An activatable fluorescence probe for rapid detection and in situ imaging of ß-galactosidase activity in cabbage roots under heavy metal stress.

ß-Galactosidase (ß-gal), an enzyme related to cell wall degradation, plays an important role in regulating cell wall metabolism and reconstruction. However, activatable fluorescence probes for the detection and imaging of ß-gal fluctuations in plants have been less exploited. Herein, we report an activatable fluorescent probe based on intramolecular charge transfer (ICT), benzothiazole coumarin-bearing ß-galactoside (BC-ßgal), to achieve distinct in situ imaging of ß-gal in plant cells. It exhibits high sensitivity and selectivity to ß-gal with a fast response (8 min). BC-ßgal can be used to efficiently detect the alternations of intracellular ß-gal levels in cabbage root cells with considerable imaging integrity and imaging contrast. Significantly, BC-ßgal can assess ß-gal activity in cabbage roots under heavy metal stress (Cd2+, Cu2+, and Pb2+), revealing that ß-gal activity is negatively correlated with the severity of heavy metal stress. Our work thus facilitates the study of ß-gal biological mechanisms.

Bacterial membrane vesicles: orchestrators of interkingdom interactions in microbial communities for environmental adaptation and pathogenic dynamics.

Bacterial membrane vesicles (MVs) have attracted increasing attention due to their significant roles in bacterial physiology and pathogenic processes. In this review, we provide an overview of the importance and current research status of MVs in regulating bacterial physiology and pathogenic processes, as well as their crucial roles in environmental adaptation and pathogenic infections. We describe the formation mechanism, composition, structure, and functions of MVs, and discuss the various roles of MVs in bacterial environmental adaptation and pathogenic infections. Additionally, we analyze the limitations and challenges of MV-related research and prospect the potential applications of MVs in environmental adaptation, pathogenic mechanisms, and novel therapeutic strategies. This review emphasizes the significance of understanding and studying MVs for the development of new insights into bacterial environmental adaptation and pathogenic processes. Overall, this review contributes to our understanding of the intricate interplay between bacteria and their environment and provides valuable insights for the development of novel therapeutic strategies targeting bacterial pathogenicity.

Surface-mediated fluorescent sensor array for identification of gut microbiota and monitoring of colorectal cancer.

The development of efficient, accurate, and high-throughput technology for gut microbiota sensing holds great promise in the maintenance of health and the treatment of diseases. Herein, we developed a rapid fluorescent sensor array based on surface-engineered silver nanoparticles (AgNPs) and vancomycin-modified gold nanoclusters (AuNCs@Van) for gut microbiota sensing. By controlling the surface of AgNPs, the recognition ability of the sensor can be effectively improved. The sensor array was used to successfully discriminate six gut-derived bacteria, including probiotics, neutral, and pathogenic bacteria and even their mixtures. Significantly, the sensing system has also been successfully applied to classify healthy individuals and colorectal cancer (CRC) patients rapidly and accurately within 30 min, demonstrating its clinically relevant specificity.

Fluorescent Probe for Investigating the Mitochondrial Viscosity and Hydrogen Peroxide Changes in Cerebral Ischemia/Reperfusion Injury.

Cerebral ischemia-reperfusion injury (CIRI), a cause of cerebral dysfunction during cerebral infarction treatment, is closely associated with mitochondrial viscosity and hydrogen peroxide (H2O2). However, the accurate measurement of mitochondrial viscosity and H2O2 levels in CIRI is challenging because of the lack of sufficient selectivity and blood-brain barrier (BBB) penetration of existing monitoring tools related to CIRI, hampering the exploration of the role of mitochondrial viscosity and H2O2 in CIRI. To address this issue, we designed an activatable fluorescent probe, mitochondria-targeting styryl-quinolin-ium (Mito-IQS), with excellent properties including high selectivity, mitochondrial targeting, and BBB penetration, for the visualization of mitochondrial viscosity and H2O2 in the brain. Based on the real-time monitoring capabilities of the probe, bursts of mitochondrial viscosity and H2O2 levels were visualized during CIRI. This probe can be used to monitor the therapeutic effects of butylphthalein treatment. More importantly, in vivo experiments further confirmed that CIRI was closely associated with the mitochondrial viscosity and H2O2 levels. This discovery provides new insights and tools for the study of CIRI and is expected to accelerate the process of CIRI diagnosis, treatment, and drug design.

Boronic Acid-Decorated Carbon Dot-Based Semiselective Multichannel Sensor Array for Cytokine Discrimination and Oral Cancer Diagnosis.

Cytokines are essential components of the immune system and are recognized as significant biomarkers. However, detection of a single cytokine is not precise and reliable enough to satisfy the requirements for diagnosis. Herein, we developed a pattern recognition-based method for the multiplexed sensing of cytokines, which involves three-color-emitting boronic acid-decorated carbon dots (BCDs) and arginine-modified titanium carbide (Ti3C2 MXenes) as the sensor array. Initially, the fluorescence signals of the three BCDs were quenched by Ti3C2 MXenes. In the presence of cytokines, the fluorescence intensity of the BCDs was restored or further quenched by different cytokines. The fluorescence response occurs in two steps: first, boronic acid interacts with cis-diol functional groups of cytokines, and second, arginine headgroup selectively interacts with glycans. By exploiting the different competing binding of the BCDs and the cytokines toward Ti3C2 MXenes, seven cytokines and their mixtures can be effectively discriminated at a concentration of 20 ng mL-1. Furthermore, our sensor array demonstrated an excellent performance in classifying human oral cancer saliva samples from healthy individuals with clinically relevant specificity. The noninvasive method offers a rapid approach to cytokine analysis, benefiting early and timely clinical diagnosis and treatment.

NPAS2 dampens chemo-sensitivity of lung adenocarcinoma cells by enhancing DNA damage repair.

Chemotherapeutic agents, including cisplatin, have remained a cornerstone of lung adenocarcinoma (LUAD) treatment and continue to play an essential role in clinical practice, despite remarkable progress in therapeutic strategies. Hence, a thorough comprehension of the molecular mechanisms underlying chemotherapeutic agent resistance is paramount. Our investigation centered on the potential involvement of the NPAS2 gene in LUAD, which is highly expressed in tumors and its high expression has been associated with unfavorable overall survival rates in patients. Intriguingly, we observed that the depletion of NPAS2 in LUAD cells resulted in increased susceptibility to cisplatin treatment. Furthermore, mRNA sequencing analysis revealed that NPAS2 deficiency downregulated genes crucial to DNA repair. Additionally, NPAS2 depletion significantly impairs γH2AX accumulation, a pivotal component of the DNA damage response. Further investigation demonstrates that NPAS2 plays a crucial role in DNA double-strand breakage repair via homology-directed repair (HDR). Our inquiry into the molecular mechanisms underlying NPAS2 regulation of DDR revealed that it may enhance the stability of H2AX mRNA by binding to its mRNA, thereby upregulating the DNA damage repair pathway. In-vivo experiments further confirmed the crucial role of NPAS2 in modulating the effect of cisplatin in LUAD. Taken together, our findings suggest that NPAS2 binds to and enhances the stability of H2AX mRNA, thereby decreasing the sensitivity of tumor cells to chemotherapy by augmenting DNA damage repair.

Antimicrobial Agent Functional Gold Nanocluster-Mediated Multichannel Sensor Array for Bacteria Sensing.

Urinary tract infections (UTIs) have greatly affected human health in recent years. Accurate and rapid diagnosis of UTIs can enable a more effective treatment. Herein, we developed a multichannel sensor array for efficient identification of bacteria based on three antimicrobial agents (vancomycin, lysozyme, and bacitracin) functional gold nanoclusters (AuNCs). In this sensor, the fluorescence intensity of the three AuNCs was quenched to varying degrees by the bacterial species, providing a unique fingerprint for different bacteria. With this sensing platform, seven pathogenic bacteria, different concentrations of the same bacteria, and even bacterial mixtures were successfully differentiated. Furthermore, UTIs can be accurately identified with our sensors in ∼30 min with 100% classification accuracy. The proposed sensing systems offer a rapid, high-throughput, and reliable sensing platform for the diagnosis of UTIs.

A sensitive electrochemical sensor for glutathione based on specific recognition induced collapse of silver-contained metal organic frameworks.

An electrochemical sensor capable of detecting glutathione (GSH) with high sensitivity and selectivity was developed based on the unique novel electroactive silver-based metal organic framework (Ag-MOF). The Ag-MOF obtained by silver nitrate and 1,3,5-benzoic acid (H3BTC) was thoroughly characterized and was modified onto the electrode via facile drop-casting method. The electrochemical response of GSH on the Ag-MOF modified electrode showed a significant reduction in the current signal because the Ag-GSH complex had stronger specific affinity than Ag-H3BTC and resulted in the collapse of the Ag-MOF. This sensor demonstrated an extensive linear dynamic range of 0.1 nM-1 µM, along with the low detection limit of 0.018 nM. Additionally, it exhibited good reproducibility, stability, and resistance to interfering compounds. The Ag-MOF modified electrode demonstrated superior performance attributed to its rapid electron transfer rate, outstanding electrochemical redox activity, and specific recognition/competitive reaction. These factors improved both sensitivity and selectivity. The high anti-interference ability allowed for the selective detection of GSH in intricate surroundings. In the real sample testing, the RSD was lower than 3.1% and the recovery was between 98.1 and 103%. This research highlights the potential of Ag-MOFs in developing electrochemical sensors and their promising applications in determining GSH for food screening and early disease diagnosis.

DNA Sensor-Based Strategy to Visualize the TRPM7 mRNA-Mg2+ Signaling Pathway in Cancer Cells.

Technological advances and methodological innovations in cell signaling pathway analysis will facilitate progress in understanding biological processes, intervening in diseases, and screening drugs. In this work, an elaborate strategy for visualizing and monitoring the transient receptor potential melastatin 7 (TRPM7)-Mg2+ signaling pathway in living cells was constructed through the logical analysis of upstream mRNA and downstream molecules by two individual DNA sensors. The DNA sensors are constructed by modifying the dye-labeled DNA sequences on the surface of gold nanoparticles. By hybridizing with upstream mRNA, Cy5-modified DNA sensor 1 can detect and silence it simultaneously, outputting a red fluorescence signal. When the upstream mRNA is silenced, the concentration of downstream molecules of Mg2+ will be affected and down-regulated. The FAM-modified DNA sensor 2 detects this change and emits a green fluorescence as a signal. Therefore, the dynamic information on TRPM7 mRNA and the Mg2+-mediated signaling pathway can be successfully obtained by fluorescence imaging methods. Furthermore, the TRPM7 mRNA-Mg2+ signaling pathway also affects cell activity and migratory function through cell scratching and other experiments. More importantly, the proposed sensor also shows potential for screening signaling pathway inhibitors. Our work provides a simple and general strategy for the visualization of signaling pathways, which helps to understand the changes in the physiological activities of cancer cells and the causes of carcinogenesis and is crucial for cancer diagnosis and prognosis.

Consuming intracellular glucose and regulating the levels of O2/H2O2 via the closed cascade catalysis system of Cu-CeO2 nanozyme and glucose oxidase.

Imbalances in the intracellular environment caused by high levels of glucose, H2O2, and hypoxia can greatly impact cancer development and treatment. However, there is limited research on regulating the levels of these species simultaneously in tumor cells. Here, a pH-responsive nanozyme-enzyme hybrid system was developed to regulate intracellular glucose, H2O2 and O2. The system, named DMSN@Cu-CeO2@GOx, consists of Cu-CeO2 nanoparticles and glucose oxidase (GOx) immobilized in dendritic mesoporous silica (DMSN) spheres. GOx efficiently consumes glucose in tumor cells, causing a drop in pH and producing a significant amount of H2O2. Cu-CeO2 then catalyzes the conversion of H2O2 to O2 due to its high catalase-like (CAT) activity in weakly acidic conditions. The process was monitored by fluorescence probes, and the mechanism was investigated through fluorescence spectroscopy and confocal laser scanning microscopy. The cascade catalytic system with excellent biocompatibility continuously consumes glucose and elevates the level of O2 in cells. This hybrid nanomaterial offers a means to regulate the glucose/H2O2/O2 levels in cells and may provide insights into starvation therapy by modulating reactive species within cells.

A large Stokes shift NIR fluorescent probe for visual monitoring of mitochondrial peroxynitrite during inflammation and ferroptosis and in an Alzheimer's disease model.

The excessive formation of peroxynitrite (ONOO-) in mitochondria has been implicated in various pathophysiological processes and diseases. However, owing to short emission wavelengths and small Stokes shifts, previously reported fluorescent probes pose significant challenges for mitochondrial ONOO- imaging in biological systems. In this study, a near-infrared (NIR) fluorescent probe, denoted as DCO-POT, is designed for the visual monitoring of mitochondrial ONOO-, displaying a remarkable Stokes shift of 170 nm. The NIR fluorophore of DCO-CHO is released by DCO-POT upon the addition of ONOO-, resulting in off-on NIR fluorescence at 670 nm. This phenomenon facilitates the high-resolution confocal laser scanning imaging of ONOO- generated in biological systems. The practical applications of DCO-POT as an efficient fluorescence imaging tool are verified in this study. DCO-POT enables the fluorometric visualization of ONOO- in organelles, cells, and organisms. In particular, ONOO- generation is analyzed during cellular and organism-level (zebrafish) inflammation during ferroptosis and in an Alzheimer's disease mouse model. The excellent visual monitoring performance of DCO-POT in vivo makes it a promising tool for exploring the pathophysiological effects of ONOO-.

Self-assembled super-small AIEgen nanoprobe for highly sensitive and selective detection of protamine and trypsin.

Amphiphilic aggregation-induced emission (AIE) molecules show superior potential for fabricating novel ultrasmall nanoprobes. Here, an anionic dipyridyl tetraphenylethene (TPE) derivative is rationally designed and a super-small self-assembled AIEgen nanoprobe (TPE-2Py-SO3NaNPs, ca. 2.48 nm) is thus conveniently constructed for the supersensitive detection of protamine and trypsin. In HEPES/DMSO solution (8 : 2, v/v, pH = 7.4), negatively charged TPE-2Py-SO3NaNPs exhibited an AIE effect in the presence of positively charged protamine, presenting a fluorescence enhancement at 498 nm together with a large Stokes shift of 150 nm and a low detection limit of 8.0 ng mL-1. In addition, the in situ formed TPE-2Py-SO3Na/protamine nanocomposite can be dissociated by trypsin due to the highly selective degradation of protamine via enzymatic hydrolysis, achieving a detection limit for trypsin as low as 5.0 ng mL-1.

Molecular dynamic simulation reveals the molecular interactions of epidermal growth factor receptor with musk xylene are involved in the carcinogenicity.

Musk xylene (MX), a kind of personal care product, has become a new type of environmental contaminant in recent years. Long-term exposure to MX is associated with a variety of cancers, but the mechanism is still unclear. Meanwhile, our previous research showed that MX exposure could lead to malignant transformation of human liver cells L02 and up-regulation of multi genes which are involved in the MAPK signaling pathway, such as the epidermal growth factor receptor (EGFR). These findings indicated that the MAPK signaling pathway might be involved in the malignant transformation caused by MX, but the mechanism is also unclear. In this study, the underlying interaction mechanisms between EGFR and MX were investigated using molecular dynamics (MD) simulation. Results revealed that MX bound to the ECD of EGFR in four binding sites, which was mainly driven by van der Waals and nonpolar interactions, and the affinity of MX toward ECD was sIII > sI > sII > sIV. Further analysis through MD simulation found that s III, the site with the strongest binding, was coincidentally located at the binding area of EGF, which is the natural ligand of EGFR. Therefore, we speculated that MX may activate the MAPK signaling pathway by binding to EGFR in a similar way to EGF, and finally lead to tumorigenesis. In addition, the MM/PBSA method could also be utilized to calculate the hot residues in each binding site. The prediction of hot residues would provide some theoretical guidance for further study of the carcinogenesis mechanisms of MX both in MD simulation and experimental research.

Ag2S QDs integration with MnO2 nanosheets for the sensitive detection of Cr (VI) via the redox reaction induced photoelectrochemical variation.

The heavy metal Cr (VI) will remain, accumulate, and migrate after entering the environment or ecosystem, causing serious harm to the environment. Here, a photoelectrochemical sensor was developed for Cr (VI), utilizing the Ag2S quantum dots (QDs) and MnO2 nanosheets as photoactive components. By introducing Ag2S QDs with a narrow gap, a staggered energy level match is created which effectively prevents the carrier recombination in MnO2 nanosheets, resulting in an enhanced photocurrent response. In the presence of the electron donor, l-ascorbic acid (AA), the photocurrent of the Ag2S QDs and MnO2 nanosheets modified photoelectrode is further enhanced. As AA has the ability to convert Cr (VI) to Cr (Ⅲ), the photocurrent may decline due to the decrease in the electron donors when Cr (VI) is added. This phenomenon can be utilized for the sensitive detection of Cr (VI) over a wider linear range (100 pM-30 µM) with a lower detection limit of 6.46 pM (S/N = 3). This work using the strategy that the targets induced the variations of the electron donor shows the advantages of good sensitivity and nice selectivity. The sensor holds many advantages such as simple fabrication process, economical material expense, and consistent photocurrent signals. It also holds significant potential for environmental monitoring and serves as a practical photoelectric sensing approach for detecting Cr (VI).

General Strategy for Specific Fluorescence Imaging of Homocysteine in Living Cells and In Vivo.

The aberrantly changed level of homocysteine (Hcy) triggers a variety of pathological symptoms and subsequently Hcy-related diseases. Direct and selective visualization of Hcy in biological systems is pivotal to understanding the pathological functions of Hcy at the molecular level. Herein, a general strategy was developed for the specific fluorescence imaging of Hcy through the combination of dual-binding sites and the introduction of a nitro group at the 6-position of the 7-diethylaminocoumarin fluorophore. Also, a series of novel fluorescent probes were exploited for monitoring Hcy with excellent selectivity, high sensitivity, and far-red/near-infrared fluorescence emission. Furthermore, fluorescence imaging of endogenous Hcy dynamics in living cells and in vivo was achieved, providing direct and solid evidence for the increasement of endogenous Hcy in type 2 diabetes mellitus and Alzheimer's disease. This research will greatly advance the development and understanding of the molecular nexus between the Hcy metabolism cascade and the root causes of diseases related to Hcy.

Virulence regulation of Zn2+ uptake system znuABC on mesophilic Aeromonas salmonicida SRW-OG1.

Psychrophilic Aeromonas salmonicida could not grow above 25°C and therefore thought unable to infect mammals and humans. In our previous study, a mesophilic A. salmonicida SRW-OG1 was isolated from Epinephelus coioides with furunculosis. Through the analysis of preliminary RNA-seq, it was found that the Zn2+ uptake related genes znuA, znuB and znuC might be involved in the virulence regulation of A. salmonicida SRW-OG1. Therefore, the purpose of this study was to explore the effect of znuABC silencing on the virulence regulation of A. salmonicida SRW-OG1. The results showed that the growth of the znuA-RNAi, znuB-RNAi, and znuC-RNAi strains was severely restricted under the Fe2+ starvation, but surprisingly there was no significant difference under the Zn2+ restriction. In the absence of Zn2+ and Fe2+, the expression level of znuABC was significantly increased. The motility, biofilm formation, adhesion and hemolysis of the znuA-RNAi, znuB-RNAi, and znuC-RNAi strains were significantly reduced. We also detected the expression of znuABC under different growth periods, temperatures, pH, as well as Cu2+ and Pb2+ stresses. The results showed that znuABC was significantly up-regulated in the logarithmic phase and the decline phase of A. salmonicida. Interestingly, the trend of expression levels of the znuABC at 18, 28, and 37°C was reversed to another Zn2+ uptake related gene zupT. Taken together, these indicated that the znuABC was necessary for A. salmonicida SRW-OG1 pathogenicity and environmental adaptability, and was cross regulated by iron starvation, but it was not irreplaceable for A. salmonicida SRW-OG1 Zn2+ uptake in the host.

Glucose determination in human serum by applying inner filter effect quenching mechanism of upconversion nanoparticles.

Accurate blood glucose determination is essential to the clinical diagnosis and management of diabetes. This work establishes an inner filter effect (IFE) strategy between upconversion nanoparticles (UCNPs) and quinone-imine complex for glucose monitoring in human serum simply and efficiently. In this system, the enzyme glucose oxidase (GOx) catalyzes the reaction of glucose into hydrogen peroxide (H2O2) and gluconic acid when compulsion by oxygen. In the presence of horseradish peroxidase (HRP), the produced H2O2 can catalytically oxidize phenol and 4-amino antipyrine (4-AAP) to generate quinone-imine products. The purple-colored quinone-imine complex effectively absorbed the fluorescence of NaYF4:Yb3+, Er3+ UCNPs, leading to the strong fluorescence quenching of UCNPs through IFE. Thus, a new approach was established for glucose monitoring by determining the fluorescence intensity. Under the optimal condition, this approach shows better linearity to glucose from 2-240 µmol/L with a low detection limit at 1.0 µmol/L. Owing to the excellent fluorescence property and background-free interference of the UCNPs, the biosensor was applied for glucose measurements in human serum and got a satisfactory result. Furthermore, this sensitive and selective biosensor revealed great potential for the quantitative analysis of blood glucose or different kinds of H2O2-involved biomolecules for the application of clinical diagnosis.

Recognition Engineering-Mediated Multichannel Sensor Array for Gut Microbiota Sensing.

The composition and activity of the gut microbiota are crucial for health management and disease treatment. Herein, we develop a rapid and robust multichannel sensor array via a recognition engineering strategy using antimicrobial agent (vancomycin, bacitracin, and lysozyme) functional gold nanoclusters and gluconamide-modified Ti3C2 MXenes, which provide superior fingerprint patterns to distinguish gut-derived bacteria. The discrimination ability of the sensor array was highly improved via the synergistic recognition between the bacteria and the various antimicrobial agents. Five gut-derived bacteria, including probiotics, neutral, and pathogenic bacteria were clearly differentiated and discriminated from the bacteria mixtures. Furthermore, the sensing system was successfully applied for the accurate classification of human colorectal cancer samples from healthy individuals rapidly (30 min) with clinically relevant specificity. The rapidity, simplicity, and economic cost of this strategy offers a robust platform for gut microbiota analysis.

A turn-on NIR fluorescent probe for risk-assessing oxidative stress in cabbage roots under abiotic stress.

Oxidative stress is closely related to the crop health status under stress conditions. H2O2 is an important signaling molecule in plants under stress. Therefore, monitoring H2O2 fluctuations is of great significance when risk-assessing oxidative stress. However, few fluorescent probes have been reported for the in situ tracking of H2O2 fluctuations in crops. Herein, we designed a "turn-on" NIR fluorescent probe (DRP-B) to detect and in situ-image H2O2 in living cells and crops. DRP-B exhibited good detection performance for H2O2 and could image endogenous H2O2 in living cells. More importantly, it could semi-quantitatively visualize H2O2 in cabbage roots under abiotic stress. Visualization of H2O2 in cabbage roots revealed H2O2 upregulation in response to adverse environments (metals, flood, and drought). This study provides a new method for risk-assessing oxidative stress in plants under abiotic stress and is expected to provide guidance for the development of new antioxidant defense strategies to enhance plant resistance and crop productivity.

Mitochondria-targeting fluorescent sensor with high photostability and permeability for visualizing viscosity in mitochondrial malfunction, inflammation, and AD models.

Viscosity changes in mitochondria are closely associated with numerous cellular processes and diseases. Currently available fluorescence probes used in mitochondrial viscosity imaging are not very photostable or sufficiently permeable. Herein, a highly photostable and permeable mitochondria-targeting red fluorescent probe (Mito-DDP) was designed and synthesized for viscosity sensing. Viscosity was imaged in living cells using a confocal laser scanning microscope, and the results suggested that Mito-DDP penetrated the membrane and stained the living cells. Importantly, practical applications of Mito-DDP were demonstrated: viscosity visualization was realized for mitochondrial malfunction, cellular and zebrafish inflammation, and Drosophila Alzheimer's disease models, i.e., for subcellular organelles, cells, and organisms. The excellent analytical and bioimaging performance of Mito-DDP in vivo makes it an effective tool for exploring the physiological and pathological effects of viscosity.