Transcriptomic analyses of public datasets (TCGA and GTEx) revealed that both CD74 and Cathepsin L (CTSL) are significantly overexpressed in diffuse large B-cell lymphoma (DLBCL) compared to normal tissues, and that their expression levels are highly correlated to each other (Spearman R = 0.64, p = 3 × 10−46). Kaplan–Meier analysis showed that elevated expression of both genes is associated with reduced overall survival (OS), defining a high-risk CD74+/CTSL+ DLBCL subgroup. This is the first study demonstrating coordinated overexpression of CD74 and CTSL and proposing their dual targeting via antibody–drug conjugates (ADCs) to improve outcomes in relapsed or refractory DLBCL. Cysteine cathepsins, a family of proteases, are upregulated in many cancers, facilitating tumor invasion and metastasis. Cathepsins are overexpressed and play key roles in DLBCL progression. GB111-NH2, a potent broad-spectrum cathepsin inhibitor, significantly reduced cathepsin activity in lymphoma cell lines and patient samples. GB111-NH2 treatment increased apoptosis and caspase-3 activation in DLBCL patient cells and chronic lymphocytic leukemia (CLL) mononuclear cells. Here, we developed a modified cathepsin inhibitor, M-GB, containing a maleimide linker for site-specific antibody conjugation. While M-GB alone has poor cell permeability, when conjugated to an antibody, it forms an ADC (M-GB–ADC) that selectively induces lymphoma cell death. One M-GB–ADC demonstrated high specificity for CD74-expressing lymphoma cells while exhibiting minimal toxicity to non-target cells in vitro. Our findings highlight the potential of another M-GB–ADC as a targeted therapy for overcoming rituximab resistance and treatment failure in DLBCL. This strategy enhances therapeutic efficacy and represents a preclinical proof-of-concept treatment option by directing a cathepsin-inhibitor payload specifically to malignant B cells.

Streptococcus suis is a zoonotic pathogen that poses a significant threat to both the swine industry and human health. This bacterium utilizes a type VII secretion system (T7SS) to translocate effector proteins that mediate bacterial competition and contribute to virulence. However, the functions of T7SS effectors in S. suis remain poorly understood. In this study, we identified and characterized LXG-T2, a T7SS-secreted toxin from S. suis virulent strain WUSS351. Bioinformatics analysis revealed that LXG-T2 harbors a C-terminal glycine zipper motif, a structural feature commonly associated with membrane-disrupting toxins. Functional assays demonstrated that LXG-T2 exhibits strong bactericidal activity against E. coli and provides S. suis with a competitive advantage. Furthermore, the LXG-T2 has the capacity to compromise the integrity of bacterial membranes, as evidenced by the observed increase in membrane permeability and depolarization in target cells. Moreover, LXG-T2 exhibited cytotoxic effects on host cells and promoted S. suis survival in a murine infection model. Collectively, our findings establish LXG-T2 as a T7SS effector mediated membrane disruption to enhance both bacterial competition and virulence. This work not only reveals a novel mechanism by which S. suis manipulates microbial communities, but also highlights the significance of T7SS effectors as key mediators of pathogenesis in Gram-positive bacteria.

The effluents generated during the process of hair dyeing exhibit a complex composition, comprising chemical compounds with varying toxicity levels. While the adverse impact of hair dyes on human health is acknowledged, there is a notable absence of studies addressing the toxicity associated with effluents produced during these activities. The primary objective of this study was to assess two effluents emanating from beauty salons after brown hair dyeing: one resulting from hair washing with water, shampoo, and conditioner, referred to as the complete effluent (CE), and the other from washing the dyed hair solely with water, excluding surfactants, referred to as the dye effluent (DE). Invitro bioassays were conducted with the human hepatoma cell line (HepG2/C3A). Cytotoxicity was evaluated through 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and Trypan Blue tests, while genotoxicity and mutagenicity were assessed by comet assay and cytokinesis-block micronucleus test, respectively. The cells were exposed for 4 h to various dilutions of the two sampled effluents [100.0% (1); 50.0% (2); 25.0% (3); 12.5% (4); 6.25% (5); 3.125% (6)]. Cytotoxicity was induced by CE-1, DE-1, DE-2, and DE-3 dilutions as indicated by both assays, whereas CE-2 dilution exhibited cytotoxicity solely through the MTT assay. These findings suggest impaired cell membrane integrity, permeability, and mitochondrial activity. Nontoxic dilutions (4, 5, and 6) were viable for the comet assay and micronucleus test, revealing genotoxicity without mutagenic potential. Consequently, residual concentrations of effluents were found to induce nonlethal and reparable primary DNA damage. Moreover, the effluents decreased the cytokinesis-block proliferation index and the cell replication index, indicating interference and arrestment in the cell cycle. These outcomes highlight the potential threat posed by residual concentrations of hair DEs to environmental quality and human health, emphasizing the imperative need for pre-disposal treatments in salon settings.

Sphingosine-1-phosphate receptor-3 (S1PR3) has been implicated in the maintenance of cerebrovascular integrity during cerebral ischemia. This study aimed to elucidate the impact of S1PR3 inactivation on oxidative stress-induced cerebrovascular endothelial cell permeability and to explore the potential mechanisms. In bEnd3 cells, blockade of S1PR3 exacerbated H2O2-induced endothelial hyperpermeability, ZO-1 redistribution, reactive oxygen species (ROS) accumulation and cell viability loss. A previous study indicated that the activation of p38 and extracellular signal-regulated kinase (ERK) pathway was essential for H2O2-mediated cytosolic phospholipase A2 (cPLA2) phosphorylation in bEnd3 cells. This study revealed that the activation of c-Jun N-terminal kinase (JNK) was crucial for H2O2-induced signal transducer and activator of transcription 3 (STAT3) phosphorylation. In bEnd3 monolayer, either blockade or genetic knockdown of S1PR3 further enhanced the phosphorylation level of p38, ERK, cPLA2, JNK and STAT3 in response to H2O2 exposure. Furthermore, lentivirus-mediated knockdown of S1PR3 intensified H2O2-induced ZO-1 redistribution, paracellular hyperpermeability and ROS accumulation, while further compromising endothelial cell viability. Collectively, these results identified S1PR3 as a critical guardian of endothelial barrier integrity under oxidative stress, acting through cPLA2- and STAT3-dependent signalling pathways.

ABSTRACTGlioblastoma (GB) is a highly aggressive brain tumour with a poor prognosis and limited responsiveness to standard chemotherapy, particularly temozolomide (TMZ), due to intrinsic resistance mechanisms. This study investigates the potential of Aesculus hippocastanum, known as horse chestnut extract (HCE), to enhance the therapeutic efficacy of TMZ in GB cells through modulation of the Wnt/β‐catenin signalling pathway. Combined treatment of HCE (500 μg/mL) and TMZ (100 μM) significantly reduced cell viability and inhibited wound healing and colony formation compared to either agent alone at 48 h. Notably, the expression of β‐catenin and Wnt‐1 was significantly reduced in the combination group, followed by a significant downregulation of Nestin and β3‐tubulin, markers of glioma stem‐like cells and aggressiveness, respectively. Furthermore, apoptotic activity was significantly increased following the combined treatment. In a 3D U87‐spheroid model, the combination therapy resulted in a substantial reduction in spheroid area, suggesting impaired tumour growth. Propidium iodide (PI) staining revealed increased membrane permeability in cells treated with the combination, which was accompanied by an increase in p53 expression, supporting the induction of apoptosis. Collectively, these findings demonstrate that HCE increases the cytotoxic effects of TMZ by inhibiting Wnt/β‐catenin signalling, reducing tumour stemness, and promoting apoptotic pathways in GB cells.

Construction of a biocompatible supramolecular sensor for fluorescence imaging of Nitric oxide in plant tissues.

Aberrant protein glycosylation is known to induce immune suppression, which can contribute to cancer malignancy and metastasis. The enzymes responsible for glycan biosynthesis may be modulated as a therapeutic strategy to improve the anticancer immune response. Kifunensine is a potent inhibitor of one of these enzymes, type I α-mannosidase, which can increase cell surface glycosylation through upregulation of high mannose N-glycans over branched and complex N-glycans. However, the high polarity of Kifunensine limits its cell permeability. Here, we report two hydrophobic analogues of Kifunensine designed for improved cell permeability. The hydrophobic analogues were formulated in an ethyl oleate oil vehicle to prolong release in vivo . The therapeutic efficiency of the developed formulation was examined using an immunocompetent mice model of colon cancer, which showed a marginal delay in the tumor growth with treatment. Immunofluorescence analysis demonstrated overexpression of high mannose N-glycans after treatment with both analogues, but only a slight increase in immune cell infiltration was observed. While previously reported studies showed significant upregulation of immune cells with Kifunensine, the present study suggests that Kifunensine may have limited efficacy as a monotherapy in colon cancer. • Ethyl oleate was shown to load the lipophilic Kifunensine analogues and sustain their release • The study supports the application of cyanovirin-N lectin for the detection of cells displaying high mannose N-glycans • Lipophilic Kifunensine analogues significantly upregulated high mannose N-glycans • The most lipophilic Kifunensine analogue (JDW-II-008) showed marginal antitumor activity compared to the parent drug • A slight immune cells infiltration was observed following Kifunensine analogues treatment

Introduction. Recently, flow cytometry has gained the attention of clinical microbiologists for its ability to characterize bacterial species. This article shows how acoustic-enhanced flow cytometry, combined with the fluorescent dye SYTO 9, can differentiate between Gram-positive and Gram-negative bacteria.Gap Statement. SYTO 9 is a cell membrane-permeable dye with a high affinity for nucleic acids in both living and non-living prokaryotic and eukaryotic cells and has been used as a counterstain to discriminate between live and dead cells in combination with other dyes. However, the consistency of its cell permeability in different bacterial species and its potential application to Gram differentiation has not been fully considered.Aim. We sought to assess the suitability of the fluorescent dye, SYTO 9, to differentiate Gram-positive and Gram-negative bacteria by flow cytometry.Methodology. A range of common Gram-positive and Gram-negative bacterial species were stained with SYTO 9, then processed using an acoustic-enhanced flow cytometer (Attune NxT, Thermo Fisher). The fluorescence emission data were gated and analysed in quadrant plots.Results. Single and polymicrobial bacterial suspensions stained with SYTO 9 produced different fluorescence signals in Gram-positive and Gram-negative bacteria in the forward scatter-height/blue 3-height (FSC-H/BL3-H) quadrant. Gram-positive species (Staphylococcus aureus, Enterococcus faecalis, Staphylococcus epidermidis, Streptococcus pneumoniae and Streptococcus pyogenes) had higher fluorescence intensities in the BL3 channel than the Gram-negative species (Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa, Burkholderia thailandensis and Proteus vulgaris) in both single and mixed cultures.Conclusion. The FSC-H/BL3-H quadrant analysis of flow cytometer emission spectra from SYTO 9-stained bacterial suspensions segregated Gram-positive and Gram-negative bacteria into separate quadrants based on their different fluorescence intensities. This provides a single-dye flow cytometry method for Gram differentiation.

Endothelial cells play a central role in the pathogenesis of sepsis. Currently, effective therapeutic options for sepsis remain limited. Baicalein (BAI) is a compound with multiple bioactivities. This study aims to investigate the protective effects of BAI against lipopolysaccharide (LPS)-induced endothelial cell injury and to explore the underlying molecular mechanisms. Bioinformatics tools, including RNA sequencing (RNA-seq), immune cell infiltration analysis, Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses, were utilized to explore the molecular mechanisms of BAI in human umbilical vein endothelial cells (HUVECs) induced by LPS. The LPS-induced HUVECs model was used to assess the effects of BAI through CCK8 assays, cell permeability assays, and RT-qPCR. RNA-seq analysis revealed a set of differentially expressed genes (DEGs) shared between the Control group vs LPS group and LPS vs Baicalein group, including vascular cell adhesion molecule 1 (VCAM1), phosphoinositide-specific phospholipase C X-domain containing 1 (PLCXD1), and MIR3142host gene (MIR3142HG), a long non-coding RNA. GO, KEGG, and Reactome enrichment analyses indicated that the DEGs were primarily enriched in TNF signaling, NF-κB signaling, and immune regulation pathways. Molecular docking and molecular dynamics simulation analyses revealed that BAI exhibits a strong binding affinity for key targets VCAM1 and PLCXD1. ROC analysis revealed that these core genes, VCAM1 and PLCXD1, exhibited significant diagnostic potential for sepsis. The cell experimental results demonstrated that BAI significantly alleviated the expression levels of inflammatory markers (such as IL-6 and IL-1β) and reduced endothelial cell permeability induced by LPS in HUVECs. BAI may alleviate LPS-induced endothelial cell injury by modulating the inflammatory response and immune microenvironment through the regulation of MIR3142HG, VCAM1, and PLCXD1 targets. This study provides new molecular targets and theoretical insights for sepsis therapy.

Quercetagetin phospholipid complex self-microemulsifying delivery system for enhanced oral delivery and alcoholic liver injury protection.

Highly selective and permeable DDR membranes for CO2/CH4 separation in a wide temperature range

Effects of biochar addition and semi-permeable membrane covering on aerobic composting: Insights into carbon–nitrogen transformations and bacterial community and function succession

Oxygen permeability and stability in the entropy-stabilized Co-based perovskite oxygen permeable membranes

Synergistic channels: A review of hollow nanoparticles for enhanced selective permeability in thin film nanocomposite membranes

Enzyme-coupled polymersome microreactor for point-of-care blood urea sensing.

Elapid snakebites cause severe toxicity, predominantly neurotoxicity and general cytotoxicity. However, the specific cellular impacts of individual venom toxins remain largely underexplored. This study developed a high-throughput platform for profiling cytotoxicity from elapid venoms, focusing on nanofractionation analytics to enhance selectivity and toxin identification. Elapid Venoms were tested on four human cell lines, representing kidney (RPTEC/TERT1), liver (HepaRG), endothelial (iPSC-EC), and skin (HaCaT) tissues. Cytotoxic effects were assessed through cell coverage, viability, and metabolic assays in both crude and nanofractionated venom samples. Nanofractionation revealed selective cytotoxicity in venom components, notably phospholipases A2 (PLA2s) and three-finger toxins (3FTxs), which impaired membrane integrity and cellular metabolism. Crude B. multicinctus venom displayed specific cytotoxicity toward liver and skin cells but not kidney or endothelial cells. Cytotoxicity of nanofractionated B. multicinctus venom was lost, likely due to denaturing conditions of the reversed-phase separation. Fractionation after size exclusion chromatography (SEC) for post-column bioassaying to avoid toxin denaturation yielded bioactive fractions, with 3FTxs, PLA2s, and Kunitz-type serine protease (KUNs) likely responsible for the observed cell permeability disruption, extracellular matrix (ECM) degradation, and metabolic loss. This integrated analytical workflow, combining nanofractionation with high-throughput cytotoxicity assays and venomics, enabled rapid identification of venom components with cell type-specific toxicity. Our findings contribute to understanding elapid venom toxicity and can aid in developing targeted snakebite treatments focusing on cytotoxicity responsible for tissue-specific damage.

The stereoisomers of lactate, L- and D- are not only metabolic substrates but also signalling molecules, capable of activating and signalling through its G protein-coupled receptor, Hydroxycarboxylic acid receptor 1 (HCAR1). These stereoisomers are both produced by the gut microbiota at millimolar concentrations creating a physiological environment for lactate-sensing unique to the gut yet, poorly understood. Here we identify a role for D-/L-lactate on intestinal barrier function. A human colonic epithelial cell model, Caco2, activated Gαi signalling in response to both L- and D-lactate, although L-lactate exhibited a more potent and rapid Gαi signal profile. When differentiated, apically but not basally treated D-/L-lactate enhanced tight junctions and reduced cell permeability, consistent with the apical localization of HCAR1. This improved barrier function occurred in a Gαi-dependent manner. In addition, apical lactate rescued the reduced intestinal barrier function induced by lipopolysaccharides. This work highlights the potential for D-/L-lactate supplementation in improving gut health.

With increased demand for protein, microorganisms such as bacteria and fungi are considered potentially important protein sources. Yet, these microbial proteins are encapsulated by their cell walls, which limits their solubility and in turn restricts any possible emulsifying or foaming abilities for food applications . In this study, we aimed to improve the soluble protein recovery from a Gram-positive bacterium Clostridium tyrobutyricum and a filamentous fungus Rhizomucor pusillus and assess the quality of the soluble extracts. The suspensions were pretreated by pH adjustments followed by microfluidisation, ultrasonication or a combination of the two to permeabilise and disrupt the cells. The effectiveness of the treatments were assessed by the soluble protein recovery, particle size and microscopy while the soluble fraction was characterised based on its relative composition and size. Furthermore, the energy consumed for each treatment was computed to understand the process efficiency. pH greatly influenced the cell permeability and disruption effectiveness. Bacterial cells were more easily lysed at low pH using ultrasonication (49.6% recovery), achieving a high efficiency of 10 kJ/g protein recovered and obtaining protein-rich soluble fractions that assembled into worm-like aggregates. Conversely, the highest recovery for the filamentous fungus occurred using a combined treatment at high pH (29.1% recovery), which was less energy-efficient than ultrasonication alone (358 kJ/g protein recovered). Unlike bacteria, the soluble fungal extract contained more carbohydrates than proteins with larger hydrodynamic sizes. These findings demonstrate the influence of microbial cell structure and composition on protein extractability and its characteristics for future implementation in food applications.

Cyclic peptides are a promising class of therapeutics due to their attractive drug qualities such as increased structural stability, cell permeability, and resistance to proteolytic degradation. With recent advancements in cyclic peptide backbone generation models like CyclicCAE and RFPeptide, generating cyclic peptide backbones can be done more rapidly compared to traditional algorithm or physics based approaches. However, designing energetically favorable cyclic peptide sequences to fit generated backbones using only canonical amino acids is nontrivial. We fine-tuned the state-of-the-art deep learning model for protein sequence design, ProteinMPNN, using a combination of X-ray crystal structures from the Protein Data Bank and in silico generated cyclic peptides. Our approach surpasses ProteinMPNN in cyclic peptide sequence design, producing energetically stable sequences with a higher success rate of folding into the generated cyclic peptide backbones. We show that CyclicMPNN can be used as a motif-inpainting strategy and in de novo sequence design tasks. We propose that CyclicMPNN will enable the rapid design of energetically stable cyclic peptide sequences, increasing the success rate of therapeutic cyclic peptide development.

The in vitro antifungal effect of cocoa mucilage (Theobroma cacao L.) on Moniliophthora roreri was evaluated in order to define the most effective fermentation conditions and concentrations. A completely randomized design with a factorial arrangement was applied in the laboratory, considering fermentation of 5, 10, and 15 days and concentrations of 10, 7.5, 5, and 2.5 percent, in addition to a control. Activity was estimated by mycelial growth and changes in membrane integrity, using electrical conductivity and absorbance at 256 nanometers. Mucilage fermented for 10 and 15 days reduced fungal growth and increased cell permeability compared to 5 days, and concentrations of 10 and 7.5 percent showed the greatest response. The findings suggest that fermentation enhances the bioactivity of mucilage and, without exaggerating its scope, support its potential use as a sustainable alternative for the management of cocoa moniliasis in local cocoa production systems.