the redistribution of chemical elements in the landscape. One or another type of vegetation determines the amount and nature of soil organic matter, often affects the degree of development of browning processes, affects the chemical composition of the air, is an environmental indicator. The study and isolation of physiologically active compounds from plant raw materials and the determination of the place of their demand is the most important urgent task. Cousinia mindschelkensis B. Fedtsch. It is a rare species from the Asteraceae. Methods. In the Research Center of Medicinal Plants of the Al-Farabi KazNU, analyses were carried out for the determination of organic compounds by gas chromatography with mass spectrometric detection and analyses of 70% of alcohol extracts of the aboveground and underground parts species by the method of high-performance liquid chromatography on a liquid chromatograph. Results. Some of the components identified by GC-MS are biologically active compounds. The GC-MS method revealed the presence of 55 phytochemical compounds in the aboveground and 38 in the underground part of plants that can contribute to the medicinal properties of this species. Conclusion. Compounds have some important biological potential for future drug development. Of the aboveground part of the plant includes bicyclo[2.2.1]heptan-2-one, 1.7.7-trimethyl-, (1S)- (9.50%), hexadecanoic acid (8.05%), cis-Vaccenic acid (7.66%), 9.12-Octadecadienoic acid (Z,Z)- (7.40%), hanphyllin (7.01%), 5.8-Dihydroxy-4a-methyl-4,4a,4b,5,6,7,8,8 a,9,10-decahydro-2(3H)-phenanthrenone (6.51%), dibutyl phthalate (5.64%), while the underground part of the plant has a spike of components such as octacosane (26.84%), β-amyrin (22.35%), β-sitosterol (12.63%), 9.12-Octadecadienoic acid (Z,Z)- (8.43%).9,12-Octadecadienoic acid (Z,Z)- it is present both in the aboveground (7.40%) and in the underground part of the plant (8.43%).

Exploiting Chemical Protein Synthesis to Study the Role of Tyrosine Sulfation on Anticoagulants from Hematophagous Organisms.

ConspectusTyrosine sulfation is a post-translational modification (PTM) that modulates function by mediating key protein-protein interactions. One of the early proteins shown to possess this PTM was hirudin, produced in the salivary glands of the medicinal leech Hirudo medicinalis, whereby tyrosine sulfation led to a ∼10-fold improvement in α-thrombin inhibitory activity. Outside of this pioneering discovery, the involvement of tyrosine sulfation in modulating the activity of salivary proteins from other hematophagous organisms was unknown. We hypothesized that the intrinsic instability of the tyrosine sulfate functionality, particularly under the acidic conditions used to isolate and analyze peptides and proteins, has led to poor detection during the isolation and/or expression of these molecules.Herein, we summarize our efforts to interrogate the functional role of tyrosine sulfation in the thrombin inhibitory and anticoagulant activity of salivary peptides and proteins from a range of different blood feeding organisms, including leeches, ticks, mosquitoes, and flies. Specifically, we have harnessed synthetic chemistry to efficiently generate homogeneously sulfated peptides and proteins for detailed structure-function studies both in vitro and in vivo.Our studies began with the leech protein hirudin P6 (from Hirudinaria manillensis), which is both sulfated on tyrosine and O-glycosylated at a nearby threonine residue. Synthetically, this was achieved through solid-phase peptide synthesis (SPPS) with a late-stage on-resin sulfation, followed by native chemical ligation and a folding step to generate six differentially modified variants of hirudin P6 to assess the functional interplay between O-glycosylation and tyrosine sulfation. A one-pot, kinetically controlled ligation of three peptide fragments was used to assemble homogeneously sulfoforms of madanin-1 and chimadanin from the tick Haemaphysalis longicornis. Dual tyrosine sulfation at two distinct sites was shown to increase the thrombin inhibitory activity by up to 3 orders of magnitude through a novel interaction with exosite II of thrombin. The diselenide-selenoester ligation developed by our lab provided us with a means to rapidly assemble a library of different sulfated tick anticoagulant proteins: the andersonins, hyalomins, madanin-like proteins, and hemeathrins, thus enabling the generation of key structure-activity data on this family of proteins. We have also confirmed the presence of tyrosine sulfation in the anticoagulant proteins of Anopheles mosquitoes (anophelins) and the Tsetse fly (TTI) via insect expression and mass spectrometric analysis. These molecules were subsequently synthesized and assessed for thrombin inhibitory and anticoagulant activity. Activity was significantly improved by the addition of tyrosine sulfate modifications and led to molecules with potent antithrombotic activity in an in vivo murine thrombosis model.The Account concludes with our most recent work on the design of trivalent hybrids that tandemly occupy the active site and both exosites (I and II) of α-thrombin, with a TTI-anophelin hybrid (Ki = 20 fM against α-thrombin) being one of the most potent protease inhibitors and anticoagulants ever generated. Taken together, this Account highlights the importance of the tyrosine sulfate post-translational modification within salivary proteins from blood feeding organisms for enhancing anticoagulant activity. This work lays the foundation for exploiting native or engineered variants as therapeutic leads for thrombotic disorders in the future.

Nonequilibrium Behavior in Isoprene Secondary Organic Aerosol.

Recent studies have shown that instantaneous gas-particle equilibrium partitioning assumptions fail to predict SOA formation, even at high relative humidity (∼85%), and photochemical aging seems to be one driving factor. In this study, we probe the minimum aging time scale required to observe nonequilibrium partitioning of semivolatile organic compounds (SVOCs) between the gas and aerosol phase at ∼50% RH. Seed isoprene SOA is generated by photo-oxidation in the presence of effloresced ammonium sulfate seeds at <1 ppbv NOx, aged photochemically or in the dark for 0.3-6 h, and subsequently exposed to fresh isoprene SVOCs. Our results show that the equilibrium partitioning assumption is accurate for fresh isoprene SOA but breaks down after isoprene SOA has been aged for as short as 20 min even in the dark. Modeling results show that a semisolid SOA phase state is necessary to reproduce the observed particle size distribution evolution. The observed nonequilibrium partitioning behavior and inferred semisolid phase state are corroborated by offline mass spectrometric analysis on the bulk aerosol particles showing the formation of organosulfates and oligomers. The unexpected short time scale for the phase transition within isoprene SOA has important implications for the growth of atmospheric ultrafine particles to climate-relevant sizes.

Bisphenol-A and phthalate metabolism in children with neurodevelopmental disorders.

The etiology of autism spectrum (ASD) and Attention Deficit/Hyperactivity (ADHD) disorders are multifactorial. Epidemiological studies have shown associations with environmental pollutants, such as plasticizers. This study focused on two of these compounds, the Bisphenol-A (BPA) and Diethylhexyl Phthalate (DEHP). The major pathway for BPA and DEHP excretion is via glucuronidation. Glucuronidation makes insoluble substances more water-soluble allowing for their subsequent elimination in urine. Detoxification of these two plasticizers is compromised in children with ASD and ADHD. Consequently, their tissues are more exposed to these two plasticizers. We measured the efficiency of glucuronidation in three groups of children, ASD (n = 66), ADHD (n = 46) and healthy controls (CTR, n = 37). The children were recruited from the clinics of Rutgers-NJ Medical School. A urine specimen was collected from each child. Multiple mass spectrometric analyses including the complete metabolome were determined and used to derive values for the efficiency of glucuronidation for 12 varied glucuronidation pathways including those for BPA and MEHP. (1) Both fold differences and metabolome analyses showed that the three groups of children were metabolically different from each other. (2) Of the 12 pathways examined, only the BPA and DEHP pathways discriminated between the three groups. (3) Glucuronidation efficiencies for BPA were reduced by 11% for ASD (p = 0.020) and 17% for ADHD (p<0.001) compared to controls. DEHP showed similar, but not significant trends. ASD and ADHD are clinically and metabolically different but share a reduction in the efficiency of detoxification for both BPA and DEHP with the reductions for BPA being statistically significant.

Extract yields of two commercially valuable wood species in Central Africa: Bilinga and Tali, their nature and role in water sorption activities

ABSTRACT This paper aimed at determining the quantity, nature and role of extracts during the sorption behavior of two Central African hardwood species: Nauclea diderrichii (Bilinga) and Erythrophleum suaveolens (Tali) at 30, 40, 50, 60 and 70°C using relative humidities (RH) between 5.3% and 97%. For this purpose, extract contents and their nature were determined by solvent extraction and mass spectrometric analysis (MALDI-TOF/MS) respectively. The ABC model whose parameters were used to determine the monolayer moisture capacity (EMCm) also described type II sigmoidal sorption isotherms. The results showed that acetone extracts were the highest in Tali (7.88%) and Bilinga (6.92%). MALDI-TOF results found 10 new chemical components out of 24 in Tali (saponins, flavonoids, glycosides, alkaloids and stilbenes) while 16 were identified as saponins, acids and flavonoids in Bilinga. Acetone-soluble extracts had the most significant effect on the hygroscopic nature of wood. Extracted wood recorded higher equilibrium moisture contents (EMC), monolayer moisture capacities and lower sorption differences. For native wood, water-repellent polyphenols like alkaloids and stilbenes and newly identified components there in, rendered Tali less hygroscopic than Bilinga. The nature of extractives would be an effective alternative to explore the chemical compounds, which can best improve the hygroscopic response of wood.

Induction and metabolomic analysis of hairy roots of Atractylodes lancea.

Atractylodes lancea is an important source of traditional Chinese medicines. Sesquiterpenoids are the key active compounds in A. lancea, and their presence determines the quality of the material. Hairy hoot (HR) culture is a potential method to produce medicinally active compounds industrially; however, the induction and metabolic profiling of A. lancea HR have not been reported. We found that optimal induction of A. lancea HR was achieved by Agrobacterium rhizogenes strain C58C1 using the young leaves of tissue culture seedlings in the rooting stage as explants. Ultra-performance liquid chromatography-tandem mass spectrometric analyses of the chemical compositions of HR and normal root (NR) led to the annotation of 1046 metabolites. Over 200 differentially accumulated metabolites were identified, with 41 found to be up-regulated in HR relative to NR and 179 down-regulated in HR. Specifically, atractylodin levels were higher in HR, while the levels of β-eudesmol and hinesol were higher in NR. Metabolic pathway analyses showed a significant difference in metabolites of the shikimate acid pathway between HR and NR. Five A. lancea compounds are potential biomarkers for evaluation of HR and NR quality. This study provides an important reference for the application of HR for the production of medicinally active compounds. KEY POINTS: • We established an efficient protocol for the induction of HR in A. lancea • HR was found to have a significantly higher amount of atractylodin than did NRs • Metabolic pathway analyses showed a significant difference in metabolites of the shikimate acid pathway between HR and NR.

Mass Spectrometric Analysis of Urinary N-Glycosylation Changes in Patients with Parkinson's Disease.

Urine is thought to provide earlier and more sensitive molecular changes for biomarker discovery than blood. Numerous glycoproteins, peptides, and free glycans are present in urine through glomerular filtration of plasma, cell shedding, apoptosis, proteolytic cleavage, and exosome secretion. Urine biomarkers have enormous diagnostic potential, and the use of these biomarkers is a long-standing practice. The discovery of non-urological disease biomarkers from urine is also gaining attention due to its non-invasive sample collection and ease of analysis. Abnormal protein glycosylation in plasma or cerebrospinal fluid has been associated with Parkinson's disease, however, whether urine with Parkinson's disease has characteristic glycosylation remains to be explored. Here, we use mass spectrometry-based glycomics and glycoproteomics approaches to analyze urine samples for glycans, glycosites, and intact glycopeptides of urine samples. Reduced abundance of N-glycans was detected at the level of total glycans as well as specific glycosites of glycopeptides. The most abundant N-glycan in urine is S(6)1H5N4F1; S(6)2H5N4 and N4H4F1 are highly present in serum and urine, and 10 biantennary galactosylated N-glycans in the urine of PD patients were significantly decreased. The downregulation of sialylation may be due to the reduction of ST3GAL2. Site-specific N-glycosylation analysis revealed that AMBP, UMOD, and RNase1 have PD-specific N-glycosylation sites. GO and KEGG analysis revealed that N-glycosylation changes may provide clues to identify disease-specific glycosylation biomarkers in Parkinson's disease.

Assessing the Activity under Different Physico-Chemical Conditions, Digestibility, and Innocuity of a GAPDH-Related Fish Antimicrobial Peptide and Analogs Thereof.

The antimicrobial activity of SJGAP (skipjack tuna GAPDH-related antimicrobial peptide) and four chemical analogs thereof was determined under different physicochemical conditions, including different pH values, the presence of monovalent and divalent cations, and after a heating treatment. The toxicity of these five peptides was also studied with hemolytic activity assays, while their stability under human gastrointestinal conditions was evaluated using a dynamic in vitro digestion model and chromatographic and mass spectrometric analyses. The antibacterial activity of all analogs was found to be inhibited by the presence of divalent cations, while monovalent cations had a much less pronounced impact, even promoting the activity of the native SJGAP. The peptides were also more active at acidic pH values, but they did not all show the same stability following a heat treatment. SJGAP and its analogs did not show significant hemolytic activity (except for one of the analogs at a concentration equivalent to 64 times that of its minimum inhibitory concentration), and the two analogs whose digestibility was studied degraded very rapidly once they entered the stomach compartment of the digestion model. This study highlights for the first time the characteristics of antimicrobial peptides from Scombridae or homologous to GAPDH that are directly related to their potential clinical or food applications.

Vascular endothelial growth factor B regulates insulin secretion in β cells of type 2 diabetes mellitus mice via PLCγ and the IP3R‑evoked Ca2+/CaMK2 signaling pathway.

Vascular endothelial growth factor B (VEGFB) plays a crucial role in glucolipid metabolism and is highly associated with type 2 diabetes mellitus (T2DM). The role of VEGFB in the insulin secretion of β cells remains unverified. Thus, the present study aimed to discuss the effect of VEGFB on regulating insulin secretion in T2DM development, and its underlying mechanism. A high‑fat diet and streptozocin (STZ) were used for inducing T2DM in mice model, and VEGFB gene in islet cells of T2DM mice was knocked out by CRISPR Cas9 and overexpressed by adeno‑Associated Virus (AAV) injection. The effect of VEGFB and its underlying mechanism was assessed by light microscopy, electron microscopy and fluorescence confocal microscopy, enzyme‑linked immunosorbent assay, mass spectrometer and western blot analysis. The decrement of insulin secretion in islet β cell of T2DM mice were aggravated and blood glucose remained at a high level after VEGFB knockout (KO). However, glucose tolerance and insulin sensitivity of T2DM mice were improved after the AAV‑VEGFB186 injection. VEGFB KO or overexpression can inhibit or activate PLCγ/IP3R in a VEGFR1‑dependent manner. Then, the change of PLCγ/IP3R caused by VEGFB/VEGFR1 will alter the expression of key factors on the Ca2+/CaMK2 signaling pathway such as PPP3CA. Moreover, VEGFB can cause altered insulin secretion by changing the calcium concentration in β cells of T2DM mice. These findings indicated that VEGFB activated the Ca2+/CaMK2 pathway via VEGFR1‑PLCγ and IP3R pathway to regulate insulin secretion, which provides new insight into the regulatory mechanism of abnormal insulin secretion in T2DM.

Exploring the Accumulation Behavior and Heterogeneity of Perfluorooctanesulfonic Acid in Zebrafish Primary Organ Cells by Single-Cell Mass Cytometry.

Perfluorooctanesulfonic acid (PFOS) is a commonly found environmental pollutant with potential toxicity and health risks to biosystems and ecosystems. Study of the accumulation behavior and heterogeneity of PFOS in biological primary organ cells provides us significant insights to explore its cytotoxicity, carcinogenicity, and mutagenicity. Here a single-cell mass cytometry system was established for the high-throughput analysis of trace PFOS and the exploration of its accumulation behavior and heterogeneity in zebrafish primary organ cells. The single-cell mass cytometry system applied a ∼25 μm constant-inner-diameter capillary as the single-cell generation and transportation channel with an etched tip-end of 40 μm as the nanoelectrospray emitter for mass spectrometric analysis. The single-cell mass cytometry system showed satisfactory semiquantitative performance and sensitivity for analysis of PFOS in single cells, with a high detection throughput of ∼35 cells/min. Subsequently, the liver, intestine, heart, and brain from PFOS-exposed zebrafish (100 pg/μL, 28 days) were dissociated and prepared as cell suspensions, and the cell suspensions were introduced into the single-cell mass cytometry system for high-throughput analysis of PFOS in individual primary organ cells. Significant cellular accumulation heterogeneities were observed, with the highest content in liver cells, followed by intestine cells, then heart cells, and the lowest in brain cells. In addition, the dynamics of PFOS in the zebrafish liver, intestine, heart, and brain cells showed typical violin plot distributions and were well-described using a gamma (γ) function.

A Micropillar Array Based Microfluidic Device for Rare Cell Detection and Single-Cell Proteomics.

Advancements in single-cell-related technologies have opened new possibilities for analyzing rare cells, such as circulating tumor cells (CTCs) and rare immune cells. Among these techniques, single-cell proteomics, particularly single-cell mass spectrometric analysis (scMS), has gained significant attention due to its ability to directly measure transcripts without the need for specific reagents. However, the success of single-cell proteomics relies heavily on efficient sample preparation, as protein loss in low-concentration samples can profoundly impact the analysis. To address this challenge, an effective handling system for rare cells is essential for single-cell proteomic analysis. Herein, we propose a microfluidics-based method that offers highly efficient isolation, detection, and collection of rare cells (e.g., CTCs). The detailed fabrication process of the micropillar array-based microfluidic device is presented, along with its application for CTC isolation, identification, and collection for subsequent proteomic analysis.

Linking meteorites to their asteroid parent bodies: The capabilities of dust analyzer instruments during asteroid flybys

AbstractLinking meteorites to their asteroid parent bodies remains an outstanding issue. Space‐based dust characterization using impact ionization mass spectrometry is a proven technique for the compositional analysis of individual cosmic dust grains. Here we investigate the feasibility of determining asteroid compositions via cation mass spectrometric analyses of their dust ejecta clouds during low (7–9 km s−1) velocity spacecraft flybys. At these speeds, the dust grain mass spectra are dominated by easily ionized elements and molecular species. Using known bulk mineral volume abundances, we show that it is feasible to discriminate the common meteorite classes of carbonaceous chondrites, ordinary chondrites, and howardite–eucrite–diogenite achondrites, as well as their subtypes, relying solely on the detection of elements with ionization efficiencies of ≤700 or ≤800 kJ mol−1, applicable to low (~7 km s−1) and intermediate (~9 km s−1) flyby speed scenarios, respectively. Including the detection of water ion groups enables greater discrimination between certain meteorite types, and flyby speeds ≥10 km s−1 enhance the diagnostic capabilities of this technique still further. Although additional terrestrial calibration is required, this technique may allow more unequivocal asteroid‐meteorite connections to be determined by spacecraft flybys, emphasizing the utility of dust instruments on future asteroid missions.

Differential Spectrum of Albumin Glycation, Oxidation, and Truncation in Type 2 and Type 1 Diabetes: Clinical and Biological Implications.

Background: Albumin, the most abundant and physiologically vital serum protein, accumulates a range of chemical modifications, as consequence of encounters with large number of reactive molecules whose concentrations increase in serum under pathological conditions. Methods: In a "proof of concept" study, mass spectrometric analysis was utilized to quantitate albumin post-translational modifications (glycation, oxidation, and truncation; individual isoforms and total) in four informative subject groups [type 1 diabetes (T1DM), type 2 diabetes (T2DM), prediabetes-obesity and healthy; all with estimated glomerular filtration rate ≥60 mL/(min·m2)]. Besides glycated albumin (GA/mass spectrometry), glycated serum protein (GSP/nitro blue tetrazolium colorimetry), and glycated hemoglobin (HbA1c/high-performance liquid chromatography) were also measured. Results: A wide spectrum of albumin molecular modifications was identified in diabetes, with significant differences between T2DM and T1DM. Albumin glycation: GA correlated more strongly with HbA1c in T1DM, compared to T2DM. Higher albumin glycation isoforms (human serum albumin +3G/2G) were more stable and discriminative markers of mean glycemia. Albumin oxidation: T2DM, in comparison with T1DM, showed enhanced oxidative and dual (glycation plus oxidation) modifications, representing extreme molecular pathology. Albumin truncation: There was dramatic reduction ("deficiency") of truncated albumin isoforms in T2DM, and significant reduction in T1DM. Albumin truncation negatively correlated with severity of albumin glycation (mean glycemia) and albumin oxidation (cysteinylation). Possible mechanisms of insulin resistance, with associated increased free fatty acids binding to albumin, in stimulating albumin oxidation and inhibiting albumin glycation ("metabolic cross talks") are reviewed. Conclusions: Albumin molecular modifications in diabetes, together with significant differences between T2DM and T1DM, suggest possible role for insulin resistance in their genesis and consequent cell, tissue, and vascular dysfunction/damage. Albumin molecular fingerprinting can provide valuable insights into pathogenesis, diagnosis, monitoring, and future therapies for diabetes. Identification of biomarker battery ("albuminomics," "diabetomics") driven diverse "healthy," prediabetes, obesity, and T2DM phenotypes represents additional novel step toward precision medicine in diabetes and related disorders.