We report a one-step biosynthesis of silver nanoparticles (Fa-AgNPs) using an aqueous Fagonia indica extract and benchmark their activity at the nano-fungal biointerface against Candida albicans and the emerging, multidrug-resistant pathogen Candida auris. Fa-AgNPs were polydisperse and quasi-spherical, with a smaller mean diameter than citrate-capped chemically synthesized AgNPs (Cs-AgNPs) (19.41 ± 7.32 nm vs. 29.01 ± 2.63 nm). Fa-AgNPs outperformed Cs-AgNPs on both species, with MICs of 31.25 vs 125 µg/mL against C. albicans and 3.91 vs 7.81 µg/mL against azole-resistant C. auris. Against C. auris, Fa-AgNPs were at least 64-fold more potent than fluconazole (3.91 vs >250 µg/mL). To our knowledge, this is the first report of F. indica-mediated AgNPs achieving an MIC below 4 µg/mL against this pathogen. Mechanistically, Fa-AgNPs delayed the onset of growth, elevated intracellular ROS in C. albicans, and induced catalase-based antioxidant defense in both species; SEM showed loss of budding cells and extracellular polymeric substances, together with a transition from smooth to rough surfaces and cell-wall damage, in both species, consistent with a primarily oxidative interfacial mechanism. Hemolysis assays indicated acceptable blood compatibility at the lowest concentration tested (2.5 µg/mL), with hemolysis increasing at higher concentrations.
Pt(IV) prodrugs offer a versatile platform for the design of platinum-based antitumor drugs; yet utilizing axial ligands to actively modulate tumor bioenergetics has been less investigated. Herein, we report a mitochondria-directed Pt(IV) prodrug, LA-Pt(IV), in which lipoic acid (LA) serves as a metabolite-reactive axial ligand enabling selective tumor suppression. Upon cellular internalization, LA-Pt(IV) preferentially accumulates in the mitochondria and undergoes intracellular reduction to release cytotoxic Pt(II) species together with lipoic acid-derived dihydrolipoic acid (DHLA). While the Pt(II) species induce DNA damage, DHLA undergoes the Michael addition reaction with endogenous ubiquinone (UQ), thereby depleting this key electron carrier and disrupting mitochondrial electron transport. This process triggers profound mitochondrial dysfunction, bioenergetic collapse, and a lethal burst of reactive oxygen species (ROS). Notably, LA-Pt(IV) exhibits markedly enhanced potency and selectivity toward multiple cancer cell lines relative to cisplatin. In vivo studies confirmed that LA-Pt(IV) achieves potent tumor inhibition with substantially reduced systemic toxicity. This work establishes a dual-targeting strategy for platinum drug design by incorporating the reactive ligand to perturb endogenous metabolism associated with mitochondrial electron transport.
Chronic hepatitis B (CHB) afflicts approximately 254 million people worldwide and is responsible for more than one million deaths annually. Intrahepatic persistence of covalently closed circular DNA (cccDNA), the episomal transcriptional template of hepatitis B virus (HBV), sustains chronic replication, blunts the efficacy of antiviral therapy, and precipitates virological relapse once treatment is discontinued. While emerging cccDNA-targeted strategies include CRISPR/Cas9-mediated cleavage and epigenetic silencing, their clinical translation is impeded by delivery inefficiency, safety concerns, and limited durability. Herein, we present a sequential nano-therapeutic platform that combines small interfering RNA (siRNA) delivery using HBV-pre-activated macrophage membranes (HMs) with photothermal therapy utilizing aggregation-induced emission (AIE) agents. In the first step, HM-coated lipid nanoparticles encapsulating siRNA (siR@HM NPs) enable virus to neutralize and hepatocyte targeted delivery via surface viral receptors, achieving efficient gene silencing. At 72 h post-administration, HM-coated PLGA nanoparticles loaded with an AIE photothermal agent (TPAT@HM NPs) are introduced, facilitating near-infrared IIb fluorescence-guided 808 nm laser ablation of residual infected cells. In a cell model of chronic HBV infection, both siR@HM NPs and TPAT@HM NPs achieved targeted delivery to infected hepatocytes. Critically, sequential administration yielded superior antiviral efficacy compared with concurrent administration or either monotherapy, leading to substantial reduction in HBsAg, HBeAg, HBV DNA, cccDNA, and pgRNA levels. Biosafety assessments confirmed negligible cytotoxicity, hemolysis, and systemic toxicity. This bionic platform integrates time-programmed virus targeting, RNA interference, and photothermal ablation. The virus-preactivated membrane coating enables dual recognition of both viral particles and infected cells, offering a precise and scalable theranostic strategy for HBV and other refractory viral infections.
Oxidative stress-induced disruption of neuronal signaling remains a central barrier in the treatment of neurological disorders, where excessive reactive oxygen species (ROS) compromise ion channel function, synaptic transmission, and neuronal differentiation. Although conductive nanomaterials such as graphene oxide-silver nanoparticles (GO-AgNPs) have shown promise in promoting neurite outgrowth and neuronal maturation, their application is limited by structural instability and burst silver ion (Ag+) release under oxidative conditions. Here, we introduce a carbon dot (CD)-enabled strategy to stabilize conductive GO-Ag nanocomposites via green, in situ integration of folic acid-derived nitrogen-doped CDs. This design transforms conventional GO-AgNPs into a redox-active, self-regulating conductive platform, GO-CD-AgNPs (GC-AgNPs), that simultaneously enhances electrical stability and mitigates oxidative damage. GC-AgNPs exhibit sustained electrical conductivity under oxidative stress and markedly enhanced radical scavenging capacity. In SH-SY5Y cells, GC-AgNPs significantly reduce cytotoxicity by preventing superoxide overproduction and intracellular Ca2+ overload. Under oxidative stress, these nanocomposites promote neuronal maturation, as evidenced by increased expression of MAP2 and NF-H. In addition, GC-AgNPs suppress pro-inflammatory cytokine secretion in HMC3 microglia, demonstrating concurrent immunomodulatory effects. Collectively, by resolving the intrinsic coupling between conductivity and oxidative stability, this work establishes a design framework for redox-adaptive conductive biomaterials. This strategy provides a promising approach for designing electroactive biomaterials capable of maintaining neural network integrity in challenging pathological microenvironments.
Molecular orientation and structural topology play critical roles in determining the performance of biomacromolecular nanocarriers for controlled drug delivery. Herein, the influence of organic and silsesquioxane crosslinker topologies on hybrid nanogels composed of an allyl-functionalized silk fibroin macromer (SFM) and poly(lactic acid) macromer (PLAM) was systematically investigated. Three crosslinkers with distinct molecular architectures were employed: a linear organic framework (N,N'-methylenebisacrylamide, MBAm), a ladder-like silsesquioxane [LD-4Ph-2MeVi, (LDSQ)], and a well-defined double-decker silsesquioxane [(DDSQ-2MeVi, (DDSQ)]. Crosslinker topology markedly influenced nanogel morphology and surface characteristics, producing average particle diameters of approximately 292 nm [NG1(MBAm)], 688 nm [NG2(LDSQ)], and 315 nm [NG3(DDSQ)], while silsesquioxane-containing nanogels exhibited negative zeta potentials (-10 to -30 mV). High-resolution XPS, HRTEM, and XRD analyses revealed that the rigid-cage-like DDSQ topology promotes localized molecular ordering and more compact chain packing of β-sheet-rich SFM and PLAM chains within the hybrid nanogel network. These structural differences significantly affected pH-responsive biphasic Rhodamine B release. At physiological pH 7.4, electrostatic interactions between the nanogel matrix and cargo molecules minimized premature release, whereas acidic conditions (pH 5.5) induced protonation-mediated swelling and accelerated molecular diffusion, resulting in accelerated release. Among all formulations, NG3(DDSQ) exhibited the highest cumulative release efficiency, which was attributed to its ordered network structure and stable nanochannel-like free volume. Furthermore, all nanogels demonstrated excellent cytocompatibility (>98% viability) and significantly promoted fibroblast proliferation (p < 0.05). These findings establish a clear topology-structure-property relationship in silk fibroin/PLA hybrid nanogels and demonstrate that silsesquioxane topology is an effective molecular design parameter for developing pH-responsive biomacromolecular drug delivery systems.
Hexagonal boron nitride nanoplatelets (BNNPs) are promising nanofillers for Mg matrix composites owing to their excellent mechanical properties and low density. In this research, we report the fabrication of ZK61 magnesium composites reinforced with a hybrid of BNNPs and β-tricalcium phosphate (TCP) using spark plasma sintering (SPS). The designed composites exhibit a dense microstructure characterized by refined α-Mg grains with β-TCP particles preferentially distributed along the α-Mg particle boundaries. In addition, trace amounts of interfacial products, such as Mg3N2 and MgB2, are detected to act as chemical anchoring phases, promoting stronger interfacial bonding. Notably, the composite containing 5.0 wt % β-TCP and 0.3 wt % BNNPs demonstrates a remarkable combination of strength and ductility, exhibiting a compressive yield strength of ∼132 MPa, an ultimate compressive strength of ∼380 MPa, and an elongation of ∼28.9%. This favorable balance of high strength and substantial plastic deformability highlights the synergistic reinforcing effect of β-TCP and BNNPs in the ZK61 matrix. In addition, immersion tests reveal that the degradation rate of the 0.3BNNP/5TCP/ZK61 composite is approximately one-fourth that of the monolithic ZK61 alloy immersed in SBF for 28 d. These findings suggest that BNNP/β-TCP reinforced ZK61 composites are promising candidates for biodegradable implant materials.
This research article analyzes the state-of-the-art technologies of bio-inspired, next-generation humanoid electronic skin (e-skin) electrochemical aptasensors developed for the detection of alprazolam. Current wearable devices suffer from poor flexibility, low durability, limited sensitivity, and discomfort during long-term use, thereby necessitating the development of sensors for flexible, sensitive, and real-time health monitoring applications. Here, 3D-printed flexible humanoid e-skin (silicon sheet) mimics human skin and holds the potential to replicate realistic skin for wearable and artificial interface systems to achieve noninvasive, real-time monitoring of physiological parameters. The 3D-printed foundation consisted of a polylactic (PLA) filament that is not conductive, offering a high-tech design and support system for the sensitive conductive humanoid e-skin electrode. Chemically synthesized gold nano-bullets (AuNBs) were utilized to raise the sensor's sensitivity by accelerating electron transport. The synthesis of gold nano-bullets (AuNBs) has been confirmed using colorimetric change, UV-vis absorption at 529 nm, FTIR analysis of their functional groups, and transmission electron microscopy and the selected area electron diffraction (TEM-SAED) investigations demonstrating their homogeneously distributed with a mean particle size of 13.379 ± 1.029 nm and a lattice parameter of 4.078 Å, providing a large active surface for aptamer immobilization and sensitive electrochemical detection. Cyclic voltammetry (CV), linear sweep voltammetry (LSV), and electrochemical impedance spectroscopy (EIS) were used for the detection of the drug analyte, i.e., alprazolam (ALP), the estimated limit of detection (LOD) and the linear range were as low as 0.001 and 0.001-10 µg/mL, respectively. The performance of reproducibility (5 times) and low relative standard deviation (RSD) and stability (20 days) were satisfactory. Kinetic parameters of the interaction of the humanoid e-skin aptamer with the analyte were also investigated using current response, effective electrode surface area, charge transfer resistance (Rct), double-layer capacitance (Cdl), and apparent electron transfer rate constant (Kapp). The AuNBs layer improved electron transfer kinetics, whereas aptamer immobilization and subsequent alprazolam binding modified the interfacial electrochemical characteristics, hence validating effective drug analyte identification by the aptasensor. This humanoid e-skin aptasensor showed adequate performance by estimation of cross-reactivity and spiked analyte concentrations in beverages. It is envisaged that humanoid e-skin electrochemical sensors are anticipated to significantly impact future healthcare by providing continuous, non-invasive monitoring and promoting a transition from traditional blood-based diagnostics to decentralized, personalized, and remote diagnostic systems.
Viral oncoprotein HPV16 E6 is a critical mediator of cervical malignancy and a promising biomarker for early-stage detection of cervical cancer. Current diagnostic approaches centered around morphology-based screening and DNA-based assays emphasize the need for noninvasive, affordable, and highly sensitive urine-based detection methods targeting HPV16 E6 oncoprotein for early diagnosis. This study introduces a noninvasive diagnostic strategy using a gelatin-PVA (G-PVA) hydrogel composite-based electrochemical biosensor for HPV16 E6 oncoprotein detection in spiked urine samples. The proposed platform integrates chemically crosslinked G-PVA hydrogel as a transducing matrix for biosensing offering superior biocompatibility, electrochemical stability, and enhanced surface area for antibody immobilization. The biosensor platform demonstrates robust electrochemical performance with a broad dynamic range (1 pg mL-1 to 1 μg mL-1), ultra-low detection limit (0.56 pg mL-1), and quantification limit (1.88 pg mL-1). Furthermore, it exhibits a sensitivity of 37 μA cm-2 log-1 (pg mL-1) along with good reproducibility and specificity toward E6 oncoprotein even in complex media such as artificial urine. Overall, this study demonstrates the feasibility of the G-PVA hydrogel composite-based biosensor as an affordable, noninvasive, point-of-care (POC) detection platform representing significant advancement toward early detection of HPV16-associated cervical cancer.
Neurodegenerative diseases are a group of progressive disorders characterized by the degeneration of neurons with significant cognitive decline and motor and autonomic dysfunction. They are linked to protein aggregation that leads to cellular toxicity, neuronal death, and brain atrophy due to the accumulation of amyloid plaques and tau tangles in the brain. Recent advances in biomaterials have introduced several biocompatible polymers and conjugated biomolecules as potential inhibitors for protein aggregation. Conjugation to biocompatible polyethylene glycol (PEG) is the most widely used strategy for enhancing bioavailability, biocompatibility, and pharmacokinetic properties of small molecular drugs. In this work, we have designed and synthesized two biomolecule-polymer conjugates, namely, Tre-CA-mPEG and LA-CA-mPEG, by combining a biocompatible, nontoxic polymer monomethoxy PEG (mPEG), a steroid bile acid, cholic acid (CA), and a functional biomolecule, trehalose (Tre) or lipoic acid (LA). These building blocks have significant protein stabilization and neuroprotective attributes, which are important for protein aggregation inhibition. The size distributions of the self-assemblies formed by the Tre-CA-mPEG and LA-CA-mPEG bioconjugates were determined by dynamic light scattering (DLS) measurements. Both the PEGylated bioconjugate assemblies show promising results for inhibition of protein aggregation. The hydrophobic interactions provided predominantly by the steroidal core moiety, together with hydrogen bonding interactions within the PEG corona of the self-assemblies, play a crucial role in protein stabilization and fibrillation inhibition. LA-CA-mPEG bioconjugate exhibits superior effects in delaying aggregation kinetics and enhancing the inhibition of protein aggregation compared to the Tre-CA-mPEG bioconjugate. This enhanced protein fibrillation inhibition can be attributed to the larger hydrophobic surface and stronger interactions arising from the LA-CA-mPEG nanoassemblies.
Polymeric nanogels are attractive biomaterial platforms for preparing aqueous dispersions of hydrophobic bioactive compounds, including essential oils. However, incorporating chemically complex, terpene-rich oils into these polymeric systems may modify their supramolecular organization, colloidal behavior, and biological responses. Such physicochemical changes do not, by themselves, demonstrate controlled release or quantitative delivery performance. Here, we developed and characterized a Pluronic F127/Carbopol 974P nanogel incorporating a nominal concentration of 1% (w/w) Duguetia stelechantha essential oil droplets (EODs) and evaluated the developmental safety profile of the complete formulation using zebrafish (Danio rerio) embryos as a vertebrate screening model. Chemical analysis showed that EODs were dominated by sabinene (41.44%), terpinen-4-ol (17.61%), and spathulenol (9.21%). The optimized EOD-incorporated nanogel (nGDs) provided a homogeneous aqueous dispersion of the hydrophobic oil, whereas FTIR, DSC, DLS, SEM, and AFM revealed formulation-dependent differences in spectral behavior, thermal transitions, hydrodynamic parameters, dried-sample morphology, and apparent nanomechanical properties. These differences are compatible with non-covalent association and changes in polymer organization but do not establish molecular localization or a specific self-assembly mechanism. The nominal oil content should not be interpreted as an experimentally determined encapsulation efficiency or analytical loading capacity. Exposure to nGDs at 50-200 μg mL-1 induced concentration-dependent developmental toxicity, including increased mortality, altered hatching, reduced spontaneous tail coiling, cardiac impairment, and morphological abnormalities. In contrast, the oil-free polymeric matrix did not reproduce the toxicity profile observed for nGDs, indicating that the carrier alone did not account for the adverse responses induced by the complete formulation. These findings characterize the developmental safety profile of nGDs as a water-dispersible formulation designed for biological application without the addition of an organic cosolvent or another external dispersing vehicle. Without a free-EOD comparator, the results cannot determine whether formulation modified the intrinsic toxicity of the oil.
Proteases cleave peptide bonds and generate shorter polypeptide products, finding broad applications. For applications in peptide sequencing, protein mapping, and production of bioactive polypeptides, control over where proteolysis initiates and how it proceeds is critical. Natural proteolytic selectivity primarily originates from local recognition of short amino-acid motifs. However, higher-order or regional selectivity (i.e., cleavage biased toward specific termini or domains) remains difficult even with sophisticated protein engineering or complex multi-enzyme, cascade proteolytic design. We recently reported that trypsin@Ca-BPDC preferentially cleaved the N-terminus of T4 phage lysozyme (T4L) [ACS Appl. Mater. Interfaces 2023, 15 (7), 8927-8936], indicating a promising alternative to introduce directional or regional proteolysis through enzyme immobilization via a unique MOF, Ca-BPDC. To confirm such preference, as a follow-up of that work, here, we extend the MOF-based immobilization to two commercial serine proteases (mixture products with unreported compositions). Our time-resolved electron paramagnetic resonance (EPR) shows that, despite differing or weakly defined selectivity in solution, both commercial proteases exhibit a consistent shift toward N-terminal cleavage of T4L upon immobilized in Ca-BPDC through co-crystallization. Control experiments indicate negligible adsorption of the substrate to the Ca-BPDC alone, suggesting that directional selectivity arises from the coupled MOF-protease environment rather than substrate-MOF interactions. These results indicate that Ca-BPDC may be able to impose an additional, material-derived layer of proteolytic selection on commercial serine proteases. Thus, it may offer a practical way for directional or programmable proteolysis without enzyme chemical or genetic modification. Our immobilization platform also offers a reusable matrix compatible with real-time mechanistic analysis. This approach opens opportunities for controlled peptide generation, proteolytic selectivity tuning, and design of programmable proteolytic bioreactors.
Understanding how the components of bioactive scaffolds regulate coordinated inflammatory and regenerative signaling is critical for the development of advanced wound healing materials. Here, zinc metal particle-embedded polycaprolactone (PZ) electrospun nanofiber scaffolds were evaluated as an immunomodulatory platform that directs protein expression to influence the wound healing process. Scaffolds containing 0, 0.5, and 1 wt % Zn were assessed during fibroblast monoculture, macrophage monoculture, and fibroblast-macrophage coculture models under sustained inflammatory stimulation. Zn incorporation enhanced cell viability and metabolic activity while maintaining low cytotoxicity across all culture systems. Fibroblasts cultured on Zn-containing scaffolds exhibited reduced inflammatory signaling, suppressed COX-2 expression, and increased secretion of proangiogenic and pro-proliferative factors such as EGF, bFGF, and VEGF. Macrophages displayed restrained inflammatory activation, accompanied by elevated growth factor output, consistent with a pro-healing phenotype. Coculture studies revealed that Zn scaffolds selectively dampened inflammatory amplification while preserving cooperative angiogenic signaling, evidenced by reduced IL-6 and COX-2 expression. Altogether, these results establish Zn-embedded PCL scaffolds as a tunable platform for directing inflammatory and regenerative signaling relevant to wound healing applications.
Exosomes, as key mediators of intercellular communication, exhibit promising prospects in the biomedical field, particularly in stem cell-based replacement therapy. Current exosome isolation techniques face challenges in achieving batch enrichment and nondestructive release, which severely restrict their applications. Traditional specific enrichment strategies rely primarily on antibodies; however, aptamers can specifically recognize targets and serve as effective alternatives to antibodies, offering advantages such as small molecular size, high structural stability, and low cost, making them more suitable for scalable target isolation. Herein, we established the Graphene Oxide-anchored Releasable Aptamer Scaffold (GRAS) technique targeting CD63, a characteristic surface biomarker of exosomes. Leveraging the high affinity of single-stranded nucleic acids for graphene oxide (GO), along with the specific binding capacity of aptamers, we fabricated aptamer-displayed oligonucleotide scaffolds (Apt-Scaffold) and immobilized them onto GO to prepare Apt-Scaffold@GO. This configuration allows effective presentation of the CD63 aptamer on the GO interface for specific exosome recognition and capture. Different methods were further designed to achieve efficient and controllable exosome release. Our experimental results demonstrated that the GRAS technique outperformed the gold-standard ultracentrifugation (UC) in enriching exosomes from mesenchymal stem cell (MSC) culture medium. This method enables quantitative and specific enrichment of exosomes. With simple operation, low cost, and independence on large sophisticated instruments, it provides a robust technical foundation for advancing exosome-related biomedical applications.
The therapeutic efficacy of growth factor-mediated bone regeneration is frequently compromised by structural destabilization and loss of bioactivity following adsorption onto biomaterial surfaces. Addressing this challenge requires biomaterials capable of not only delivering osteogenic cues but also preserving protein functionality at the nanobiointerface. Herein, a hyaluronic acid-conjugated Zn, Se codoped hydroxyapatite nanobiocomposite (HA-Zn, Se-HAP) was engineered to investigate the molecular determinants governing growth factor stabilization and cellular response. Zn and Se incorporation modulated the apatite lattice and surface reactivity, while hyaluronic acid established a hydrated extracellular matrix-mimetic interface conducive to protein interaction. Using BMP-2 as a model osteogenic growth factor, spectroscopic, calorimetric, and computational analyses revealed a thermodynamically favorable and reversible adsorption process dominated by hydrogen bonding, electrostatic interactions, and desolvation effects, while preserving the native conformational architecture of the protein. Density functional theory (DFT) and molecular docking further elucidated the electronic redistribution, binding orientation, and intermolecular interaction motifs responsible for BMP-2 stabilization. The nanobiocomposite exhibited excellent cytocompatibility toward both L929 fibroblasts and MG-63 osteoblast-like cells, promoting cellular metabolic activity, proliferation, and collagen-rich extracellular matrix formation. The findings presented herein reveal the critical role of nanobiointerface engineering in governing growth factor adsorption, structural preservation, and downstream cellular behavior. By bridging interfacial physicochemistry with biological function, this work provides fundamental mechanistic insight into growth factor-biomaterial interactions and advances the design of bioactive osteoinductive platforms that extend beyond conventional carrier systems toward the active modulation of protein fate and regenerative outcomes.
Bacterial colonization and biofilm formation remain major limitations of airway stenting, contributing to device obstruction and infection-related complications. Here, we translated to silicone tracheal stents a previously developed plasma-assisted micro/nanostructured silver interface designed to provide hybrid anti-biofilm activity through reduced bacterial adhesion and controlled silver release. The coated stents were characterized to confirm the generation of the nanostructured silver coating and were evaluated against Gram-positive and Gram-negative bacteria in vitro, followed by in vivo assessment in a miniature pig airway model. In vitro, the coating reproduced the dual bacteriophobic and silver-mediated antimicrobial behavior previously observed on model silicone surfaces. In vivo, animals implanted with silver-coated stents showed lower endpoint bacterial loads in bronchial washes than those receiving commercial stents (1.0 × 108 vs 1.2 × 109 CFU/mL, p =0.0085). Bacterial adhesion on explanted stents was also markedly lower for coated devices (109 vs 1011 CFU /cm2, 99.8% reduction, p < 0.0001), and scanning electron microscopy showed a thinner and less mature biofilm structure. Potentially pathogenic bacteria, including Klebsiella pneumoniae and Pseudomonas aeruginosa, were more frequently detected on commercial stents, whereas coated stents were mainly associated with the native respiratory microbiota. Overall, these results indicate that nanostructured silver-coated silicone tracheal stents display hybrid anti-biofilm functionality in a challenging in vivo airway setting.
Calcium is a strictly regulated physiological ion whose levels in blood and saliva act as biomarkers for a wide range of ailments, including renal, endocrine, and bone diseases. Current diagnostic approaches mainly rely on centralised laboratory assays that are accurate but slow and depend on infrastructural support. Thus, robust biosensors capable of rapid, selective calcium detection in complex biofluids are needed to enable real-time diagnostics with minimal invasive sampling. In this study, we engineered the soluble fluorescent sensor NCaMP7, a fusion of calmodulin and split neongreen fluorescent proteins, to self-assemble and bioconjugate to biopolymer particles (BPs) serving as robust and low-cost particulate calcium sensors. These particles were produced inside recombinant E. coli through a one-step, cost-effective, and scalable production process. Upon purification, the BPs retained their natural spherical structure, with the fusion proteins retaining their native conformation. Calcium-sensing functionality was achieved only when NCaMP7 was fused to the N-terminus of the biopolymer assembly protein domain (CAM-BP), enabling detection of free calcium in buffered media as well as complex biological matrices such as artificial saliva and plasma. These observations were further corroborated by fluorescence microscopy, which revealed a pronounced increase in fluorescence intensity following calcium addition, along with minimal signal decay over time, indicating low levels of photobleaching. The BP-based biosensor was also found to be moderately thermostable, retaining its calcium detection sensitivity for four weeks at 25 and 37 °C. Overall, these results position CAM-BPs as a robust, scalable, and cost-efficient platform for in vitro calcium sensing with broad application potential.
Fungal infections are majorly biofilm-associated infections characterized by dynamic microenvironmental pH fluctuations. Nystatin's hydrophobicity, poor bioavailability, and susceptibility to degradation restrict its clinical utility. In this study, electrospun pectin-carboxymethyl cellulose (CMC)-polyvinyl alcohol (PVA) nanofibers loaded with nystatin (Nys-NF) were developed as a pH-responsive antifungal biointerface. The optimization of electrospinning parameters enabled the fabrication of uniform, porous nanofibers with average fiber diameters of 193 ± 16 nm for blank nanofibers (NF2) and 264 ± 25 nm for Nys-NF. Comprehensive physicochemical characterization confirmed effective physical crosslinking between polymers and loading of nystatin with an encapsulation efficiency of 60.6%. Nys-NF demonstrated pH-responsive behavior, where acidic conditions (pH 5.5) promoted protonation-induced deswelling and matrix dissolution, driving enhanced nystatin release. Nys-NF displayed significant antifungal and antibiofilm activity against Candida albicans (CA), Candida glabrata (CG), and Cryptococcus neoformans (CN). Morphological analysis revealed disruption in biofilm architecture and loss of extracellular matrix components. Mechanistic assays indicated depletion in ergosterol and cell surface hydrophobicity. Nys-NF demonstrated considerable antioxidant activity and cytocompatibility with HEK-293 cells. Conclusively, the developed pH-adaptive polyelectrolyte-based nanofibrous matrices represented a promising localized antifungal delivery system for combatting resilient mucosal fungal biofilms.
Heavy-atom-free photosensitizers, particularly those containing thiocarbonyl groups, hold significant promise for photodynamic therapy, yet their full potential remains untapped. The unique photophysical and photochemical properties of sulfur-modified carbonyl fluorophores are not fully understood. Additionally, challenges such as limited aqueous solubility, aggregation-induced quenching, and inadequate tumor-targeting efficiency hinder their effectiveness in biomedical applications. In this study, we designed and synthesized two thionated naphthalimides and evaluated their potential for photodynamic therapy. Our results showed that transforming carbonyl groups into thiocarbonyl moieties within the naphthalimide structure caused a significant red shift in the absorption profile. This modification also resulted in increased molar absorptivity and a marked improvement in singlet oxygen generation efficiency. Interestingly, substituting at the imide nitrogen of the naphthalimide core had little effect on absorption characteristics and singlet oxygen generation. In contrast, replacing the dimethylamino group with a triphenylamine moiety at the para position of the naphthalimide led to a hypsochromic shift in the absorption spectrum, along with lower molar absorptivity and reduced singlet oxygen yield. Additionally, we successfully prepared biocompatible MANIH-S nanoparticles (MANIH-S NPs) by encapsulating MANIH-S in DSPE-PEG(2000)-biotin to enhance the system's biomedical potential. Upon red light irradiation, MANIH-S NPs effectively generated reactive oxygen species via both type I and type II photodynamic mechanisms. Additionally, these nanoparticles exhibited strong phototoxic effects against HeLa cells when exposed to red light while showing minimal toxicity in the dark. This work highlights the potential of sulfur-substituted naphthalimide derivatives, combined with DSPE-PEG(2000)-biotin nanoencapsulation, as a versatile platform for advanced photodynamic cancer therapy.
Dopamine and xanthine are essential biomolecules for neurological disorders and oxidative stress, respectively, and their abnormal concentrations are associated with Parkinson's disease, schizophrenia, Alzheimer's disease, and renal dysfunction. To overcome the limitations of invasive conventional diagnostics, a flexible MoB/PVA hydrogel on nickel foam is fabricated for the simultaneous, non-invasive electrochemical detection of dopamine and xanthine in human sweat. It is prepared through selective alkali etching and solvothermal synthesis. Characterization of the optimized MoB-MBene hydrogel (7 wt %), such as FTIR, confirms characteristic Mo-B vibrational bands, while SEM analysis reveals a sponge-like porous morphology favorable for rapid ion transport and enhanced electroactive sites. Using the DPV technique, the MoB/PVA hydrogel/NF sensor, which has a higher active surface area, demonstrates the simultaneous detection of dopamine, with a linear range from 50 nM to 500 μM, detection limit of 0.35 nM, and sensitivity of 158 μA nM-1 cm-2, and xanthine, with a linear range of 100 nM to 1 mM, detection limit of 0.098 nM, and sensitivity of 95.2 μA nM-1 cm-2, with a peak separation (ΔEp) of 285 mV. The superior electrochemical performance is attributed to the synergistic effect arising from abundant active sites in the ion-rich MoB/PVA hydrogel, facilitating higher charge-transfer kinetics and resulting in enhanced peak currents and well-resolved peaks for simultaneous sensing with 99.6-101.3% recovery for dopamine and 99.3-101.4% recovery for xanthine. Repeatability tests (N = 4) yielded dopamine with a 0.6% RSD and xanthine with a 1.93% RSD. The flexible sensor exhibits excellent selectivity and stability for up to 4 weeks and holds potential for point-of-care and flexible wearable devices.
Bladder cancer is among the most common malignancies of the urinary system and is characterized by high recurrence rates and unsatisfactory clinical outcomes. Current therapeutic strategies are limited by inadequate local efficacy, rapid drug loss caused by the bladder microenvironment, and systemic toxicity associated with conventional chemotherapy. In this study, we developed a localized therapeutic platform for bladder cancer based on a hyaluronic acid-phenylboronic acid/poly(vinyl alcohol) (HA-PBA/PVA) dual-network hydrogel incorporating liposomes co-loaded with resveratrol (Res) and gemcitabine (Gem). The hydrogel system was designed to provide sustained local drug delivery and improved intravesical retention. Physicochemical characterization demonstrated favorable injectability, self-healing capacity, mechanical stability, and pH-responsive sustained-release behavior. The embedded liposomes exhibited a uniform nanoscale size distribution and high drug encapsulation efficiency. In vitro studies showed that the composite hydrogel displayed good biocompatibility and significantly inhibited MB49 bladder cancer cell proliferation, induced apoptosis, and suppressed migration and invasion. In vivo evaluation further demonstrated favorable biosafety and enhanced antitumor efficacy of the hydrogel-based local delivery system. These findings indicate that the HA-PBA/PVA hydrogel-liposome composite system is a promising localized therapeutic strategy for bladder cancer.
