Endogenous Inflammatory Mediators Produced by Injury Activate TRPV1 and TRPA1 Nociceptors to Induce Sexually Dimorphic Cold Pain That Is Dependent on TRPM8 and GFRα3.

The detection of environmental temperatures is critical for survival, yet inappropriate responses to thermal stimuli can have a negative impact on overall health. The physiological effect of cold is distinct among somatosensory modalities in that it is soothing and analgesic, but also agonizing in the context of tissue damage. Inflammatory mediators produced during injury activate nociceptors to release neuropeptides, such as calcitonin gene-related peptide (CGRP) and substance P, inducing neurogenic inflammation, which further exasperates pain. Many inflammatory mediators induce sensitization to heat and mechanical stimuli but, conversely, inhibit cold responsiveness, and the identity of molecules inducing cold pain peripherally is enigmatic, as are the cellular and molecular mechanisms altering cold sensitivity. Here, we asked whether inflammatory mediators that induce neurogenic inflammation via the nociceptive ion channels TRPV1 (vanilloid subfamily of transient receptor potential channel) and TRPA1 (transient receptor potential ankyrin 1) lead to cold pain in mice. Specifically, we tested cold sensitivity in mice after intraplantar injection of lysophosphatidic acid or 4-hydroxy-2-nonenal, finding that each induces cold pain that is dependent on the cold-gated channel transient receptor potential melastatin 8 (TRPM8). Inhibition of CGRP, substance P, or toll-like receptor 4 (TLR4) signaling attenuates this phenotype, and each neuropeptide produces TRPM8-dependent cold pain directly. Further, the inhibition of CGRP or TLR4 signaling alleviates cold allodynia differentially by sex. Last, cold pain induced by both inflammatory mediators and neuropeptides requires TRPM8, as well as the neurotrophin artemin and its receptor GDNF receptor α3 (GFRα3). These results are consistent with artemin-induced cold allodynia requiring TRPM8, demonstrating that neurogenic inflammation alters cold sensitivity via localized artemin release that induces cold pain via GFRα3 and TRPM8.SIGNIFICANCE STATEMENT The cellular and molecular mechanisms that generate pain are complex with a diverse array of pain-producing molecules generated during injury that act to sensitize peripheral sensory neurons, thereby inducing pain. Here we identify a specific neuroinflammatory pathway involving the ion channel TRPM8 (transient receptor potential cation channel subfamily M member 8) and the neurotrophin receptor GFRα3 (GDNF receptor α3) that leads to cold pain, providing select targets for potential therapies for this pain modality.

Serological Exosome Metabolic Biopsy of Hepatocellular Carcinoma via Designed Core-Shell Nanoparticles.

Exosome metabolite-based liquid biopsy is a promising strategy for large-scale application in practical clinics toward precise medicine. Given the current challenges in successive isolation and analysis of exosomes and their metabolites in this field, we established a low-cost, high-throughput, and rapid platform for serological exosome metabolic biopsy of hepatocellular carcinoma (HCC) via designed core-shell nanoparticles. It starts with the efficient extraction of high-quality serum exosomes and exosome metabolic features, based on which significantly obvious sample clusters are observed by unsupervised cluster analysis. The following integration of feature selection and supervised machine learning enables the identification of six key metabolites and achieves high-performance prediction between HCC, liver cirrhosis, and healthy controls. Specifically, both sensitivity and accuracy achieve 100% among any pairwise intergroup discrimination in a blind test. The quality and reliability of six key metabolites are further evaluated and validated by using different machine learning algorithms and pathway exploration. Our platform contributes to the future growth of new liquid biopsy technologies for precision diagnosis and real-time monitoring of HCC, among other conditions.

Designed Directional Growth of Ti-Metal-Organic Frameworks for Decoding Alzheimer's Disease-Specific Exosome Metabolites.

Exosomes, a growing focus for liquid biopsies, contain diverse molecular cargos. In particular, exosome metabolites with valuable information have exhibited great potential for improving the efficiency of liquid biopsies for addressing complex medical conditions. In this work, we design the directional growth of Ti-metal-organic frameworks on polar-functionalized magnetic particles. This design facilitates the rapid synergistic capture of exosomes with the assistance of an external magnetic field and additionally synergistically enhances the ionization of their metabolites during mass spectrometry detection. Benefiting from this dual synergistic effect, we identified three high-performance exosome metabolites through the differential comparison of a large number of serum samples from individuals with Alzheimer's disease (AD) and normal cognition. Notably, the accuracy of AD identification ranges from 93.18 to 100% using a single exosome metabolite and reaches a flawless 100% with three metabolites. These findings emphasize the transformative potential of this work to enhance the precision and reliability of AD diagnosis, ushering in a new era of improved diagnostic accuracy.

Electromagnetic Field-Assisted Frozen Tissue Planarization Enhances MALDI-MSI in Plant Spatial Omics.

Plant samples with irregular morphology are challenging for longitudinal tissue sectioning. This has restricted the ability to gain insight into some plants using matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI). Herein, we develop a novel technique termed electromagnetic field-assisted frozen tissue planarization (EMFAFTP). This technique involves using a pair of adjustable electromagnets on both sides of a plant tissue. Under an optimized electromagnetic field strength, nondestructive planarization and regularization of the frozen tissue is induced, allowing the longitudinal tissue sectioning that favors subsequent molecular profiling by MALDI-MSI. As a proof of concept, flowers, leaves and roots with irregular morphology from six plant species are chosen to evaluate the performance of EMFAFTP for MALDI-MSI of secondary metabolites, amino acids, lipids, and proteins among others in the plant samples. The significantly enhanced MALDI-MSI capabilities of these endogenous molecules demonstrate the robustness of EMFAFTP and suggest it has the potential to become a standard technique for advancing MALDI-MSI into a new era of plant spatial omics.

2-Hydroxy-5-nitro-3-(trifluoromethyl)pyridine as a Novel Matrix for Enhanced MALDI Imaging of Tissue Metabolites.

Matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI), which is a label-free imaging technique, determines the spatial distribution and relative abundance of versatile endogenous metabolites in tissues. Meanwhile, matrix selection is generally regarded as a pivotal step in MALDI tissue imaging. This study presents the first report of a novel MALDI matrix, 2-hydroxy-5-nitro-3-(trifluoromethyl)pyridine (HNTP), for the in situ detection and imaging of endogenous metabolites in rat liver and brain tissues by MALDI-MS in positive-ion mode. The HNTP matrix exhibits excellent characteristics, including strong ultraviolet absorption, µm-scale matrix crystals, high chemical stability, low background ion interference, and high metabolite ionization efficiency. Notably, the HNTP matrix also shows superior detection capabilities, successfully showing 185 detectable metabolites in rat liver tissue sections. This outperforms the commonly used matrices of 2,5-dihydroxybenzoic acid and 2-mercaptobenzothiazole, which detect 145 and 120 metabolites from the rat liver, respectively. Furthermore, a total of 152 metabolites are effectively detected and imaged in rat brain tissue using the HNTP matrix, and the spatial distribution of these compounds clearly shows the heterogeneity of the rat brain. The results demonstrate that HNTP is a new and powerful positive-ion mode matrix to enhance the analysis of metabolites in biological tissues by MALDI-MSI.

Straightforward Creation of Multishell Hollow Hybrids for an Integrated Metabolic Monitoring System in Disease Management.

Multidimensional metabolic analysis has become a new trend in establishing efficient disease monitoring systems, as the constraints associated with relying solely on a single dimension in refined monitoring are increasingly pronounced. Here, coordination polymers are employed as derivative precursors to create multishell hollow hybrids, developing an integrated metabolic monitoring system. Briefly, metabolic fingerprints are extracted from hundreds of serum samples and urine samples, encompassing not only membranous nephropathy but also related diseases, using high-throughput mass spectrometry. With optimized algorithm and initial feature selection, the established combined panel demonstrates enhanced accuracy in both subtype differentiation (over 98.1%) and prognostic monitoring (over 95.6%), even during double blind test. This surpasses the serum biomarker panel (≈90.7% for subtyping, ≈89.7% for prognosis) and urine biomarker panel (≈94.4% for subtyping, ≈76.5% for prognosis). Moreover, after attempting to further refine the marker panel, the blind test maintains equal sensitivity, specificity, and accuracy, showcasing a comprehensive improvement over the single-fluid approach. This underscores the remarkable effectiveness and superiority of the integrated strategy in discriminating between MN and other groups. This work has the potential to significantly advance diagnostic medicine, leading to the establishment of more effective strategies for patient management.

High precision microwave measurement based on nitrogen-vacancy color center and application in velocity detection.

Wide-range high-precision velocity detection with nitrogen-vacancy (NV) color center has been realized. By treating the NV color center as a mixer, the high-precision microwave measurement is realized. Through optimization of acquisition time, the microwave frequency resolution is improved to the mHz level. Combined with the frequency-velocity conversion model, velocity detection is realized in the range of 0-100 cm/s, and the velocity resolution is up to 0.012 cm/s. The maximum deviation in repeated measurements does not exceed 1/1000. Finally, combined with the multiplexed microwave reference technique, the range of velocity can be extended to 7.4 × 105 m/s. All of the results provide reference for high-precision velocity detection and play a significant role in various domains of quantum precision measurement. This study provides a crucial technical foundation for the development of high-dynamic-range velocity detectors and novel quantum precision velocity measurement technologies.

A quick and effective method for thermostability differentiation in cucumber (Cucumis sativus L.).

High temperature affects the growth and production of cucumber. Selecting thermotolerant cucumber cultivars is conducive to coping with high temperatures and improving production. Thus, a quick and effective method for screening thermotolerant cucumber cultivars is needed. In this study, four cucumber cultivars were used to identify heat resistance indexes. The morphological, physiological and biochemical indexes were measured. When exposed to high temperatures, thermotolerant cucumber had a more stable photosystem, membrane, and oxidation-reduction systems. The impact of high temperatures on plants is multifaceted, and the accurate discrimination of heat resistance cannot be achieved solely based on a single or multiple indicators. Therefore, principal component analysis (PCA) was employed to comprehensively evaluate the heat resistance of cucumber plants. The results showed that the heat resistance obtained by PCA was significantly correlated with the heat injury index. In addition, the stepwise regression equation identified two heat-related indices, hydrogen peroxide content (H2 O2 ) and photosynthetic operating efficiency (Fq'/Fm'), and they can quickly distinguish the heat resistance of the other 8 cucumber cultivars. These results will help to accelerate the selection of thermotolerant resources and assist in cucumber breeding.

Immunoglobulin somatic hypermutation in a defined biochemical system recapitulates affinity maturation and permits antibody optimization.

We describe a purified biochemical system to produce monoclonal antibodies (Abs) in vitro using activation-induced deoxycytidine deaminase (AID) and DNA polymerase η (Polη) to diversify immunoglobulin variable gene (IgV) libraries within a phage display format. AID and Polη function during B-cell affinity maturation by catalyzing somatic hypermutation (SHM) of immunoglobulin variable genes (IgV) to generate high-affinity Abs. The IgV mutational motif specificities observed in vivo are conserved in vitro. IgV mutations occurred in antibody complementary determining regions (CDRs) and less frequently in framework (FW) regions. A unique feature of our system is the use of AID and Polη to perform repetitive affinity maturation on libraries reconstructed from a preceding selection step. We have obtained scFv Abs against human glucagon-like peptide-1 receptor (GLP-1R), a target in the treatment of type 2 diabetes, and VHH nanobodies targeting Fatty Acid Amide Hydrolase (FAAH), involved in chronic pain, and artemin, a neurotropic factor that regulates cold pain. A round of in vitro affinity maturation typically resulted in a 2- to 4-fold enhancement in Ab-Ag binding, demonstrating the utility of the system. We tested one of the affinity matured nanobodies and found that it reduced injury-induced cold pain in a mouse model.

Genes in human obesity loci are causal obesity genes in C. elegans.

Obesity and its associated metabolic syndrome are a leading cause of morbidity and mortality. Given the disease's heavy burden on patients and the healthcare system, there has been increased interest in identifying pharmacological targets for the treatment and prevention of obesity. Towards this end, genome-wide association studies (GWAS) have identified hundreds of human genetic variants associated with obesity. The next challenge is to experimentally define which of these variants are causally linked to obesity, and could therefore become targets for the treatment or prevention of obesity. Here we employ high-throughput in vivo RNAi screening to test for causality 293 C. elegans orthologs of human obesity-candidate genes reported in GWAS. We RNAi screened these 293 genes in C. elegans subject to two different feeding regimens: (1) regular diet, and (2) high-fructose diet, which we developed and present here as an invertebrate model of diet-induced obesity (DIO). We report 14 genes that promote obesity and 3 genes that prevent DIO when silenced in C. elegans. Further, we show that knock-down of the 3 DIO genes not only prevents excessive fat accumulation in primary and ectopic fat depots but also improves the health and extends the lifespan of C. elegans overconsuming fructose. Importantly, the direction of the association between expression variants in these loci and obesity in mice and humans matches the phenotypic outcome of the loss-of-function of the C. elegans ortholog genes, supporting the notion that some of these genes would be causally linked to obesity across phylogeny. Therefore, in addition to defining causality for several genes so far merely correlated with obesity, this study demonstrates the value of model systems compatible with in vivo high-throughput genetic screening to causally link GWAS gene candidates to human diseases.

Associations and attributable burdens in late-life exposure to PM2.5 and its major components and depressive symptoms in middle-aged and older adults: A nationwide cohort study.

BACKGROUND: Depression in late life has been associated with reduced quality of life and increased mortality. Whether the chronic fine particular matter (PM2.5) and its components exposure are contributed to the older depression symptoms remains unclear. METHOD: Middle-aged and older adults (>45 years) were selected from the China Health and Retirement Longitudinal Study during the four waves of interviews. The concentrations of PM2.5 and its major constituents were calculated using near real-time data at a spatial resolution of 10 km during the study period. The depressive symptom was evaluated by the Depression Center for Epidemiologic Studies Depression (CES-D)-10 score. The fix-effect model was applied to evaluate the association between PM2.5 and its major constituents with depressive symptoms. Three three-step methods were used to explore the modification role of sleep duration against the depressive symptoms caused by PM2.5 exposure. RESULTS: In our study, a total of 52,683 observations of 16,681 middle-aged and older adults were assessed. Each interquartile range (IQR) level of PM2.5 concentration exposure was longitudinally associated with a 2.6 % (95 % confidence interval [CI]: 1.3 %, 4.0 %) increase in the depression CES-D-10 score. Regarding the major components of PM2.5, OM, NO3-, and NH4+ showed the leading toxicity effects, which could increase the depression CES-D-10 score by 2.2 % (95 %CI: 1.0 %, 3.4 %), 2.2 % (0.6 %, 3.9 %), and 2.0 % (95 %CI: 0.6 %, 3.4 %) correspondingly. Besides, males were more susceptible to the worse depressive symptoms caused by PM2.5 and its major components exposure than female subpopulations. Shortened sleep duration might be the mediator of PM2.5-associated depressive symptoms. CONCLUSION: Our results suggest that long-term exposure to PM2.5 and its major components were associated with an increased risk for depressive symptoms in middle-aged and older adults. Reducing the leading components of PM2.5 may cost-effectively alleviate the disease burden of depression and promote healthy longevity in heavy pollutant countries.

A nomogram for predicting choledochal cyst with perforation.

BACKGROUND: Choledochal cyst with perforation (CC with perforation) rarely occurs, early diagnosis and timely treatment plan are crucial for the treatment of CC with perforation. This study aims to forecast the occurrence of CC with perforation. METHODS: All 1111 patients were conducted, who underwent surgery for choledochal cyst at our hospital from January 2011 to October 2022. We conducted univariate and multivariate logistic regression analysis to screen for independent predictive factors for predicting CC with perforation, upon which established a nomogram. The predictive performance of the nomogram was evaluated using receiver operating characteristic (ROC) curves, calibration plots, and decision curve analysis (DCA) curves. RESULTS: The age of children with choledochal cyst perforation is mainly concentrated between 1 and 3 years old. Logistic regression analysis indicates that age, alanine aminotransferase, glutamyl transpeptidase, C-reactive protein, vomiting, jaundice, abdominal distension, and diarrhea are associated with predicting the occurrence of choledochal cyst perforation. ROC curves, calibration plots, and DCA curve analysis curves demonstrate that the nomogram has great discriminative ability and calibration, as well as significant clinical utility. CONCLUSION: The age of CC with perforation is mainly concentrated between 1 and 3 years old. A nomogram for predicting the perforation of choledochal cyst was established.

A derivative of trihydroxynaphthalenone and a pyrone metabolite from the endophytic fungus Talaromyces purpurpgenus.

A newly discovered trihydroxynaphthalenone derivative, epoxynaphthalenone (1) involving the condensation of ortho-hydroxyl groups into an epoxy structure, and a novel pyrone metabolite characterized as pyroneaceacid (2), were extracted from Talaromyces purpurpgenus, an endophytic fungus residing in Rhododendron molle. The structures of these compounds were elucidated through a comprehensive analysis of their NMR and HRESIMS data. The determination of absolute configurations was accomplished using electronic circular dichroism (ECD) calculations and CD spectra. Notably, these recently identified metabolites exhibited a moderate inhibitory activity against xanthine oxidase (XOD).

Bi-allelic Loss-of-function Variants in CFAP58 Cause Flagellar Axoneme and Mitochondrial Sheath Defects and Asthenoteratozoospermia in Humans and Mice.

Multiple morphological abnormalities of the sperm flagella (MMAF) is a severe form of asthenoteratozoospermia. Although recent studies have revealed several MMAF-associated genes and demonstrated MMAF to be a genetically heterogeneous disease, at least one-third of the cases are still not well understood for their etiology. Here, we identified bi-allelic loss-of-function variants in CFAP58 by using whole-exome sequencing in five (5.6%) unrelated individuals from a cohort of 90 MMAF-affected Chinese men. Each of the men harboring bi-allelic CFAP58 variants presented typical MMAF phenotypes. Transmission electron microscopy demonstrated striking flagellar defects with axonemal and mitochondrial sheath malformations. CFAP58 is predominantly expressed in the testis and encodes a cilia- and flagella-associated protein. Immunofluorescence assays showed that CFAP58 localized at the entire flagella of control sperm and predominantly concentrated in the mid-piece. Immunoblotting and immunofluorescence assays showed that the abundances of axoneme ultrastructure markers SPAG6 and SPEF2 and a mitochondrial sheath protein, HSP60, were significantly reduced in the spermatozoa from men harboring bi-allelic CFAP58 variants. We generated Cfap58-knockout mice via CRISPR/Cas9 technology. The male mice were infertile and presented with severe flagellar defects, consistent with the sperm phenotypes in MMAF-affected men. Overall, our findings in humans and mice strongly suggest that CFAP58 plays a vital role in sperm flagellogenesis and demonstrate that bi-allelic loss-of-function variants in CFAP58 can cause axoneme and peri-axoneme malformations leading to male infertility. This study provides crucial insights for understanding and counseling of MMAF-associated asthenoteratozoospermia.

2,5-Dihydroxyterephthalic Acid: A Matrix for Improved Detection and Imaging of Amino Acids.

Amino acids (AAs), which are low-molecular-weight (low-MW) metabolites, serve as essential building blocks not only for protein synthesis but also for maintaining the nitrogen balance in living systems. In situ detection and imaging of AAs are crucial for understanding more complex biological processes. Matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI) is a label-free mass spectrometric imaging technique that enables the simultaneous detection and imaging of the spatial distribution and relative abundance of different endogenous/exogenous compounds in biological samples. The excellent efficiency of MALDI-MSI is attributed to the choice of the MALDI matrix. However, to the best of our knowledge, no matrix has been specifically developed for AAs. Herein, we report a MALDI matrix, 2,5-dihydroxyterephthalic acid (DHT), which can improve the detection and imaging of AAs in biological samples by MALDI-MS. Our results indicated that DHT exhibited strong ultraviolet-visible (UV-vis) absorption, uniform matrix deposition, and high vacuum stability. Moreover, the matrix-related ion signals produced from DHT were reduced by 50 and 71.8% at m/z < 500 compared to the commonly used matrices of 2,5-dihydroxybenzoic acid (DHB) and α-cyano-4-hydroxycinnamic acid (CHCA), respectively, in their respective organic solvents. In terms of quantitative performance, arginine, glutamic acid, glutamine, and proline can be detected with limits of detection of 6, 4, 6, and 4 ng/mL, respectively, using the DHT as the matrix. Using DHT as the matrix, all 20 protein AAs were successfully detected in human serum by MALDI-MS, whereas only 7 and 10 AAs were detected when DHB and CHCA matrices were used, respectively. Furthermore, 20 protein AAs and taurine were successfully detected and imaged in a section of edible Crassostrea gigas (oyster) tissue for the first time. Our study demonstrates that using DHT as a matrix can improve the detection and imaging of AAs in biological samples by MALDI-MS.

Engineering Unsymmetrically Coordinated Fe Sites via Heteroatom Pairs Synergetic Contribution for Efficient Oxygen Reduction.

Single-atom Fe catalysts are considered as the promising catalysts for oxygen reduction reaction (ORR). However, the high electronegativity of the symmetrical coordination N atoms around Fe site generally results in too strong adsorption of *OOH intermediates on the active site, severely limiting the catalytic performance. Herein, a "heteroatom pair synergetic modulation" strategy is proposed to tailor the coordination environment and spin state of Fe sites, enabling breaking the shackles of unsuitable adsorption of intermediate products on the active centers toward a more efficient ORR pathway. The unsymmetrically Co and B heteroatomic coordinated Fe single sites supported on an N-doped carbon (Fe─B─Co/NC) catalyst perform excellent ORR activity with high half-wave potential (E1/2 ) of 0.891 V and a large kinetic current density (Jk ) of 60.6 mA cm-2 , which is several times better than those of commercial Pt/C catalysts. By virtue of in situ electrochemical impedance and synchrotron infrared spectroscopy, it is observed that the optimized Fe sites can effectively accelerate the evolution of O2 into the *O intermediate, overcoming the sluggish O─O bond cleavage of the *OOH intermediate, which is responsible for fast four-electron reaction kinetics.

Monitoring surface dynamics of electrodes during electrocatalysis using in situ synchrotron FTIR spectroscopy.

Monitoring the surface dynamics of catalysts under working conditions is important for a deep understanding of the underlying electrochemical mechanisms towards efficient energy conversion and storage. Fourier transform infrared (FTIR) spectroscopy with high surface sensitivity has been considered as a powerful tool for detecting surface adsorbates, but it faces a great challenge when being adopted in surface dynamics investigations during electrocatalysis due to the complication and influence of aqueous environments. This work reports a well designed FTIR cell with tunable micrometre-scale water film over the surface of working electrodes and dual electrolyte/gas channels for in situ synchrotron FTIR tests. By coupling with a facile single-reflection infrared mode, a general in situ synchrotron radiation FTIR (SR-FTIR) spectroscopic method is developed for tracking the surface dynamics of catalysts during the electrocatalytic process. As an example, in situ formed key *OOH is clearly observed on the surface of commercial benchmark IrO2 catalysts during the electrochemical oxygen evolution process based on the developed in situ SR-FTIR spectroscopic method, which demonstrates its universality and feasibility in surface dynamics studies of electrocatalysts under working conditions.

High-throughput MALDI-MSI metabolite analysis of plant tissue microarrays.

A novel metabolomics analysis technique, termed matrix-assisted laser desorption/ionization mass spectrometry imaging-based plant tissue microarray (MALDI-MSI-PTMA), was successfully developed for high-throughput metabolite detection and imaging from plant tissues. This technique completely overcomes the disadvantage that metabolites cannot be accessible on an intact plant tissue due to the limitations of the special structures of plant cells (e.g. epicuticular wax, cuticle and cell wall) through homogenization of plant tissues, preparation of PTMA moulds and matrix spraying of PTMA sections. Our study shows several properties of MALDI-MSI-PTMA, including no need of sample separation and enrichment, high-throughput metabolite detection and imaging (>1000 samples per day), high-stability mass spectrometry data acquisition and imaging reconstruction and high reproducibility of data. This novel technique was successfully used to quickly evaluate the effects of two plant growth regulator treatments (i.e. 6-benzylaminopurine and N-phenyl-N'-1,2,3-thiadiazol-5-ylurea) on endogenous metabolite expression in plant tissue culture specimens of Dracocephalum rupestre Hance (D. rupestre). Intra-day and inter-day evaluations indicated that the metabolite data detected on PTMA sections had good reproducibility and stability. A total of 312 metabolite ion signals in leaves tissues of D. rupestre were detected, of which 228 metabolite ion signals were identified, they were composed of 122 primary metabolites, 90 secondary metabolites and 16 identified metabolites of unknown classification. The results demonstrated the advantages of MALDI-MSI-PTMA technique for enhancing the overall detection ability of metabolites in plant tissues, indicating that MALDI-MSI-PTMA has the potential to become a powerful routine practice for high-throughput metabolite study in plant science.

Identification method of raindrops and hailstones based on digital holographic interference.

The identification of raindrops and hailstones is of great significance to the study of precipitation characteristics from the aspect of microphysics and can provide important data support for weather modification. In this paper, an identification method of raindrops and hailstones based on digital holographic interference is proposed. The grayscale gradient variance method is used to obtain the focus position of the particles. By means of binarization and morphological processing, digital holograms are processed to obtain clear profiles of the particles. Then the contour parameters of the particles are used to obtain the equivalent volume diameter and roundness. Finally, according to the equivalent volume diameter, roundness and lens-like effect of the particles, the phase states of the raindrop and hailstone are identified by the algorithm. Experiments show that the method proposed in this paper has a good identification effect on raindrops and hailstones. The research results can provide reference for the research of the identification method of raindrops and hailstones and the acquisition of accurate characteristic parameters.

A concentration-descending washing strategy with methanol for the enhancement of protein imaging in biological tissues by MALDI-MS.

Matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI) is a powerful approach that has been widely used for in situ detection of various endogenous compounds in tissues. However, there are still challenges with in situ analysis of proteins using MALDI-MSI due to the ion suppression effects of small molecules in tissue sections. Therefore, tissue-washing steps are crucial for protein MALDI tissue imaging to remove these interfering molecules. Here, we successfully developed a new method named the concentration-descending washing strategy (CDWS) with methanol (MeOH), i.e., washing of biological tissue with 100%, 95%, and 70% MeOH solutions, for the enhancement of endogenous in situ protein detection and imaging in tissues using MALDI-MS. The method of MeOH-based CDWS (MeOH-CDWS) led to the successful in situ detection of 272 ± 3, 185 ± 4, and 134 ± 2 protein ion signals from rat liver, rat brain, and germinating Chinese-yew seed tissue sections, respectively. By comparison, 161 ± 2, 121 ± 1, and 114 ± 2 protein ions were detected by three commonly used methods, i.e., Carnoy's wash, ethanol (EtOH)-based CAWS (i.e., concentration-ascending washing strategy, 70% EtOH followed by 90% EtOH/9% AcOH), and isopropanol (iPrOH)-based CAWS (70% iPrOH followed by 95% iPrOH), respectively, in rat liver tissue sections, indicating that 68.9 ± 3.1%, 124.8 ± 3.3%, and 138.6 ± 4.4% more protein ion signals could be detected by the use of MeOH-CDWS than the three abovementioned washing strategies. Our results show that the use of MeOH-CDWS improves the performance of MALDI-MSI for in situ protein detection such as the number and intensity of proteins. The use of MeOH-CDWS improves the fixation of proteins and thus reduces the loss of proteins, which significantly reduces protein delocalization in tissue and enhances the performance of MALDI tissue imaging of protein. Thus, the use of MeOH-CDWS improves the quality of protein images in tissue sections through MALDI-MSI and has the potential to be used as standard practice for MALDI tissue imaging of proteins.