Search Results (301)

Diabetes is a risk factor for periodontitis. Increasing evidence suggests that a central player in this link is the vacuolar H+-ATPase (V-ATPase), which provides a physical and functional core for regulation by the catabolic lysosomal AMP-activated protein kinase complex (L-AMPK) and the anabolic mammalian target of rapamycin complex 1 (mTORC1). These complexes detect levels of various cellular nutrients, including glucose at the lysosome, and promote cellular responses to restore homeostasis. The high-glucose conditions of diabetes foster anabolic mTORC1 signaling that increases inflammation and inflammatory bone resorption in response to periodontal infections. Here, we review the structure and composition of V-ATPase, L-AMPK, mTORC1, and other elements of the energy-sensing platform. Mechanisms by which V-ATPase passes signals to the complexes are examined and recent data are reviewed. Current anti-bone resorptive therapeutics, bisphosphonates and denosumab, enhance the risk of medicine-related osteonecrosis of the jaw (MRONJ) and are not used to treat periodontal bone loss. Accumulating data suggest that it may be possible to target inflammatory bone resorption through agents that stimulate L-AMPK, including metformin and glucagon-like peptide-1 agonists. This approach may reduce inflammatory bone resorption without major effects on overall bone remodeling or increased risk of MRONJ. Full article

Vesicle-associated membrane protein (VAMP)-associated protein B (VAPB) serves as a tethering factor that interacts with various proteins and recruits these proteins to the ER surface, exerting multiple functions, such as organelle membrane tethering, lipid transfer between organelles, regulation of calcium homeostasis, autophagy, and the unfolded protein response (UPR). Its interaction is often mediated by its MSP (major sperm) domain, which binds with FFAT (two phenylalanines in an acidic tract)-motif-containing proteins. However, pathogenic variations, such as P56S, P56H, and T46I, in the VAPB MSP domain lead to the familial form of amyotrophic lateral sclerosis (ALS8). Still, the underlying pathophysiology of ALS8 due to pathogenic variations in the VAPB MSP domain remains elusive. In this study, we conducted molecular dynamics (MD) simulations to understand the pathogenic-variant-derived changes in the structural dynamics of the VAPB MSP domain. We found that pathogenic variants altered the fluctuations and conformational dynamics of the VAPB protein. Analyzing the organizations of the secondary structure revealed that pathogenic variants changed the composition of secondary structure elements, especially increasing the proportion of α-helix while reducing β-sheet formation, which might affect the organelle tethering and other functions of VAPB, as well as VAPB homodimer and heterodimer formation. Taken together, these findings can be further investigated through in vivo and/or in vitro studies to not only clarify the pathophysiology of ALS8 resulting from VAPB MSP domain pathogenic variants but also develop novel therapeutics for the disease that restore the native structural organizations as well as fluctuations and motions. Full article

Factor VIII (FVIII) interacts with Endoplasmic Reticulum (ER) chaperones Calnexin (CANX) and Calreticulin (CALR) and with ER-Golgi Intermediate Compartment (ERGIC) transporters, Lectin, mannose-binding 1 (LMAN1) and Multiple Coagulation Deficiency 2 (MCFD2). We previously reported that the Gamma-aminobutyric Acid Receptor-associated proteins (GABARAPs) also influence FVIII secretion. Here, we further investigated the intracellular dynamics of FVIII using single and double CRISPR/Cas9 Knockout (KO) models of the abovementioned chaperones as well as the GABARAP proteins in HEK293 cells expressing FVIII. Cellular pathways were manipulated by Brefeldin A (BFA), Chloroquine (CQ), a Rab7 inhibitor, and subjected to glucose starvation. The effect of each KO on FVIII secretion and organelle distribution was assessed by a two-stage chromogenic assay and immunofluorescence (IF) microscopy, prior and upon cell treatments. Using these approaches, we first observed distinct effects of each studied protein on FVIII trafficking. Notably, intracellular localization patterns revealed clustering of FVIII phenotypes in GABARAP^KO, CANX^KO, and CALR^KO cells together under both basal and treated conditions, an observation that was also reflected in their respective double KO combinations. Besides, a clear involvement of additional components of the endomembrane system was evident, specifically at the trans-Golgi space, as marked by FVIII colocalization with the Ras-like proteins in brain (Rab8 and Rab7) and with the Vesicle-Associated Membrane Protein (VAMP8), along with the observed impact of the selected cell treatments on FVIII phenotypes. These outcomes enhance our understanding of the molecular mechanisms regulating FVIII and pave the way for new perspectives, which could be further projected into FVIII replacement, cell and gene therapies. Full article

The development of novel antimicrobial peptides (AMPs) with broad-spectrum activity represents a promising strategy to overcome multidrug resistance in pathogenic bacteria. In this study, we investigated the antimicrobial activity of three designed peptides—R44K^S*, V31K^S*, and R23F^S*—engineered to incorporate an amyloidogenic fragment from the S1 protein of Staphylococcus aureus and one or two cell-penetrating peptide (CPP) fragments to enhance cellular uptake. The antimicrobial efficacy of these peptides and their combinations was assessed against Pseudomonas aeruginosa, Escherichia coli, Staphylococcus aureus, methicillin-resistant S. aureus (MRSA), and Bacillus cereus. The results demonstrated that all three peptides exhibited significant antibacterial activity in a concentration-dependent manner, with R44K^S* being the most potent. Peptide combinations, particularly V31K^S*/R23F^S* and R44K^S*/V31K^S*, showed enhanced inhibitory effects and reduced minimum inhibitory concentrations (MICs), suggesting synergistic or additive interactions. Fractional inhibitory concentration index (FICI) analysis confirmed that most combinations exhibited synergy or additive effects. These findings highlight the potential of CPP-modified peptides as antimicrobial agents and underscore the importance of optimizing peptide combinations for therapeutic applications. Full article

This study evaluated immune gene expression and locomotor behavior across five Apis mellifera subspecies (Carniolan, Caucasian, Syrian, Muğla ecotype, and Yığılca ecotype) following controlled Vairimorpha ceranae infection. Six days post-infection, Caucasian, Carniolan, and Yığılca bees exhibited a significant upregulation of antimicrobial peptide (AMP) transcripts—hymenoptaecin, abaecin, defensin, and apidaecin—indicating a robust humoral response. Conversely, Syrian and Muğla bees showed weaker AMP expression and higher V. ceranae mRNA levels, indicating lower immunity and higher susceptibility. Positive correlations among AMP transcripts, especially in Caucasian, Carniolan, and Yığılca bees, suggested a coordinated response. Eater gene expression, critical for cellular immunity, decreased in infected Caucasian and Yığılca bees, coinciding with AMP upregulation. Vitellogenin expression, linked to immunity and longevity, increased in Carniolan and Syrian bees, correlating with higher early locomotor activity. Locomotor analysis revealed subspecies-specific behavioral responses. Syrian bees maintained the highest activity despite elevated V. ceranae mRNA and minimal AMP expression, suggesting unique resilience possibly mediated by vitellogenin. Muğla bees, despite high pathogen loads, exhibited decreased activity. Caucasian bees showed strong immune responses but reduced activity post-infection, reflecting potential physiological trade-offs. Overall, these findings underscore the role of genetic variability in shaping honey bee immune and behavioral responses to Vairimorpha and support subspecies-targeted breeding and disease management strategies to enhance resilience. Full article

Tetanus, a severe and life-threatening illness caused by Clostridium tetani, produces symptoms such as muscle spasms, muscle stiffness and seizures caused by the production of tetanus neurotoxin (TeNT). TeNT causes spastic paralysis through the inhibition of neurotransmission in spinal inhibitory interneurons. This is achieved, in part, through pH-triggered membrane insertion of the translocation (HCT) domain, which delivers the catalytic light-chain (LC) domain to the cytosol. While the function of HCT is well defined, the mechanism by which it accomplishes this task is largely unknown. Based on the crystal structure of tetanus neurotoxin, we identified potential polar interactions between arginine 711, tryptophan 715 and aspartate 821 that appear to be evolutionarily conserved across the clostridial neurotoxin family. We show that the disruption of the Asp-Arg pair in a beltless HCT variant (bHCT) results in changes in thermal stability without significant alterations to the overall secondary structure. ANS (1-anilino-8-napthalene sulfonate) binding studies, in conjunction with liposome permeabilization assays, demonstrate that mutations at R711 or D821 trigger interactions with the membrane at higher pH values compared to wildtype bHCT. Interestingly, we show that the introduction of the D821N mutation into LH_NT (LC-HCT only), but not the holotoxin, resulted in the increased cleavage of VAMP 2 in cortical neurons relative to the wildtype protein. This suggests that, as observed for botulinum toxin A, the receptor-binding domain is not necessary for LC translocation but rather helps determine the pH threshold of membrane insertion. The mutation of W715 did not result in detectable changes in the activity of either bHCT or the holotoxin, suggesting that it plays only a minor role in stabilizing the structure of the toxin. We conclude that the protonation of D821 at low pH disrupts interactions with R711 and W715, helping to drive the conformational refolding of HCT needed for membrane insertion and the subsequent translocation of the LC. Full article

Energy and environmental challenges are driving researchers to explore cost-effective and eco-friendly nanomaterial fabrication methods. In this study, Atmospheric Pressure Microplasma (AMP) was used to synthesize iron oxide nanoparticles at varying molar concentrations of ferrous sulfate (0.5 M, 1 M, and 1.5 M) under a 15 kV discharge voltage for 90 min. The X-ray diffraction (XRD) results confirmed the formation of mixed cubic and hexagonal phases of magnetite and hematite nanoparticles. The particle size, calculated using the Debye–Scherrer formula, ranged from 9 to 11 nm, depending on the precursor concentration. Scanning electron microscopy (SEM) images revealed spherical nanoparticles at 0.5 M, while agglomeration occurred at 1.5 M. The energy-dispersive X-ray spectroscopy (EDS) analysis confirmed the presence of iron and oxygen in the synthesized nanoparticles. Fourier-transform infrared (FTIR) and UV spectroscopy showed characteristic absorption bands of iron oxide. The impact of the particle size and lattice strain on the optical properties of the nanoparticles was also studied. Smaller nanoparticles exhibited an excellent specific capacitance (627) and a strong charge–discharge performance in a 3 M KOH solution, with a high energy density (67.72) and power density (2227). As photocatalysts, the nanoparticles demonstrated a 97.5% and 96.8% degradation efficiency against methylene blue (MB) and methyl orange (MO), respectively, with high rate constants. These results surpass previous reports. The enhanced electrochemical performance and photocatalytic activity are attributed to the high-quality iron oxide nanoparticles, showing an excellent cyclic stability, making them promising for supercapacitors and environmental remediation. Full article

The ubiquitin–proteasome system (UPS) plays a pivotal role in determining protein fate, regulating signal transduction, and maintaining cellular homeostasis. Protein ubiquitination, a key post-translational modification, is orchestrated by the sequential actions of three primary enzymes, ubiquitin-activating enzyme (E1), ubiquitin-conjugating enzyme (E2), and ubiquitin protein ligase (E3), alongside the regulatory influence of deubiquitinases (DUBs) and various cofactors. The process begins with E1, which activates ubiquitin molecules. Subsequently, E2 receives the activated ubiquitin from E1 and transfers it to E3. E3, in turn, recognizes specific target proteins and facilitates the covalent attachment of ubiquitin from E2 to lysine residues on the target protein. Among the E2 enzymes, ubiquitin-conjugating enzyme E2O (UBE2O) stands out as a unique E2–E3 hybrid enzyme. UBE2O directly mediates the ubiquitination of a wide array of substrates, including 5′-AMP-activated protein kinase catalytic subunit alpha-2 (AMPKα2), MAX interactor 1 (Mxi1), and v-maf musculoaponeurotic fibrosarcoma oncogene homolog (c-Maf), among others. In this narrative review, we will explore the structural characteristics of UBE2O and elucidate its molecular functions. Additionally, we will summarize recent advancements in understanding the role of UBE2O in various tumors, Alzheimer’s disease (AD), and metabolic diseases. Finally, we will discuss the potential of targeting UBE2O as a novel therapeutic strategy for the treatment of human diseases. Full article

This study reports the synthesis and characterization of a novel carboxymethyl cellulose–N-fullerene–g-poly(co-acrylamido-2-methyl-1-propane sulfonic acid) (CMC–N-fullerene–AMPS) hydrogel for potential application in biosensing within food packaging. The hydrogel was synthesized via free radical polymerization and characterized using FTIR, SEM, and fluorescence microscopy. FTIR analysis confirmed the successful grafting of AMPS and incorporation of N-fullerenes, indicated by characteristic peaks and a shift in the N–H/O–H stretching frequency. Density Functional Theory (DFT) calculations revealed that the CMC–N-fullerene–AMPS hydrogel exhibited higher stability and a lower band gap energy (0.0871 eV) compared to the CMC–AMPS hydrogel, which means a high reactivity of CMC–N-fullerene–AMPS. The incorporation of N-fullerenes significantly enhanced the hydrogel’s antibacterial activity, demonstrating a 22 mm inhibition zone against E. coli and a 24 mm zone against S. aureus, suggesting potential for active food packaging applications. Critically, the hydrogel displayed a unique “turn-on” fluorescence response in the presence of bacteria, with distinct color changes observed upon interaction with E. coli (orange-red) and S. aureus (bright green). This fluorescence enhancement, coupled with the porous morphology observed via SEM (pore size 377–931 µm), suggests the potential of this hydrogel as a sensing platform for bacterial contamination within food packaging. These combined properties of enhanced antibacterial activity and a distinct, bacteria-induced fluorescence signal make the CMC–N-fullerene–AMPS hydrogel a promising candidate for developing intelligent food packaging materials capable of detecting bacterial spoilage. Full article

A family of improved low-voltage switched-capacitor circuits is introduced. It is based on the utilization of multiple-output class AB/AB op-amp architectures that provide true sample and hold outputs that are not subject to a reset phase as with conventional switched-capacitor circuits. This feature essentially relaxes the op-amp slew rate requirements, allowing a higher speed and simple low-voltage operation. A power-efficient GB boosting technique based on resistive local common mode feedback is used to significantly improve the GB and internal/external slew rate of the op-amps with only a 36.5% additional power dissipation. Full article

Despite its ecological and economic importance, many aspects of Varroa destructor’s biology remain poorly understood, particularly its defense mechanisms against pathogens. The limited knowledge of Varroa’s immunity has hindered the development of RNA interference (RNAi)-based strategies targeting immune-related genes. In this study, we investigated the immune gene repertoire of V. destructor by querying its NCBI nr protein database and comparing it to model species of ticks (Ixodes scapularis) and mites (Galendromus occidentalis and Tetranychus urticae). Transcription of candidate immune genes was confirmed by analyzing a de novo assembled transcriptome of V. destructor. Our findings reveal that V. destructor shares key immunological traits with ticks, including lysozymes, chitinases, and thioester-containing proteins (TEPs), but also shares the absence of transmembrane peptidoglycan recognition proteins (PGRPs), Gram-negative binding proteins, and several lectin families involved in pathogen recognition. Additionally, Varroa mites, like ticks, lack homologs of crucial immune signaling components, such as the unpaired ligand (JAK/STAT), Eiger (JNK), and multiple elements of the IMD pathway. They also do not encode canonical antimicrobial peptides (AMPs) like defensins but possess putative homologs of ctenidins, AMPs previously identified in spiders and ticks, which may be adopted as a novel genetic readout for immune response in mites. Our findings lay the groundwork for future functional studies on mite immunity and open new avenues for RNAi-based biocontrol strategies targeting immune pathways to enhance Varroa management. Full article

Objective: This in vitro study aimed to investigate the impact of folic acid on DNA methylation and gene expression in adipocytes from subcutaneous adipose tissue of patients with systemic lupus erythematosus (SLE), with a focus on the influence of obesity on these epigenetic changes. Methods: Tissue biopsies were collected from patients with normal weight (NW) and obesity (OBS). Adipocytes were isolated via enzymatic digestion and density separation. Each group was divided into control (standard medium) and folic acid treatment (2 mg/24 h for 48 h) conditions. After treatment, DNA methylation levels were analyzed using the Infinium Methylation EPIC v2.0 Kit, and gene expression analyses were performed by RT-qPCR. A pathway enrichment analysis was conducted using the KEGG database for functional insight. Results: Folic acid induced differential methylation at 755 CpG sites in NW adipocytes, which were associated with immune regulation, including MAPK signaling. Also, OBS adipocytes showed methylation changes at 92 CpG sites, affecting pathways related to metabolic regulation, such as cAMP signaling. LEP gene expression was upregulated (5.2-fold) in OBS adipocytes, while CREM2 expression was increased (2.8-fold) in NW adipocytes after treatment. These gene expression differences underscore weight-dependent responses to folic acid, with LEP upregulation in OBS cells suggesting links to metabolic dysregulation and CREM2 upregulation in NW cells potentially contributing to immune modulation. Conclusions: Folic acid treatment exerts distinct epigenetic and gene expression effects in adipocytes of SLE patients, modulated by obesity status. This weight-dependent response, marked by changes in pathways relevant to immune and metabolic function, highlights the need for further investigation into how nutrient-based interventions might support SLE management. From a clinical perspective, this study underscores the potential of targeted nutrient-based interventions to address immunometabolic dysfunctions in SLE patients. Further research could explore folic acid supplementation as a complementary approach to personalized treatment strategies, particularly for patients with obesity. Full article

Background/Objectives: Motile aeromonads are ubiquitous aquatic Gram-negative opportunistic pathogens with environmental, animal, aquatic, and human health implications. Methods: Motile aeromonads were isolated from village pond water samples (n = 100) of the Hisar district of Haryana state in India. Selective isolation and enumeration were followed by biochemical and genotypic identification using gyrB gene; evaluation of seven putative virulence factors and antimicrobial resistance studies and determination of extended spectrum beta lactamase (ESBL) and AmpC beta lactamase (ACBL) enzyme-producing abilities took place. Results: The viable counts of motile aeromonads varied from 1.6 × 10² CFU/mL to 1.2 × 10⁸ CFU/mL. Six species of Aeromonas were identified with high prevalence of A. veronii (74.7%), followed by A. caviae (8.9%), A. hydrophila (7.6), A. jandaei (5%), A. sobria (2.5%), and A. dhakensis (1.3%). PCR amplification of seven genes related to virulence indicated that the majority of the isolates were positive for enolase (eno, 98%), cytotoxic enterotoxin (act, 88%), and hemolysin (asa1, 86%). Many isolates were also positive for type III secretion system inner membrane component (ascV, 53%), ADP-ribosylating toxin (aexT, 47%), and extracellular hemolysin (ahh1, 4%). The antimicrobial resistance (AMR) profile of the isolated Aeromonas isolates indicated the high resistance observed to nalidixic acid (40.2%), cefoxitin (33%), and imipenem (6.2%). In addition, the occurrence of 10.3% ESBL, 32% ACBL, and 29.9% multi-drug resistant (MDR) isolates is alarming. Phylogenetic analysis of gyrB sequences of A. veronii isolates (n = 59) together with GenBank sequences of A. veronii from different geographical regions of the world indicated high genotypic diversity. Conclusions: the village aquaculture ponds in Hisar district have a high occurrence of MDR A. veronii, A. hydrophila, and A. caviae, posing significant animal and public health concern. Full article

The membranes surrounding the eukaryotic cell and its organelles are continuously invaginating, budding, and undergoing membrane fusion–fission events, which enable them to perform functions not found in prokaryotic cells. In addition, organelles come into close contact with each other at membrane contact sites (MCSs), which involve many types of proteins, and which regulate the signaling and transport of various molecules. Vesicle-associated membrane protein (VAMP)-associated protein (VAP) is an important factor involved in the tethering and contact of various organelles at MCSs in almost all eukaryotes and has attracted attention for its association with various diseases, mainly neurodegenerative diseases such as amyotrophic lateral sclerosis (ALS). However, the detailed mechanism of its functional expression remains unclear. In this review, we quantitatively discuss the structural dynamics of the entire molecule, including intrinsically disordered regions and intramolecular and intermolecular interactions, focusing on the vertebrate VAP paralogs VAPA and VAPB. Molecular phylogenetic and biophysical considerations are the basis of the work. Full article

Antimicrobial peptides (AMPs) are biocompatible and biodegradable, making them an attractive alternative to traditional antimicrobial agents and chemical preservatives. Here, a novel α-helix amphiphilic anionic AMP Lc149 was screened from a large yellow croaker (Larimichthys crocea) using a Bacillus subtilis expression system. Lc149 is a hypothesized protein fragment not annotated in the genome of a large yellow croaker. Both extracellular protein and recombinant Lc149 (rLc149) exhibited significant killing effects against Gram-negative Escherichia coli and Vibrio harveyi. Scanning and transmission electron microscopy revealed that rLc149 had the ability to disrupt bacterial cell membranes, causing irregular cell morphology, severe cell membrane damage, cytoplasm agglutination, and intracellular content leakage. Confocal laser scanning microscopy and flow cytometry further confirmed bacterial cell destruction and mortality rates of over 80%. Gel retardation assays and SDS-PAGE electrophoresis showed that rLc149 was unable to bind to bacterial DNA, but did reduce bacterial protein contents. Additionally, rLc149 maintained antibacterial activity against E. coli and V. harveyi upon exposure to temperatures of 25–100 °C, UV radiation time of 0–60 min, pH levels of 3–12, and different proteases. Biosafety assays revealed low hemolytic toxicity to erythrocytes of large yellow croaker, rabbit, and shrimp, and low cytotoxicity to large yellow croaker kidney cells and HEK 293T cells. More deeply, rLc149 also possessed significant killing activity against parasites. Therefore, rLc149 can be considered an antibacterial and antiparasitic drug in fisheries. Full article