AAV gene replacement therapy for treating MPS IIIC: Facilitating bystander effects via EV-mRNA cargo.

MPS IIIC is a lysosomal storage disease caused by mutations in heparan-α-glucosaminide N-acetyltransferase (HGSNAT), for which no treatment is available. Because HGSNAT is a trans-lysosomal-membrane protein, gene therapy for MPS IIIC needs to transduce as many cells as possible for maximal benefits. All cells continuously release extracellular vesicles (EVs) and communicate by exchanging biomolecules via EV trafficking. To address the unmet need, we developed a rAAV-hHGSNATEV vector with an EV-mRNA-packaging signal in the 3'UTR to facilitate bystander effects, and tested it in an in vitro MPS IIIC model. In human MPS IIIC cells, rAAV-hHGSNATEV enhanced HGSNAT mRNA and protein expression, EV-hHGSNAT-mRNA packaging, and cleared GAG storage. Importantly, incubation with EVs led to hHGSNAT protein expression and GAG contents clearance in recipient MPS IIIC cells. Further, rAAV-hHGSNATEV transduction led to the reduction of pathological EVs in MPS IIIC cells to normal levels, suggesting broader therapeutic benefits. These data demonstrate that incorporating the EV-mRNA-packaging signal into a rAAV-hHGSNAT vector enhances EV packaging of hHGSNAT-mRNA, which can be transported to non-transduced cells and translated into functional rHGSNAT protein, facilitating cross-correction of disease pathology. This study supports the therapeutic potential of rAAVEV for MPS IIIC, and broad diseases, without having to transduce every cell.

Highly selective and additive-free Pd(OAc)2/CPP catalyzed hydroaminocarbonylation of alkynes.

Herein, the synthesis of branched α,ß-unsaturated amides by a hydroaminocarbonylation reaction of alkynes with various amine substrates such as aromatic amines, aliphatic amines, solid amine sources like NH4HCO3, and even strongly basic piperidines is reported, using a Pd(OAc)2/hybrid N-heterocyclic carbene-phosphine-phosphine (CPP) catalytic system. The reactions feature no additives, wide substrate scope, high selectivity (b/l > 99 : 1) and excellent yields. Mechanistic studies have disclosed that the reaction takes place via a palladium hydride pathway. CPP adopts a hybrid bidentate ligand conformation with a carbene-phosphine coordination mode, wherein one phosphorus atom remains externally accessible, potentially serving as a stabilizing auxiliary during catalytic cycles.

m6A RNA methylation drives kidney fibrosis by upregulating ß-catenin signaling.

N6-methyladenosine (m6A) methylation plays a crucial role in various biological processes and the pathogenesis of human diseases. However, its role and mechanism in kidney fibrosis remain elusive. In this study, we show that the overall level of m6A methylated RNA was upregulated and the m6A methyltransferase METTL3 was induced in kidney tubular epithelial cells in mouse models and human kidney biopsies of chronic kidney disease (CKD). Proximal tubule-specific knockout of METTL3 in mice protected kidneys against developing fibrotic lesions after injury. Conversely, overexpression of METTL3 aggravated kidney fibrosis in vivo. Through bioinformatics analysis and experimental validation, we identified ß-catenin mRNA as a major target of METTL3-mediated m6A modification, which could be recognized by a specific m6A reader, the insulin-like growth factor 2 mRNA binding protein 3 (IGF2BP3). METTL3 stabilized ß-catenin mRNA, increased ß-catenin protein and induced its downstream profibrotic genes, whereas either knockdown of IGF2BP3 or inhibiting ß-catenin signaling abolished its effects. Collectively, these results indicate that METTL3 promotes kidney fibrosis by stimulating the m6A modification of ß-catenin mRNA, leading to its stabilization and its downstream profibrotic genes expression. Our findings suggest that targeting METTL3/IGF2BP3/ß-catenin pathway may be a novel strategy for the treatment of fibrotic CKD.

AntDAS-GCMS: A New Comprehensive Data Analysis Platform for GC-MS-Based Untargeted Metabolomics with the Advantage of Addressing the Time Shift Problem.

Over the years, a number of state-of-the-art data analysis tools have been developed to provide a comprehensive analysis of data collected from gas chromatography-mass spectrometry (GC-MS). Unfortunately, the time shift problem remains unsolved in these tools. Here, we developed a novel comprehensive data analysis strategy for GC-MS-based untargeted metabolomics (AntDAS-GCMS) to perform total ion chromatogram peak detection, peak resolution, time shift correction, component registration, statistical analysis, and compound identification. Time shift correction was specifically optimized in this work. The information on mass spectra and elution profiles of compounds was used to search for inherent landmarks within analyzed samples to resolve the time shift problem across samples efficiently and accurately. The performance of our AntDAS-GCMS was comprehensively investigated by using four complex GC-MS data sets with various types of time shift problems. Meanwhile, AntDAS-GCMS was compared with advanced GC-MS data analysis tools and classic time shift correction methods. Results indicated that AntDAS-GCMS could achieve the best performance compared to the other methods.

Efficient and selective external activator-free cobalt catalyst for hydroboration of terminal alkynes enabled by BiPyPhos.

Herein, a robust catalyst system, composed of a bipyridine-based diphosphine ligand (BiPyPhos) and a cobalt precursor Co(acac)2, is successfully developed and applied in the hydroboration of terminal alkynes, exclusively affording various versatile ß-E-vinylboronates in high yields at room temperature.

Factors influencing pathological changes in the liver tissue in hepatitis B virus carriers with low-level viremia.

OBJECTIVES: To investigate the optimal timing for initiating antiviral therapy in hepatitis B virus (HBV) carriers with low-level viremia (LLV). METHODS: We retrospectively enrolled 126 HBV carriers with LLV who underwent liver biopsy. Patients' clinical data, routine blood test results, portal vein diameter, splenic vein diameter and thickness, and measurements (LSM) within 1 week before liver biopsy were obtained. Single-factor and multifactor statistical methods were used to analyze factors that affected inflammation and fibrosis in pathological liver tissues. The receiver operating characteristic curve was used to analyze liver stiffness and HBV DNA levels to determine liver tissue inflammation and fibrosis. R -Studio software was used to draw nomograms, calibration plots, and model decision curves. RESULTS: Infection duration and HBV DNA levels affected liver tissue inflammation. Albumin(ALB), aspartate aminotransferase (AST), HBV DNA, liver stiffness, age, and splenic thickness affected liver fibrosis. The best cutoff value of the LSM for diagnosing liver inflammation and fibrosis was 7.45 (specificity, 92%). The best cutoff value of HBV DNA for diagnosing liver inflammation and fibrosis was 39.5 (specificity, 96%). HBV DNA,and splenic thickness affected the treatment decision in naive chronic hepatitis Bpatients with LLV CONCLUSIONS: HBV carriers with LLV have high incidences of liver tissue inflammation and fibrosis. The infection duration and HBV DNA levels affected liver inflammation whereas the ALB, AST levels, HBV DNA, LSM, age, and splenic thickness affected liver fibrosis. Eligible expansion of antiviral treatment indications is necessary, however, a universal treatment approach may be inefficient. HBV DNA can be a reference for initiating antiviral therapy.

Organoboron-Promoted Electrochemical Chlorosulfonylation of Alkenes with Sulfonyl Chlorides.

Although extraordinary advances have been achieved by the transition-metal catalysis system, there is an urgent need to explore and develop alternative methodologies that are more environmentally friendly. Herein, we report an electrochemical chlorosulfonylation of alkenes using a wide range of sulfonyl chlorides with an inexpensive, degradable, and commercially available organoboron as a promoter. Furthermore, this protocol employs convergent paired electrolysis, reducing the need for sacrificial anodes and minimizing the extent of hydrogen evolution.

Shift in microorganism and functional gene abundance during completely autotrophic nitrogen removal over nitrite (CANON) process.

Nitrification-denitrification process has failed to meet wastewater treatment standards. The completely autotrophic nitrite removal (CANON) process has a huge advantage in the field of low carbon/nitrogen wastewater nitrogen removal. However, slow start-up and system instability limit its applications. In this study, the time of the start-up CANON process was reduced by using bio-rope as loading materials. The establishing of graded dissolved oxygen improved the stability of the CANON process and enhanced the stratification effect between functional microorganisms. Microbial community structure and the abundance of nitrogen removal functional genes are also analyzed. The results showed that the CANON process was initiated within 75 days in the complete absence of anaerobic ammonium oxidizing bacteria (AnAOB) inoculation. The ammonium and nitrogen removal efficiencies of CANON process reached to 94.45% and 80.76% respectively. The results also showed that the relative abundance of nitrogen removal bacterial in the biofilm gradually increases with the dissolved oxygen content in the solution decreases. In contrast, the relative abundance of ammonia oxidizing bacteria was positively correlated with the dissolved oxygen content in the solution. The relative abundance of g__Candidatus_Brocadia in biofilm was 15.56%, and while g__Nitrosomonas was just 0.6613%. Metagenomic analysis showed that g__Candidatus_Brocadia also contributes 66.37% to the partial-nitrification functional gene Hao (K10535). This study presented a new idea for the cooperation between partial-nitrification and anammox, which improved the nitrogen removal system stability.

Esters-targeted colorimetric sensor array for the authenticity discrimination of strong-aroma baijiu with different origins.

Baijiu authenticity has been a frequent problem driven by economic interests in recent years, so it is important to discriminate against baijiu with different origins. Herein, we proposed a simple and efficient esters-targeted colorimetric sensor array mediated by hydroxylamine hydrochloride. Esters undergo a nucleophilic addition reaction with hydroxylamine hydrochloride to form hydroxamic acid, which rapidly forms a purplish red ferric hydroxamate under FeCl3·6H2O. Bromophenol blue and rhodamine B enrich the color effects. The array detected 12 esters with a detection limit on the order of 10-5 of most esters and 16 mixed esters with R2 > 0.999 and recoveries close to 100%. Otherwise, for discriminating 34 strong-aroma baijius (SABs), the array has an accuracy of 98% according to the origin, and 95% according to the grades, with a response time of 1 min. This study provides a new strategy for authenticity determination and quality control of baijiu.

Fibroblast expression of transmembrane protein smoothened governs microenvironment characteristics after acute kidney injury.

The smoothened (Smo) receptor facilitates hedgehog signaling between kidney fibroblasts and tubules during acute kidney injury (AKI). Tubule-derived hedgehog is protective in AKI, but the role of fibroblast-selective Smo is unclear. Here, we report that Smo-specific ablation in fibroblasts reduced tubular cell apoptosis and inflammation, enhanced perivascular mesenchymal cell activities, and preserved kidney function after AKI. Global proteomics of these kidneys identified extracellular matrix proteins, and nidogen-1 glycoprotein in particular, as key response markers to AKI. Intriguingly, Smo was bound to nidogen-1 in cells, suggesting that loss of Smo could affect nidogen-1 accessibility. Phosphoproteomics revealed that the 'AKI protector' Wnt signaling pathway was activated in these kidneys. Mechanistically, nidogen-1 interacted with integrin ß1 to induce Wnt in tubules to mitigate AKI. Altogether, our results support that fibroblast-selective Smo dictates AKI fate through cell-matrix interactions, including nidogen-1, and offers a robust resource and path to further dissect AKI pathogenesis.

A colorimetric sensor based on 4-MPBA Au@AgNPs for accurately identification of EnshiYulu tea grade.

Enshi Yulu green tea (ESYL) is the most representative traditional steamed green tea in Enshi, Hubei. Different ESYL grades exhibit distinct flavors, tastes, and prices. In this study, a visual sensor based on 4-MPBA Au@AgNPs was developed for the rapid and accurate identification of ESYL grades. The recognition mechanism involved the binding of 4-MPBA Au@AgNPs with polyphenolic compounds in ESYL to form borate esters and the conversion of Ag+ to Ag0, with the generated Ag0 depositing on the surface of 4-MPBA Au@AgNPs. The results showed that the sensor can amplify the color differences of different grades of ESYL. The visual results were also validated by the partial least squares discriminant analysis model, demonstrating an enhancement in recognition accuracy from 68.2 % to 95.5 % compared to the original extraction solution. The colorimetric sensor developed in this study is expected to provide a new approach for traceability research of other foods.

Myeloid-derived Wnts play an indispensible role in macrophage and fibroblast activation and kidney fibrosis.

Wnt/ß-catenin signaling plays a pivotal role in the pathogenesis of chronic kidney diseases (CKD), which is associated with macrophage activation and polarization. However, the relative contribution of macrophage-derived Wnts in the evolution of CKD is poorly understood. Here we demonstrate a critical role of Wnts secreted by macrophages in regulating renal inflammation and fibrosis after various injuries. In mouse model of kidney fibrosis induced by unilateral ureteral obstruction (UUO), macrophages were activated and polarized to M1 and M2 subtypes, which coincided with the activation of Wnt/ß-catenin signaling. In vitro, multiple Wnts were induced in primary cultured bone marrow-derived macrophages (BMDMs) after polarization. Conversely, Wnt proteins also stimulated the activation and polarization of BMDMs to M1 and M2 subtype. Blockade of Wnt secretion from macrophages in mice with myeloid-specific ablation of Wntless (Wls), a cargo receptor that is obligatory for Wnt trafficking and secretion, blunted macrophage infiltration and activation and inhibited the expression of inflammatory cytokines. Inhibition of Wnt secretion by macrophages also abolished ß-catenin activation in tubular epithelium, repressed myofibroblast activation and reduced kidney fibrosis after either obstructive or ischemic injury. Furthermore, conditioned medium from Wls-deficient BMDMs exhibited less potency to stimulate fibroblast proliferation and activation, compared to the controls. These results underscore an indispensable role of macrophage-derived Wnts in promoting renal inflammation, fibroblasts activation and kidney fibrosis.

An ion synergism fluorescence probe via Cu2+ triggered competition interaction to detect glyphosate.

The widespread use of glyphosate (Gly) poses significant risks to environmental and human health, underscoring the urgent need for its sensitive and rapid detection. In this work, we innovated by developing a novel material, ionic liquids, which formed the ionic probe "[P66614]2[2,3-DHN]-Cu2+ (PDHN-Cu2+)" through coordination with Cu2+. This probe capitalized on the distinctive fluorescence quenching properties of ionic liquids in the presence of Cu2+, driven by synergistic interactions between anions and cations. Glyphosate disrupted the PDHN-Cu2+ coordination structure due to its stronger affinity for Cu2+, triggering a "turn-on" fluorescence response. Impressively, PDHN-Cu2+ enabled the sensitive detection of glyphosate within just one minute, achieving a detection limit as low as 71.4 nM and excellent recovery rates of 97-103% in diverse samples. This groundbreaking approach, utilizing ionic probes, lays a robust foundation for the accurate and real-time monitoring of pesticides, employing a strategy based on synergism and competitive coordination.

Maresin1 improves hippocampal neuroinflammation and cognitive function in septic rats by activating the SLC7A11 / GPX4 ferroptosis signaling pathway.

Sepsis-associated encephalopathy (SAE) is a prevalent complication of sepsis, with hippocampal neuroinflammation playing a crucial role in SAE-induced cognitive impairment. Maresin1 (MaR1), a bioactive docosahexaenoic acid (DHA) metabolite, demonstrates comprehensive anti-inflammatory and neuroprotective attributes. Yet, its protective efficacy against SAE-induced cognitive decline remains unexplored. In this investigation, we implemented a rat SAE model via cecal ligation and puncture (CLP), while lipopolysaccharide (LPS) stimulation of HT22 cells simulated an in vitro SAE model; both models were pre-treated with MaR1. We evaluated rat learning and memory using a water maze, assessed hippocampal neuron damage via Nissl and FJC staining, and observed mitochondrial alterations through TEM. In vivo and in vitro assays gauged levels of Fe2+, MDA, GSH, and SOD. Additionally, Iba1 expression in the hippocampus was examined via immunofluorescence, while SLC7A11 and GPX4 protein expression levels were determined using western blot. Our findings indicated CLP-induced learning and memory impairment in rats, along with heightened ROS, Fe2+, and MDA levels in hippocampal neurons, diminished GSH and SOD levels, and down-regulated ferroptosis-related proteins (GPX4 and SLC7A11). Remarkably, MaR1 treatment attenuated these adverse effects. In LPS-stimulated HT22 cells, MaR1 lowered lipid ROS and bolstered mitochondrial membrane potential. Nonetheless, the ferroptosis inducer Erastin reversed MaR1's protective effects. Transwell experiments further showed MaR1's potential to inhibit microglia activation triggered by ferroptosis in HT22 cells. Consequently, MaR1 may mitigate hippocampal neuroinflammation via activating the SLC7A11/GPX4 ferroptosis signaling pathway, thus ameliorating SAE-related cognitive impairment.

A fluorescence ionic probe utilizing Cu2+ assisted competition for detecting glyphosate abused in green tea.

Food fraud caused by the violation of glyphosate use in tea is frequently exposed, posing a potential health risk to consumers and undermining trust in food safety. In the work, an ionic fluorescent probe "[P66614] [4HQCA]-Cu2+ (PHQCA-Cu2+)" was constructed using Cu2+ and ionic liquids coordination through a competitive coordination strategy to detect glyphosate. This probe exhibited a prominent "turn-on" fluorescence response in glyphosate detection. PHQCA-Cu2+was destroyed by glyphosate with its strong coordination capability, and a new complex re-formed simultaneously between glyphosate and the Cu2+ in it, where Cu2+ served as an "invisible indicator" influencing fluorescence changes. Remarkably, PHQCA-Cu2+formed rapidly within 5 s, demonstrated exceptional sensitivity and selectivity, and satisfactory detection performance on paper strips impregnated withPHQCA-Cu2+.Importantly,PHQCA-Cu2+showed excellent recoveries in various green tea, which offered a viable method for identifying contaminated products from the supply chain quickly to enhance overall food safety surveillance.

A novel organic acids-targeted colorimetric sensor array for the rapid discrimination of origins of Baijiu with three main aroma types.

Given the severe problem of Baijiu authenticity, it is essential to discriminate Baijiu from different origins quickly and effectively. As organic acids (OAs) are the most dominant taste-imparting substances in Baijiu, we proposed a simple, fast, and effective OAs-targeted colorimetric sensor array based on the colorimetric reaction of 4-aminophenol (AP)/4-amino-3-chlorophenol (ACP) under oxidation of Cu(NO3)2 for the rapid discrimination of origins of Baijiu with three main aroma types. Hydrogen ions ionized from OAs induced the protonation of the amino group, which blocked the colorimetric reaction, and the different levels of OAs in Baijiu enabled the array to discriminate different origins of Baijiu. The array was implemented to analyze 10 simple OAs and 16 mixed OAs and further for the discrimination of 42 Baijius with an accuracy of 98%. This method provided an efficient research strategy for a basis for rapid quality analysis of Baijiu.

[Correlation between diaphragmatic-rapid shallow breathing index and lung ultrasound score in elderly patients with mechanical ventilation and its predictive value for weaning results].

OBJECTIVE: To investigate the correlation between diaphragmatic-rapid shallow breathing index (D-RSBI) and lung ultrasound score (LUS) in elderly patients with mechanical ventilation and its predictive value for weaning results. METHODS: A retrospective study was conducted. The clinical data of elderly patients (age > 60 years old) with invasive positive pressure ventilation (IPPV) admitted to the department of intensive care unit (ICU) of the First Affiliated Hospital of Jinzhou Medical University from January 2021 to June 2022 were enrolled. According to the outcome of withdrawal, the patients were divided into successful and failed groups. The differences in gender, age, acute physiology and chronic health evaluation II (APACHE II), D-RSBI and LUS before weaning and extubation were compared between the two groups. Pearson correlation was used to analyze the correlation between D-RSBI and LUS. The predictive value of D-RSBI and LUS on weaning results of elderly patients with IPPV was analyzed by receiver operator characteristic curve (ROC curve). RESULTS: A total of 398 elderly patients with IPPV were enrolled, including 300 successful weaning patients and 98 failed weaning patients. There were no significant differences in gender and age between the failed group and successful group [male: 55.1% (54/98) vs. 59.0% (177/300), age (years old): 67.02±5.03 vs. 66.96±4.99, both P > 0.05]. APACHE II score in the failed group was significantly higher than that in the successful group (17.09±3.30 vs. 16.06±3.81, P < 0.05), and the D-RSBI and LUS score before extubation were significantly higher than those in the successful group [D-RSBI (time×min-1×mm-1): 2.19±0.33 vs. 1.60±0.22, LUS: 17.30±3.04 vs. 11.97±3.20, both P < 0.01]. All patients showed a significant positive correlation between D-RSBI and LUS score (r = 0.406, P = 0.000). ROC curve analysis showed that the area under the curve (AUC) of D-RSBI for predicting weaning outcomes in elderly IPPV patients was 0.920, with a 95% confidence interval (95%CI) of 0.881-0.958 and P = 0.000. When the cut-off value was 1.85 times×min-1×mm-1, the sensitivity was 88.7% and the specificity was 86.7%. The AUC of LUS score for predicting weaning outcome in elderly IPPV patients was 0.875, with a 95%CI of 0.839-0.912 and P = 0.000. When the cut-off value was 14.50, the sensitivity was 75.7% and the specificity was 84.7%. CONCLUSIONS: There is a significant correlation between D-RSBI and LUS score in elderly mechanically ventilated patients, both of them can predict weaning outcome in elderly patients with mechanical ventilation.

Discrimination and prediction of Qingzhuan tea storage year using quantitative chemical profile combined with multivariate analysis: Advantages of MRMHR based targeted quantification metabolomics.

The duration of storage significantly influences the quality and market value of Qingzhuan tea (QZT). Herein, a high-resolution multiple reaction monitoring (MRMHR) quantitative method for markers of QZT storage year was developed. Quantitative data alongside multivariate analysis were employed to discriminate and predict the storage year of QZT. Furthermore, the content of the main biochemical ingredients, catechins and alkaloids, and free amino acids (FAA) were assessed for this purpose. The results show that targeted marker-based models exhibited superior discrimination and prediction performance among four datasets. The R2Xcum, R2Ycum and Q2cum of orthogonal projection to latent structure-discriminant analysis discrimination model were close to 1. The correlation coefficient (R2) and the root mean square error of prediction of the QZT storage year prediction model were 0.9906 and 0.63, respectively. This study provides valuable insights into tea storage quality and highlights the potential application of targeted markers in food quality evaluation.

Three-dimensional fluorescence combined with alternating trilinear decomposition and random forest algorithm for the rapid prediction of species, geographical origin and main components of Glycyrrhizae Radix et Rhizoma (Gancao).

Glycyrrhizae Radix et Rhizoma (Gancao) is a functional food whose quality varies significantly between distinct geographical sources owing to the influence of genetics and the geographical environment. This study employed three-dimensional fluorescence coupled with alternating trilinear decomposition (ATLD) and random forest (RF) algorithms to rapidly predict Gancao species, geographical origins, and primary constituents. Seven fluorescent components were resolved from the three-dimensional fluorescence of the ATLD for subsequent analysis. Results indicated that the RF model distinguished Gancao from various species and origins better than other algorithms, achieving an accuracy of 94.4 % and 88.9 %, respectively. Furthermore, the RF regressor algorithm was used to predict the concentrations of liquiritin and glycyrrhizic acid in Gancao, with 96.4 % and 95.6 % prediction accuracies compared to HPLC, respectively. This approach offers a novel means of objectively evaluating the origin of food and holds substantial promise for food quality assessment.

UHPLC-QTOF-MS-based untargeted metabolomic authentication of Chinese red wines according to their grape varieties.

Wine is a very popular alcoholic drink owing to its health benefits of antioxidant effects. However, profits-driven frauds of wine especially false declarations of variety frequently occurred in markets. In this work, an UHPLC-QTOF-MS-based untargeted metabolomics method was developed for metabolite profiling of 119 bottles of Chinese red wines from four varieties (Cabernet Sauvignon, Merlot, Cabernet Gernischt, and Pinot Noir). The metabolites of red wines from different varieties were assessed using orthogonal partial least-squares discriminant analysis (OPLS-DA) and analyzed using KEGG metabolic pathway analysis. Results showed that the differential compounds among different varieties of red wines are mainly flavonoids, phenols, indoles and amino acids. The KEGG metabolic pathway analysis showed that indoles metabolism and flavonoids metabolism are closely related to wine varieties. Based on the differential compounds, OPLS-DA models could identify external validation wine samples with a total correct rate of 90.9 % in positive ionization mode and 100 % in negative ionization mode. This study indicated that the developed untargeted metabolomics method based on UHPLC-QTOF-MS is a potential tool to identify the varieties of Chinese red wines.