Search Results (1,347)

Heterotetramerization of Kv1.1 and Kv1.2 α-subunits expands the functional diversity of voltage-gated potassium Kv1 channels in the central nervous system (CNS), thus necessitating the study of the properties of these heterochannels, including their interactions with ligands. We report on the expression, electrophysiological, and ligand-binding properties of human heterochannels Kv(1.1-1.2)₂ formed by dimeric concatemers Kv1.1-Kv1.2 fused with fluorescent protein mKate2 in Neuro-2a cells. Kv(1.1-1.2)₂ is a low-voltage-activated, highly active, non-inactivating channel with a fast activation rate. Its activation rate and half-maximum activation voltage are similar to that of the Kv1.1 channel, but differ from that of Kv1.2. This suggests that the membrane expression of Kv(1.1-1.2)₂ may functionally compensate for the absence of membrane presentation of homotetrameric Kv1.1 channels in CNS. Hongotoxin 1 fused with fluorescent protein GFP (HgTx-G) is shown to be a pore-blocking ligand of Kv(1.1-1.2)₂ with a dissociation constant of 100 pM. Using confocal microscopy and competitive binding assay, HgTx-G and cells expressing Kv(1.1-1.2)₂, the apparent dissociation constants of the complexes between Kv(1.1-1.2)₂ and peptides Ce1, Ce4, hongotoxin 1, MeKTx11-1, agitoxin 2, charybdotoxin, and scyllatoxin were evaluated to be 14, 33, 40, 250, 800, and >>3300 pM, respectively. Heterotetramerization of α-subunits has a different effect on the affinity of ligands compared to those for Kv1.1 and Kv1.2 channels. Full article

The synergistic effect of using D2EHPA and TOPO together to enhance the extraction of samarium(III) from aqueous media via emulsion liquid membrane (ELM) technology was explored. D2EHPA in binary mixtures with TBP and in ternary mixtures with TOPO and TBP was also tested. Among the tested extractants, a binary mixture of 0.1% (w/w) D2EHPA and 0.025% (w/w) TOPO achieved 100% samarium(III) extraction at a low loading. This mixture outperformed D2EHPA-TBP and other systems because D2EHPA strongly binds to Sm(III) ions, while TOPO increases the solubility and transport efficiency of metal complexes. Additionally, process factors that optimize performance and minimize emulsion breakage were examined. Key insights for successfully implementing the process include the following: 5 min emulsification with 0.75% Span 80 in kerosene at pH 6.7 (natural), 250 rpm stirring, a 1:1 internal/membrane phase volume ratio, a 20:200 treatment ratio, and a 0.2 N HNO₃ stripping agent. These insights produced stable, fine droplets, enabling complete recovery and rapid carrier regeneration without emulsion breakdown. Extraction kinetics accelerate with temperature up to 35 °C but declined above this limit due to emulsion rupture. The activation energy was calculated to be 33.13 kJ/mol using pseudo-first-order rate constants. This suggests that the process is diffusion-controlled rather than chemically controlled. Performance decreases with Sm(III) feed concentrations greater than 200 mg/L and in high-salt matrices (Na₂SO₄ > NaCl > KNO₃). Integrating these parameters yields a scalable, low-loading ELM framework capable of achieving complete Sm(III) separation with minimal breakage. Full article

Background/Objectives: This study evaluated a novel PET tracer, ⁶⁸Ga-NOTA-CYT-200, which targets human granzyme B (GZB) as a biomarker for cytotoxic T-cell activation in a clinically relevant model of melanoma-bearing mice with a humanized immune system treated with immune checkpoint inhibitor (ICI) therapy. Methods: The binding affinity of the tracer was determined using an enzymatic colorimetric assay. Tumor-bearing humanized NSG mice underwent PET imaging before and during ICI monotherapy or combination therapy to assess ⁶⁸Ga-NOTA-CYT-200 uptake within tumors and other organs. The tumor growth was carefully monitored. The treatment response was evaluated based on the percentage change in tumor size at days 5 and 15 after the treatment started. A tracer biodistribution study and immunohistochemical staining of the tumors and organs were also performed. Results: The inhibition constant (Ki) of ⁶⁸Ga-NOTA-CYT-200 was estimated at 4.2 nM. PET imaging showed a significantly higher ⁶⁸Ga-NOTA-CYT-200 uptake in mice receiving the combination therapy compared to those receiving monotherapy or a vehicle (p < 0.0001 or p = 0.0005, respectively), which correlated with the greatest reduction in tumor size in the combination ICI group. Regardless of treatment, the responders presented with a significantly higher ⁶⁸Ga-NOTA-CYT-200 uptake at days 4 or 7 after the treatment began (p = 0.0002 and p = 0.0109, respectively). An increased uptake of ⁶⁸Ga-NOTA-CYT-200, especially in the intestines and liver within the combination ICI group, suggested immune-related adverse events (IrAEs). Conclusions: Our study demonstrates that ⁶⁸Ga-NOTA-CYT-200 PET imaging can predict the early treatment response in melanoma models treated with ICI and may also help in detecting IrAEs. Full article

Astringent persimmons (Diospyros kaki Thunb.) require effective postharvest deastringency treatments due to their high soluble tannin content at harvest. While high CO₂ and ethylene are commonly used to remove astringency, their different effects on fruit firmness and quality require cultivar-specific approaches. This study investigated the transcriptomic and biochemical responses of ‘Daebong’ persimmon to high CO₂ and ethylene treatments during deastringency. Both treatments significantly decreased soluble tannin and total phenolic content, enhancing fruit edibility. However, the firmness was maintained under high levels of CO₂, but it decreased quickly after exposure to ethylene. RNA-Seq analysis identified 2271 differentially expressed genes (DEGs) and revealed distinct transcriptional signatures for each treatment. CO₂ treatment activated hypoxia-responsive genes, stress-related transcription factors (e.g., WRKY, ERF14/26), and components of antioxidant defense (e.g., GSTU17, peroxidases), which contributed to oxidative stress reduction and preservation of firmness. On the other hand, ethylene treatment increased ethylene biosynthesis (ACS), signaling (EIN3-binding F-box), and ripening-related genes (polygalacturonase, laccase, ERF061/113), which promote cell wall degradation and softening. Functional enrichment analysis revealed that various regulatory mechanisms are responsible for the insolubilization of tannins, loss of antioxidants, and changes in firmness. These findings provide new insights into the molecular responses of pollination-constant astringent (PCA) persimmons, particularly the ‘Daebong’ cultivar, to postharvest deastringency treatments for the development of more effective postharvest management strategies. The results suggest that high CO₂ helps maintain fruit quality by promoting stress adaptation and suppressing pathways that lead to softening, whereas ethylene accelerates the ripening process by activating signaling pathways associated with ethylene. Full article

Schiff base ligands prepared from salicylaldehyde and 2-, 3- and 4-methoxybenzylamine were used to prepare copper(II) complexes, characterized by spectral methods, elemental analysis and X-ray crystallography in the case of complex 4a derived from 2-methoxybenzylamine. The DNA cleavage activity of the prepared complexes was exceptional, with best activities of over 95% one-strand cleavage for 4c at 3 mM and full double-strand cleavage for complex 4a at 5 mM. Absorption titration studies with ct-DNA revealed good binding constants (at 10⁵ M⁻¹) with a decrease of up to 56% light absorption. Meanwhile, the EB–DNA displacement method and viscosity studies revealed groove binding as a possible binding mode. For BSA binding studies, all three complexes showed K_BSA values in the optimal range for reversible BSA binding (10⁴ M⁻¹). The copper(II) complexes showed significant cytotoxic effects (67–96% at 1 mM) in mitochondrial activity monitoring assays. Cytotoxicity was confirmed against cancer cell lines (A549 and HepG2) and HEL cells. The complexes 4a and 4c exhibited high activity against HepG2 cancer cells (IC50 < 22 μM), comparable to cisplatin. The radical scavenging activity was determined by the INT method with the best IC50 for 4c (189 ± 11 μM). Overall, complexes 4a and 4c with a methoxy group in the ortho and para positions show high potential in most determined activities, but mainly as DNA cleavers and as cytotoxic agents with selectivity against HepG2 cells. Full article

Potato (Solanum tuberosum L.) is a globally significant staple crop that faces constant threats from Phytophthora infestans, the causative agent of late blight (LB). The battle between Phytophthora infestans and its host is driven by the molecular interplay of RXLR-encoded avirulence (PiAvr) effectors and nucleotide-binding leucine-rich repeat (NLR) immune receptors in potato. This review provides a comprehensive analysis of the structural characteristics, functional diversity, and evolutionary dynamics of RXLR effectors and the mechanisms by which NLR receptors recognize and respond to them. The study elaborates on both direct and indirect modes of effector recognition by NLRs, highlighting the gene-for-gene interactions that underlie resistance. Additionally, we discuss the molecular strategies employed by P. infestans to evade host immunity, including effector polymorphism, truncation, and transcriptional regulation. Advances in structural biology, functional genomics, and computational modeling have provided valuable insights into effector–receptor interactions, paving the way for innovative resistance breeding strategies. We also discuss the latest approaches to engineering durable resistance, including gene stacking, synthetic NLRs, and CRISPR-based modifications. Understanding these molecular mechanisms is critical for developing resistant potato cultivars and mitigating the devastating effects of LB. This review aims to bridge current knowledge gaps and guide future research efforts in plant immunity and disease management. Full article

Given health concerns, oleogels are promising substitutes for saturated fats in food products. An emulsion-templated method was used, employing rapeseed oil and hydroxypropyl methylcellulose (HPMC) as the structuring agent, to produce oleogels. Oil-in-water emulsions (50:50 w/w) were prepared with three HPMC concentrations (1.5, 2.0, and 2.5% w/w) and dried convectively at 60, 70, 80, and 90 °C to obtain oleogels. The emulsions exhibited viscoelastic behaviour with a predominant viscous character, G″ > G′. Drying kinetics showed a constant rate period followed by a falling rate period; the latter was satisfactorily modelled using a diffusion-based approach. All oleogels displayed predominantly elastic behaviour but the characteristics depended on the temperature employed during the drying operation and the HPMC content. The mechanical moduli (G″ and G′) of the oleogels increased significantly with a drying temperature below 80 °C. Higher HPMC content enhanced structural development and thermal stability. Most oleogels exhibited high oil binding capacity (>85%), which increased with the drying temperature and the HPMC content. A correlation was established between the elastic moduli, oil retention, and the hardness of the oleogels. No significant influences of the drying temperature and the polymer concentration on lipid oxidation and colour samples were determined. These results highlight the importance of selecting appropriate drying conditions based on the desired final product properties. Full article

This study systematically explored the interaction mechanisms between two KRAS G12C inhibitors (Sotorasib and Adagrasib) and human serum albumin (HSA) via UV-vis spectroscopy, fluorescence spectroscopy, three-dimensional fluorescence spectroscopy, and molecular docking methods. The experimental findings demonstrated that both drugs caused static quenching of HSA fluorescence, with binding constants of 13.64 × 10³ M⁻¹ (Sotorasib) and 63.67 × 10³ M⁻¹ (Adagrasib), demonstrating significant selectivity differences in their binding affinities. UV spectral analysis demonstrated distinct microenvironmental perturbations: Sotorasib and Adagrasib induced a shift (∆λ = 7 nm and ∆λ = 8 nm, respectively) at 211 nm, consistent with altered polarity in HSA’s binding pockets. Fluorescence spectroscopy confirmed a 1:1 binding stoichiometry, with Stern-Volmer analysis validating static quenching as the dominant mechanism. Three-dimensional fluorescence spectra further highlighted Adagrasib’s stronger conformational impact, reducing tyrosine and tryptophan residue fluorescence intensities by 16% (Peak 1) and 10% (Peak 2), respectively, compared to Sotorasib. Molecular docking revealed divergent binding modes: Sotorasib occupied Sudlow Site I via three hydrogen bonds and hydrophobic interactions (∆G = −24.60 kJ·mol⁻¹), whereas Adagrasib bound through one hydrogen bond and hydrophobic forces (∆G = −30.92 kJ·mol⁻¹), with stability differences attributed to structural characteristics. This study uses multispectral technology and molecular docking to reveal the binding mechanism of Sotorasib and Adagrasib with HSA, providing a theoretical basis for designing highly targeted albumin nanocarriers. The strong binding properties of Adagrasib and HSA may reduce the toxicity of free drugs, providing direction for the development of long-acting formulations. Full article

Goethite, a ubiquitous Fe(III) oxyhydroxide mineral, typically occurs in very small particle sizes whose interfacial properties critically influence the fate and transport of ionic species in natural systems. The surface site density of synthetic goethite increases with particle size, resulting in enhanced adsorption capacity per unit area. In the first two parts of this study, we modeled the adsorption of protons, nitrate, As(V), Pb(II), Zn(II), and phosphate on goethite as a function of particle size, adsorbate concentration, pH, and ionic strength, using unified parameters within the CD-MUSIC framework. Here, we extend this work to characterize the interfacial behavior of carbonate in goethite suspensions, using a comprehensive dataset generated previously under both closed and open CO₂ system conditions. Carbonate oxyanions, prevalent in geochemical environments, exhibit competitive and complexation interactions with other ions and mineral surfaces. Although a bidentate bridging surface carbonate complex has been successful in previous modeling efforts on goethite, we found that the size of the carbonate moiety is too small and would require extreme octahedron bending of the goethite’s singly coordinated sites to accommodate this type of binding. Here, we propose a novel complex configuration that considers structural, physicochemical, and spectroscopic evidence. Optimal unified affinity constants and charge distribution parameters for this complex simulated all experimental data successfully, providing further validation of the CD-MUSIC model for describing relevant goethite/aqueous interfacial reactions. Full article

Background/Objectives: Monoclonal antibodies targeting the PD-1/PD-L1 immune checkpoint have achieved clinical success but face drawbacks such as poor oral bioavailability, limited tumor penetration, and immune-related adverse events. Small-molecule inhibitors present a promising alternative that may overcome these challenges. Methods: Here, an integrated computational framework combining ligand-based pharmacophore modeling and structure-based molecular docking was utilized to screen a comprehensive library consisting of traditional Chinese medicine-derived compounds and clinically approved drugs. The binding affinity between identified candidate compounds and PD-L1 was quantitatively assessed using bio-layer interferometry (BLI). In vitro cytotoxicity assays were conducted on A549 human lung carcinoma and LLC mouse lung carcinoma cell lines. In vivo antitumor efficacy was evaluated in LLC tumor-bearing mice through measurement of tumor growth inhibition, serum cytokine levels (IFN-γ and IL-4) by ELISA, and expression levels of IFN-γ and granzyme B (GZMB) within tumor tissues via immunohistochemistry. Results: In vitro, anidulafungin exhibited anti-tumor effects against both human lung cancer A549 cells and mouse Lewis lung carcinoma (LLC) tumor cells, with IC₅₀ values of 170.6 µg/mL and 160.9 µg/mL, respectively. The BLI analysis revealed a dissociation constant (K_D) of 76.9 μM, indicating a high affinity of anidulafungin for PD-L1. In vivo, anidulafungin significantly increased serum levels of IFN-γ and IL-4 in tumor-bearing mice and elevated expression of IFN-γ and granzyme B (GZMB) in tumor tissues, confirming its immune-mediated anti-tumor effects. Conclusions: Anidulafungin represents a promising small-molecule PD-L1 inhibitor, demonstrating significant anti-tumor potential via immune activation and highlighting the feasibility of repurposing approved drugs for cancer immunotherapy. Full article

Most astrophysical systems (except for very compact objects such as, e.g., black holes and neutron stars) in our Universe are characterized by shallow gravitational potentials, with dimensionless compactness $|\Phi |\equiv r_s/R\ll 1$, where $r_s$ and R are their Schwarzschild radius and typical size, respectively. While the existence and characteristic scales of such virialized systems depend on gravity, we demonstrate that the value of $|\Phi |$—and thus the non-relativistic nature of most astrophysical objects—arises from microphysical parameters, specifically the fine structure constant and the electron-to-proton mass ratio, and is fundamentally independent of the gravitational constant, G. In fact, the (generally extensive) gravitational potential becomes ‘locally’ intensive at the system boundary; the compactness parameter corresponds to the binding energy (or degeneracy energy, in the case of quantum degeneracy pressure-supported systems) per proton, representing the amount of work that needs to be done in order to allow proton extraction from the system. More generally, extensive properties of gravitating systems depend on G, whereas intensive properties do not. It then follows that peak rms values of large-scale astrophysical velocities and escape velocities associated with naturally formed astrophysical systems are determined by electromagnetic and atomic physics, not by gravitation, and that the compactness, $|\Phi |$, is always set by microphysical scales—even for the most compact objects, such as neutron stars, where $|\Phi |$ is determined by quantities like the pion-to-proton mass ratio. This observation, largely overlooked in the literature, explains why the Universe is not dominated by relativistic, compact objects and connects the relatively low entropy of the observable Universe to underlying basic microphysics. Our results emphasize the central but underappreciated role played by dimensionless microphysical constants in shaping the macroscopic gravitational landscape of the Universe. In particular, we clarify that this independence of the compactness, $|\Phi |$, from G applies specifically to entire, virialized, or degeneracy pressure-supported systems, naturally formed astrophysical systems—such as stars, galaxies, and planets—that have reached equilibrium between self-gravity and microphysical processes. In contrast, arbitrary subsystems (e.g., a piece cut from a planet) do not exhibit this property; well within/outside the gravitating object, the rms velocity is suppressed and G reappears. Finally, we point out that a clear distinction between intensive and extensive astrophysical/cosmological properties could potentially shed new light on the mass hierarchy and the cosmological constant problems; both may be related to the large complexity of our Universe. Full article

The SARS-CoV-2 spike (S1) protein facilitates viral entry through binding to angiotensin-converting enzyme 2 (ACE2), but it also contains an Arg–Gly–Asp (RGD) motif that may enable interactions with RGD-binding integrins on ACE2-negative cells. Here, we provide quantitative evidence for this alternative binding pathway using a live-cell, label-free resonant waveguide grating (RWG) biosensor. RWG technology allowed us to monitor real-time adhesion kinetics of live cells to RGD-displaying substrates, as well as cell adhesion to S1-coated surfaces. To characterize the strength of the integrin–S1 interaction, we determined the dissociation constant using two complementary approaches. First, we performed a live-cell competitive binding assay on RGD-displaying surfaces, where varying concentrations of soluble S1 were added to cell suspensions. Second, we recorded the adhesion kinetics of cells on S1-coated surfaces and fitted the data using a kinetic model based on coupled ordinary differential equations. By comparing the results from both methods, we estimate that approximately 33% of the S1 molecules immobilized on the Nb₂O₅ biosensor surface are capable of initiating integrin-mediated adhesion. These findings support the existence of an alternative integrin-dependent entry route for SARS-CoV-2 and highlight the effectiveness of label-free RWG biosensing for quantitatively probing virus–host interactions under physiologically relevant conditions without the need of the isolation of the interaction partners from the cells. Full article

Background: When an antigen molecule is exposed to serum, many different kinds of antibodies bind to it. The complexity of these binding events is only poorly characterized by assays that generate a single variable, generally reflecting the fractional saturation of the antigen, as the readout. Methods: We have previously devised an assay that delivers the essential biochemical variables to determine fractional saturation as the output: an equilibrium dissociation constant for affinity, the ratio of antibody concentration to the equilibrium constant and the concentration of bound antibodies under reference conditions. Here we propose a visualization method for the practical and informative display of these variables. Results: Using total antigen concentration and free and bound antibody concentration as coordinates in a three-dimensional space, a surface plot can depict the behavior of serum antibodies in the measurement range and identify the values of the key variables of binding activity. This surface display (antibody binding in 3-concentration display, Ab3cD) was used for the characterization of antibody binding to the SARS-CoV-2 spike protein in seronegative and seropositive sera. We demonstrate that this visualization scheme is suitable for presenting both individual and group differences and that epitope density changes, not commonly measured by immunoassays, are also revealed by the method. Conclusions: We recommend the use of 3D visualization whenever detailed, informative and characteristic differences in serum antibody reactivity are studied. Full article

Objective: The development of natural and new P-gp modulators to reverse tumor multidrug resistance (MDR). Methods: Test compounds were prepared from the plant Metaplexis japonica, and their ability to reverse P-glycoprotein (P-gp)-mediated MDR was investigated in HepG2/Dox cells. Their effects on P-gp expression and function and their interaction modes with P-gp were also investigated. Results: Natural product 3β,12β,14β, 17β,20(S)-pentahydroxy-5α-pregnan-12β-O-(E)-cinnamate (1) and its new semisynthetic derivative 3β12β,14β,17β,20(S)-pentahydroxy-5α-pregnan-3β-O-nicotinate-12β-O-(E)-cinnamate (1a) were obtained. At non-cytotoxic concentrations of 5 or 10 μM, they significantly reversed the resistance of HepG2/Dox cells to P-gp substrate drugs doxorubicin, paclitaxel, and vinblastine, with reversal folds of 7.1, 118.5, and 198.3 (1), and 18.8, 335.8, and 140.0 (1a), respectively, at 10 μM. Cell apoptosis and expression of caspase 9 were both triggered by the combination of 10 μM of compound 1 or 1a and 500 nM of paclitaxel (p < 0.001). Compound 1 or 1a did not affect P-gp expression, but it did significantly suppress the efflux of Rhodamine 123 out of HepG2/Dox cells (p < 0.001). On the Caco-2 cell monolayer, 1 and 1a were shown to be non-substrates of P-gp, with efflux ratios of 0.83 and 0.89. Molecular docking revealed their strong binding energies (−8.2 and −8.4 kcal/mol) with P-gp, and their direct binding to P-gp was confirmed by their dissociation constants (5.53 µM for 1 and 3.72 µM for 1a), determined using surface plasmon resonance. Conclusions: Compounds 1 and 1a are potential P-gp modulators; they may reverse P-gp-MDR through interacting with P-gp to interfere with substrate binding and transporting, and have the potential to improve the efficacy of paclitaxel or vinblastine drugs for combating P-gp-mediated MDR in tumor cells. Full article

Background/Objectives: Radiolabeled interleukin-2 (IL2) could allow for imaging activated T-lymphocytes in the tumor microenvironment (TME). The aims of this study were to assess the shelf life of a lyophilized kit containing THP-desIL2 to allow for the labeling of IL2 with ⁶⁸Ga at room temperature and to test the in vitro binding of ⁶⁸Ga-THP-desIL2 on different T-cell populations in order to determine which specific T-cell subset expresses the CD25 subunit of the IL2 receptor (IL2R). Methods: desIL2 was conjugated with THP and lyophilized. ⁶⁸Ga labeling was performed and several quality controls, including HPLC, iTLC and SDS-PAGE, were carried out at different storage times (1, 3 and 6 months) and temperatures (4 °C and −80 °C). Moreover, flow cytometric analysis on different T-cell populations and the in vitro and competitive binding of ⁶⁸Ga-THP-desIL2 were performed. Results: The lyophilized kit of THP-desIL2 was stable up to 6 months at −80 °C, preserving its sterility, integrity and acceptable values of labeling yield (51.80 ± 3.74%), radiochemical purity (>96%) and specific activity (5.59 ± 0.40 MBq/µg). Binding of ⁶⁸Ga-THP-desIL2 on activated lymphocytes was specific and exhibited a low dissociation constant from IL2R on stimulated Tregs (Kd: 10⁻⁹–10⁻¹⁰ mol/L). Conclusions: We assessed the shelf life of a lyophilized kit containing THP-desIL2 for the easy labeling of IL2 with ⁶⁸Ga at room temperature. The kit can be stored at −80 °C up to 6 months, thus facilitating the adoption of ⁶⁸Ga-THP-desIL2 into clinical practice. ⁶⁸Ga-THP-desIL2 showed high affinity and specificity for CD25 on activated T-lymphocytes, particularly Tregs, thus opening new opportunities for imaging immune cells trafficking in the TME. Full article