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In this work, an economical and easy-to-use microcapsule array fabricated by ice printing technique has been realized for ultrasensitive fluorescence quantification of copper ions employing functional nucleic acid strategy. With ice printing, the detection reagents are sealed by polystyrene (PS) film isolation and photopolymer, which guarantees a stable and contamination-free environment for functional nucleic acid reaction. Our microcapsule arrays have shown long-term stability (20 days) under −20 °C storage in frozen form before use. During the Cu 2+ on-site detection, 1 μL sample is simply injected into the thawy microcapsule by a microliter syringe under room temperature, and after 20 minutes the fluorescence result can be obtained by an LED transilluminator. This method can realize the detection limit to 100 nM (100 fmol/μL) with high specificity.

Background Breast tumorigenesis is a complex and multistep process accompanied by both genetic and epigenetic dysregulation. In contrast to the extensive studies on DNA epigenetic modifications 5-hydroxymethylcytosine (5hmC) and 5-methylcytosine (5mC) in malignant breast tumors, their roles in the early phases of breast tumorigenesis remain ambiguous. Results DNA 5hmC and 5mC exhibited a consistent and significant decrease from usual ductal hyperplasia to atypical ductal hyperplasia and subsequently to ductal carcinoma in situ (DCIS). However, 5hmC showed a modest increase in invasive ductal breast cancer compared to DCIS. Genomic analyses showed that the changes in 5hmC and 5mC levels occurred around the transcription start sites (TSSs), and the modification levels were strongly correlated with gene expression levels. Meanwhile, it was found that differentially hydroxymethylated regions (DhMRs) and differentially methylated regions (DMRs) were overlapped in the early phases and accompanied by the enrichment of active histone marks. In addition, TET2-related DNA demethylation was found to be involved in breast tumorigenesis, and four transcription factor binding sites (TFs: ESR1, FOXA1, GATA3, FOS) were enriched in TET2-related DhMRs/DMRs. Intriguingly, we also identified a certain number of common DhMRs between tumor samples and cell-free DNA (cfDNA). Conclusions Our study reveals that dynamic changes in DNA 5hmC and 5mC play a vital role in propelling breast tumorigenesis. Both TFs and active histone marks are involved in TET2-related DNA demethylation. Concurrent changes in 5hmC signals in primary breast tumors and cfDNA may play a promising role in breast cancer screening.

Edible starch-based film was developed for packaging seasoning applied in instant noodles. The edible film can quickly dissolve into hot water so that the seasoning bag can mix in the soup of instant noodles during preparation. To meet the specific requirements of the packaging, such as reasonable high tensile properties, ductility under arid conditions, and low gas permeability, hydroxypropyl cornstarch with various edible additives from food-grade ingredients were applied to enhance the functionality of starch film. In this work, xylose was used as a plasticizer, cellulose crystals were used as a reinforcing agent, and laver was used to decrease gas permeability. The microstructures, interface, and compatibility of various components and film performance were investigated using an optical microscope under polarized light, scanning electron microscope, gas permeability, and tensile testing. The relationship was established between processing methodologies, microstructures, and performances. The results showed that the developed starch-based film have a modulus of 960 MPa, tensile strength of 36 Mpa with 14% elongation, and water vapor permeability less than 5.8 g/m2.h under 20% RH condition at room temperature (25 °C), which meets the general requirements of the flavor bag packaging used in instant noodles.

Purpose Sleep staging is critical for assessing sleep quality and diagnosing sleep disorders. However, manual annotation is both time-consuming and labor-intensive, highlighting the need for efficient automated solutions. This study proposes Kolmogorov–Arnold Networks (KAN)-SleepNet, a hybrid deep learning model designed for automated sleep stage classification using single-channel electroencephalogram (EEG) signals. Methods The KAN-SleepNet consists of two primary components: (1) a ConvKAN block that integrates convolutional neural networks with KAN to effectively extract discriminative features from EEG signals, and (2) a Bidirectional Long Short-Term Memory layer to capture temporal dependencies across sleep stages. The model was trained and evaluated using two publicly available datasets: the SleepEDF-78 dataset, comprising 153 recordings from 78 subjects, and the ISRUC-S1 dataset, consisting of 100 recordings from 100 subjects. SleepEDF-78 was annotated according to the Rechtschaffen and Kales criteria, whereas ISRUC-S1 followed the American Academy of Sleep Medicine guidelines. Performance was assessed using accuracy, F1-score, and Cohen's Kappa (κ), and results were compared against baseline models, including SleepEEGNet, DeepSleepNet, TinySleepNet, AttnSleep, GraphSleepNet, and MSTGCN. Results Experimental results demonstrate that KAN-SleepNet outperforms existing baseline models across both datasets (all p < 0.05 except vs. AttnSleep, p = 0.051). The KAN-SleepNet achieved an accuracy of 85.1%, F1-score of 80.0%, and Kappa of 0.792 on SleepEDF-78, and an accuracy of 82.8%, F1-score of 80.5%, and Kappa of 0.778 on ISRUC-S1. The model also exhibited strong performance in the challenging N1 stage, with F1-scores of 53.2% and 57.4% on SleepEDF-78 and ISRUC-S1, respectively. Conclusion The KAN-SleepNet demonstrates superior performance in automated sleep staging, highlighting its potential as an efficient and supportive tool for clinical sleep analysis.

As a model pathogen, Salmonella invades both phagocytic and non-phagocytic host cells and adopts an intracellular lifestyle in a membrane-bound compartment during infection. Therefore, a systemic overview of Salmonella adaptations to distinct host cells together with host remodeling will assist us in charting the landscape of host-pathogen interactions. Central to the Salmonella-host interplay are bacterial virulence factors (effectors) that are injected into host cells by type III secretion systems (T3SSs). Despite great progress, functional studies of bacterial effectors have experienced daunting challenges as well. In the last decade, mass spectrometry-based proteomics has evolved into a powerful technological platform that can quantitatively measure thousands of proteins in terms of their expression as well as post-translational modifications. Here, we will review the applications of high-throughput proteomic technologies in understanding the dynamic reprogramming of both Salmonella and host proteomes during the course of infection. Furthermore, we will summarize the progress in utilizing affinity purification-mass spectrometry to screen for host substrates of Salmonella T3SS effectors. Finally, we will critically discuss some limitations/challenges with current proteomic platforms in the context of host-pathogen interactions and highlight some emerging technologies that may offer the promise of tackling these problems.

Activated B cells increase central carbon metabolism to fulfill their bioenergetic demands, yet the mechanistic basis for this, as well as metabolic regulation in B cells, remains largely unknown. Here, we demonstrate that B-cell activation reprograms the tricarboxylic acid cycle and boosts the expression of fumarate hydratase (FH), leading to decreased cellular fumarate abundance. Fumarate accumulation by FH inhibition or dimethyl-fumarate treatment suppresses B-cell activation, proliferation and antibody production. Mechanistically, fumarate is a covalent inhibitor of tyrosine kinase LYN, a key component of the BCR signaling pathway. Fumarate can directly succinate LYN at C381 and abrogate LYN activity, resulting in a block to B-cell activation and function in vitro and in vivo. Therefore, our findings uncover a previously unappreciated metabolic regulation of B cells, and reveal LYN is a natural sensor of fumarate, connecting cellular metabolism to B-cell antigen receptor signaling.B-cell activation reprograms the TCA cycle and reduces cellular fumarate levels. Increased fumarate caused by fumarase inhibition or dimethyl-fumarate treatment directly succinates and inhibits LYN, leading to impaired B-cell activation and function.

The online combination of capillary electrophoresis (CE) with mass spectrometry (MS) has long been desired for the capability of direct and simultaneous separation and detection with high efficiency, accuracy, and throughput. In this work, a novel CE-MS interface was developed, using dielectric barrier discharge ionization (DBDI). The interface employed a spray tip with a coaxial three-layer structure, into which the CE sample solution, the sheath liquid, and the nebulizing gas were introduced. The spray tip was put between the DBDI outlet and the MS inlet, thus the CE sample solution could be blended with the sheath liquid, then nebulized. The nebulized sample could be ionized by DBDI, and finally analyzed by MS. The key parameters of the interface were optimized. Then, proof-of-concept experiments separating and detecting the mixture of metronidazole and acetaminophen solutions were conducted. The results showed high separation efficiency, low time consumption, high reproducibility, and convenience in operation. In addition, the interface exhibited a high tolerance of non-volatile salts and surfactants, which would be widely used in CE analyses. All of these results demonstrated that the newly developed CE-DBDI-MS interface could be successfully used in CE-MS studies, and could be further utilized in multiple areas involving efficient separation and detection.

When DNA is mixed with the cationic polyelectrolyte poly(diallyldimethyl ammonium chloride) (PDDA), the DNA/PDDA complex is formed instantaneously at room temperature. This complex is much more efficient in enhancing the fluorescence of Hoechst 33258 (H 33258) than DNA alone. Based on the interaction of H 33258 with the DNA/PDDA complex, a new fluorescence assay for DNA is described. At pH 7.3 in Tris-HCl buffered solution, the DNA/PDDA complex causes a sharp enhancement in fluorescence intensity of H 33258. Simultanously, the emission maximum wavelength of H 33258 blueshifts from 490 nm to 450 nm, while the excitation redshifts from 345 to 350 nm. The calibration graphs for calf thymus DNA (ctDNA) and herring sperm DNA (hsDNA) are both linear up to 5.0 µg mL−1 when the concentration of H 33258 and PDDA are fixed at 1.5×10−6 and 1.6×10−5 mol L−1, respectively. The method is specific for native DNA. The 3σ detection limits for ctDNA and hsDNA are 1.8 and 5.6 ng mL−1, respectively, i.e. much lower than in the presence of H 33258 alone. Four synthetic samples were determined satisfactorily. This method can also be developed to investigate the formation and the nature of the complexes between DNA and polycations, which have recently been widely applied in some fields such as genetic engineering and gene therapy.

Emerging evidence has demonstrated that gut microbiome plays essential roles in the pathogenesis of human diseases in distal organs. Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease characterized by the progressive loss of motor neurons. Treatment with the only drug approved by the US Food and Drug Administration for use in ALS, riluzole, extends a patient׳s life span by only a few months. Thus, there is an urgent need to develop novel interventions that for alleviate disease progression and improve quality of life in patients with ALS. Here we present evidence that intestinal dysfunction and dysbiosis may actively contribute to ALS pathophysiology. We used G93A transgenic mice as a model of human ALS. The G93A mice show abnormal intestinal microbiome and damaged tight junctions before ALS disease onset. The mice were given 2% butyrate, a natural bacterial product, in the drinking water. In mice fed with butyrate, intestinal microbial homeostasis was restored, gut integrity was improved, and life span was prolonged compared with those in control mice. At the cellular level, abnormal Paneth cells—specialized intestinal epithelial cells that regulate the host–bacterial interactions—were significantly decreased in the ALS mice treated with butyrate. In both ALS mice and intestinal epithelial cells cultured from humans, butyrate treatment was associated with decreased aggregation of the G93A superoxide dismutase 1 mutated protein. The findings from this study highlight the complex role of the gut microbiome and intestinal epithelium in the progression of ALS and present butyrate as a potential therapeutic reagent for restoring ALS-related dysbiosis.

There remains a critical need for the precise control of CRISPR (clustered regularly interspaced short palindromic repeats)-based technologies. Here, we engineer a set of inducible CRISPR-based tools controllable by focused ultrasound (FUS), which can penetrate deep and induce localized hyperthermia for transgene activation. We demonstrate the capabilities of FUS-inducible CRISPR, CRISPR activation (CRISPRa), and CRISPR epigenetic editor (CRISPRee) in modulating the genome and epigenome. We show that FUS-CRISPR-mediated telomere disruption primes solid tumours for chimeric antigen receptor (CAR)-T cell therapy. We further deliver FUS-CRISPR in vivo using adeno-associated viruses (AAVs), followed by FUS-induced telomere disruption and the expression of a clinically validated antigen in a subpopulation of tumour cells, functioning as “training centers” to activate synthetic Notch (synNotch) CAR-T cells to produce CARs against a universal tumour antigen to exterminate neighboring tumour cells. The FUS-CRISPR(a/ee) toolbox hence allows the noninvasive and spatiotemporal control of genomic/epigenomic reprogramming for cancer treatment. There remains a critical need for precise control of CRISPR-based technologies. Here, the authors develop a focused ultrasound (FUS)-controllable CRISPR toolbox, allowing the noninvasive and spatiotemporal control of genomic/epigenomic reprogramming for cancer treatment combined with CAR-T therapy.