Search Results (339)

Intervertebral disc degeneration (IVDD) is the leading cause of low back pain, a global public health burden for which current pharmacological and surgical treatments provide symptomatic relief but fail to reverse the underlying degenerative process. The uniquely avascular, hypoxic, acidic, and mechanically demanding disc microenvironment poses formidable barriers to the survival and function of therapeutic cells and genes, emphasizing the critical need for bioengineered delivery systems. In this review, we introduce the structure and microenvironment of the intervertebral disc, as well as the molecular mechanisms underlying IVDD. We then provide a critical comparative analysis of delivery platforms, including hydrogels, microspheres, nanoparticles, nanofibrous scaffolds, and viral and non-viral vectors, around five core delivery challenges: mechanical protection, retention and leakage prevention, targeted intracellular delivery, controlled release kinetics, and metabolic support. Furthermore, we examine the fabrication technologies and material considerations that determine platform performance, and we analyze the translational barriers that have impeded clinical adoption, such as the limitations of small-animal models and unresolved cell leakage. Finally, we highlight emerging strategies, including gene-cell combination therapy and endplate preconditioning, to accelerate the clinical translation of precision therapies for IVDD. Full article

Background/Objectives: Boron neutron capture therapy (BNCT) relies on the selective delivery of boron-10 to tumor cells. Although 4-[¹⁰B]borono-L-phenylalanine (BPA) is currently the only clinically approved BNCT agent, it is limited by poor L-type amino acid transporter 1 (LAT1)/LAT2 selectivity and aqueous solubility. We previously developed 3-borono-5-fluoro-α-methyl-L-phenylalanine (5F-αMe-3BPA), a novel BPA derivative designed to be a LAT1-targeted BNCT/positron emission tomography theranostic agent. This study comprehensively characterizes its pharmacological profile and explores its pharmacokinetic optimization by modulating renal organic anion transporter 1 (OAT1). Methods: Transport kinetics of BPA, related analogs, and 5F-αMe-3BPA were analyzed in HEK293 cells stably expressing LAT1 or LAT2 using Michaelis–Menten analysis. Time-dependent cellular uptake and intracellular retention of BPA and 5F-αMe-3BPA were evaluated in T3M-4 pancreatic cancer cells with or without the LAT1 inhibitor JPH203. In vivo biodistribution was examined in T3M-4 tumor-bearing mice after intravenous administration of 5F-αMe-3BPA or BPA, with assessment of probenecid pretreatment. Results: 5F-αMe-3BPA retained LAT1 affinity comparable to that of BPA while showing markedly reduced LAT2-mediated transport, indicating improved LAT1/LAT2 selectivity. In T3M-4 cells, 5F-αMe-3BPA showed stronger LAT1 dependence, higher steady-state accumulation, and better intracellular retention than BPA under amino acid-containing conditions. Although 5F-αMe-3BPA achieved favorable tumor-to-plasma and tumor-to-muscle ratios in vivo, it was rapidly cleared from circulation. Probenecid pretreatment increased plasma exposure, reduced early renal accumulation, and significantly enhanced tumor boron accumulation, reaching approximately twofold higher levels than control. Conclusions: These findings establish 5F-αMe-3BPA as a highly LAT1-selective BNCT candidate and identify probenecid pretreatment as a clinically translatable pharmacokinetic strategy for maximizing therapeutic boron delivery. Full article

Feline infectious peritonitis virus (FIPV) remains one of the most challenging viral diseases in veterinary medicine, largely owing to the absence of a consistently effective and safe vaccine. Despite widespread feline coronavirus infection, only a subset of infected cats progresses to feline infectious peritonitis, indicating that host immune responses are key determinants of disease outcomes. Accumulating evidence indicates that disease severity is driven not only by viral replication but also by macrophage- and monocyte-centered immune signaling, leading to excessive inflammation and systemic immunopathology in the host. Previous vaccine approaches against FIPV have failed to provide consistent protection and, in some cases, have been associated with enhanced disease. These outcomes suggest that vaccine-induced immune responses that recapitulate pathogenic signaling patterns may exacerbate disease rather than confer protection. In this review, we discuss the current knowledge of immune cell signaling pathways implicated in FIPV infection, including innate sensing through Toll-like receptors, downstream mitogen-activated protein kinases and NF-κB signaling, cytokine production profiles, Fc receptor-associated processes, and intracellular pathways such as autophagy, and how these mechanisms shape vaccine-induced immunity. By integrating insights from immune signaling kinetics, antibody functionality, adjuvant-driven pathway engagement, and platform-specific immune signatures, this review emphasizes the need to reframe FIPV vaccine development strategies that actively shape host immune responses. Rather than maximizing immunogenicity, successful vaccine design is likely to depend on limiting sustained macrophage activation and pro-inflammatory cytokine amplification while supporting antiviral immune functions, thereby reducing the risk of antibody-dependent enhancement and immunopathology. Beyond feline diseases, these considerations provide broader lessons for vaccine design in settings where immune-mediated pathology contributes to disease severity. Full article

The limited regenerative capacity of articular cartilage (AC) following injury has led to a high prevalence of degenerative AC-related disorders, including osteoarthritis (OA). Current clinical treatments for OA have failed to halt disease progression, driving growing interest in cartilage tissue engineering (CTE) strategies aimed at developing biomimetic substitutes to regenerate damaged AC tissue. Among the available biofabrication techniques, electrospinning has gained attention due to its ability to generate fibrous scaffolds that closely mimic the architecture of the native AC extracellular matrix, while also serving as versatile drug delivery platforms with high surface area and elevated drug loading efficiency. Small molecules, low-molecular-weight therapeutic agents capable of interacting with both cell membrane and intracellular components, can be incorporated into these scaffold systems to target the underlying mechanisms of OA. This review examines the current state of the art of small molecule-loaded electrospun scaffolds for CTE applications. Small molecules targeting pain, inflammation, and cartilage function restoration show considerable therapeutic potential, and their incorporation into coaxial and other advanced electrospinning setups enables controlled and sustained drug release. Recent examples of small molecule-loaded electrospun scaffolds for AC repair demonstrate enhanced chondrogenic differentiation and neo-cartilage formation, supporting their potential as viable CTE strategies. Nevertheless, challenges related to drug release kinetics, scaffold load-bearing properties, manufacturing scalability, reproducibility, and regulatory approval remain critical barriers to clinical translation. Emerging fabrication strategies, AI-assisted optimization, personalized medicine approaches, and stimuli-responsive drug delivery systems offer promising avenues to overcome these limitations and advance the clinical adoption of these platforms. Full article

Poly(amidoamine) (PAMAM) dendrimers are widely recognized as versatile nanocarriers due to their tunable architecture and ability to associate with bioactive molecules. In this study, generation 3 PAMAM dendrimers functionalized with triphenylphosphonium (TPP) were employed to investigate the association of structurally related phenolic compounds—caffeic acid, p-coumaric acid, and cinnamic acid—through either covalent conjugation or non-covalent encapsulation. Physicochemical characterization by NMR, dynamic light scattering, and zeta potential measurements revealed the formation of supramolecular aggregates rather than isolated dendrimer units, with hydrodynamic diameters ranging from 127 to 260 nm and positive surface charge across all formulations. Encapsulation efficiencies determined by HPLC reached 93.8% for caffeic acid, 78.9% for p-coumaric acid, and 71% for cinnamic acid, indicating differential association behavior. Molecular dynamics simulations over 1 μs supported these findings, showing stronger and more stable interactions for polar antioxidants, particularly caffeic acid, driven by hydrogen bonding and electrostatic interactions, while cinnamic acid displayed preferential binding in more hydrophobic dendrimer regions. Radical scavenging assays (DPPH• and ABTS•+) demonstrated that all formulations retained antioxidant capacity, although dendrimer association modulated scavenging kinetics. In cellular assays under oxidative stress, free caffeic acid exhibited the strongest immediate reduction of intracellular reactive oxygen species, whereas dendrimer-associated systems showed reduced but significant activity, consistent with decreased solvent accessibility and slower release predicted by simulations. Overall, these results highlight a trade-off between molecular retention and immediate biological efficacy, demonstrating that the mode of association governs antioxidant accessibility and performance. This combined experimental and computational approach provides a mechanistic framework for the rational design of dendrimer-based delivery systems aimed at balancing stability and functional activity. Full article

The Mycobacterium tuberculosis bacterium encodes two active potassium (K⁺)-uptake transport systems, the Trk and the Kdp. The Trk is the low-affinity K⁺ transporter, consisting of two TrkA proteins, while the Kdp consists of the high-affinity K⁺ transporter KdpFABC and the two-component system KdpDE. Both transporters are utilised by the bacteria for growth and survival. During growth, the bacteria utilise the constitutively expressed Trk and suppress the Kdp, but upregulate both transporters during survival. In the current study, we investigated the interactive effects of these systems on bacterial growth and survival. This was achieved by first constructing a M. tuberculosis mutant strain in which both the Trk and Kdp systems were inactivated by homologous recombination. The mutant was evaluated for its growth kinetics in planktonic cultures, as well as survival in biofilm and macrophage cultures. The constructed M. tuberculosis mutant showed faster growth rates in planktonic cultures, but was attenuated for both biofilm formation and intracellular survival in isolated human monocyte-derived macrophages. These results illustrate that both K⁺-uptake systems are essential to sustain slow rates of bacterial growth, as well as for bacterial persistence in hostile environments via optimisation of biofilm formation, and intracellular survival in macrophages. (Words: 194) Full article

Background/Objectives: Multidrug resistance (MDR) remains a major pathophysiological barrier to effective chemotherapy based on anthracyclines, including daunorubicin (DNR), in the treatment of leukemia. However, conventional population-level measurements of drug uptake do not resolve variability in uptake kinetics among individual leukemia cells, which may influence intracellular drug accumulation and therapeutic response. Methods: In this study, real-time DNR uptake was quantified at the single-cell level using a microfluidic biochip that enabled long-term cellular retention and continuous monitoring. Both wild-type drug-sensitive leukemia cells and a multidrug-resistant mutant overexpressing the P-glycoprotein (P-gp) efflux pump were examined. Results: Kinetic analysis revealed that DNR uptake in drug-sensitive cells was well described by a single dominant uptake process, whereas uptake in MDR cells required a model incorporating two kinetically distinct processes. In both cell populations, pronounced cell-to-cell variation was observed in uptake rates and intracellular drug retention, indicating substantial functional heterogeneity within phenotypically similar cells. This variability persisted following the treatment with an MDR inhibitor and obscured the differences between inhibitor-treated and untreated cells when the uptake was compared across different single cells. To overcome this limitation, a same-single-cell analysis (SASCA) approach was employed, enabling direct comparison of DNR uptake in the same individual cell before and after inhibitor exposure, thereby revealing enhanced intracellular DNR retention and accelerated uptake kinetics following inhibition. Conclusions: Together, these results demonstrate that real-time single-cell kinetic analysis reveals functionally relevant heterogeneity in multidrug-resistant leukemia cells and provides insight into the pathophysiology of MDR that cannot be obtained from population-averaged measurements. Full article

Bioactive glass (BAG) S53P4 is a clinically approved bone substitute with antibacterial, osteoconductive and osteostimulatory properties. Its antibacterial effect is associated with ion release, local pH elevation and osmolality, but the precise biochemical and biophysical mode-of-action is unclear. This study investigates the antibacterial mechanism of BAG S53P4 eluates. BAG eluates, collected at 2, 4, 8, and 24 h, eradicated Staphylococcus aureus. Elemental analysis revealed an early increase in concentrations of Si and Na, a later rise in Ca, depletion of P over time and rapid loss of Mg. Membrane disturbances occurred within 5 min, evident by permeability for SYTOX, aligning with time-kill kinetics for S. aureus and Bacillus subtilis. In B. subtilis, 2h-BAG-eluate induced rapid delocalization of marker proteins for cell division and DNA repair, signaling membrane potential collapse and nucleoid condensation. Transcriptomics revealed early transcription remodeling reflecting ionic and energetic imbalance, including disruption of central metabolism, redox homeostasis, and translational stability. Scanning electron microscopy revealed severe cell surface damage and particulate deposits on S. aureus. Transmission electron microscopy showed cell envelop disruptions and cytoplasmic leakage. Energy dispersive X-ray analysis identified Si on bacterial cell surface at 4 h and intracellular accumulation in punctured, empty cells at 24 h. Overall, BAG ionic dissolution products kill bacteria through a stepwise mechanism involving membrane damage, protein delocalization and metabolic impairment, accompanied by Si deposition on bacterial surfaces and loss of Mg. This finally leads to cell wall degradation, cytoplasmic content leakage and further Si deposition on the cells and inside cell ghosts. Full article

Background/Objectives: Chronic metabolic acidosis (CMA) is a mild, persistent acid–base imbalance characterized by low serum bicarbonate and urinary pH and is common in chronic illness, aging, and metabolic disorders such as type 2 diabetes (T2D). This review highlights the critical, yet often overlooked, role of CMA in T2D (CMAD) and its contribution to disease pathophysiology. Methods: We conducted a comprehensive review of the systemic impacts of CMA, from molecular mechanisms to organ-specific dysfunction. The analysis covers physiological pH dynamics in intracellular (IC) and extracellular (EC) fluids and explores their effects on cellular processes, including the cell cycle and apoptosis. Results: At the molecular level, acidosis significantly alters enzyme kinetics, macromolecule metabolism, and ion conductance. Cell-level analysis shows that pH shifts impact proliferation and programmed cell death. Systemically, the manifestations of CMA align closely with T2D features in vital organs, including the pancreas, liver, skeletal muscle, adipose tissue, and the renal, nervous, and immune systems. Our findings indicate that the pathophysiological landscape of T2D largely mirrors the biological effects of chronic acidosis. Conclusions: The alignment between the effects of CMA and the clinical features of T2D suggests that T2D pathophysiology is worth revisiting through the lens of CMAD. This perspective is further supported by therapeutic interventions showing preliminary efficacy signals in limited studies of acid-neutralization in managing T2D symptoms and progression. Full article

Postbiotics derived from Bacillus species are recognized as promising natural antimicrobial agents. This study aimed to systematically evaluate the inhibitory activity of postbiotics derived from B. velezensis 906 against L. monocytogenes, elucidate the underlying antibacterial mechanisms using agar diffusion assays, broth microdilution, growth kinetics, flow cytometry, phospholipid competition assays, whole-genome mining, and non-targeted metabolomics, and characterize the bioactive metabolites responsible for their antibacterial effects. The postbiotics exhibited significant antagonistic activity against Gram-positive bacteria, Gram-negative bacteria, and fungi. They also inhibited pathogens such as Salmonella and Enterobacter sakazakii. Against L. monocytogenes, the minimum inhibitory concentration was 0.0083 mg/mL. At 1× MIC, the OD600 after 24 h remained at approximately 0.8, compared with 1.3–1.4 in the untreated control, whereas treatment at 4× MIC almost completely inhibited bacterial growth. Mechanistic analyses suggested that the postbiotics interact with membrane phospholipids, resulting in membrane disruption, increased intracellular reactive oxygen species accumulation, and enhanced membrane permeability. Integrated genome mining and non-targeted metabolomics indicated that the antibacterial activity was associated with a coordinated antimicrobial network involving lipopeptides, polyketides, bacteriocin-related compounds, and siderophore-associated metabolites. These findings provide insight into the antibacterial basis of B. velezensis 906 postbiotics and support their potential application in food safety control. Full article

Objectives: Limited intracellular exposure can reduce the in vitro activity of pazopanib (PAZ) and enzalutamide (ENZ). This study developed bovine serum albumin (BSA) particles co-encapsulating PAZ and ENZ (PE-BSA) and evaluated physicochemical properties, release kinetics, 4T1 cellular uptake, and in vitro cytotoxicity versus free drugs and single-drug particles. Methods: Drug-loaded BSA particles were prepared using a crosslinking-based method. Particle size (PS), polydispersity index (PDI), zeta potential (ZP), and encapsulation efficiency (EE) were determined. In vitro release was assessed over 48 h and fitted to kinetic models. 4T1 uptake was quantified after 2 and 4 h by intracellular drug levels. Cytotoxicity was measured by MTT at 24 and 72 h (1–100 µg/mL). Moreover, cell death analyses were conducted. Stability studies at +4 °C and serum were also carried out. Results: PE-BSA was nanoscale and monodisperse (PS 128.7 ± 2.6 nm; PDI 0.026 ± 0.01) with ZP −31.65 ± 1.13 mV and high EE (PAZ 98.59 ± 1.78%; ENZ 69.79 ± 0.02%). At 24/48 h, cumulative release from PE-BSA was 11.96/12.31% for PAZ and 52.26/85.95% for ENZ. The release kinetics were best described by the Korsmeyer–Peppas model for PAZ (r² = 0.9578) and the Higuchi model for ENZ (r² = 0.9605), indicating diffusion-controlled release. PE-BSA increased 4T1 uptake versus free drugs (2 h: 10.02% PAZ and 21.9% ENZ; 1.77-fold and 4.15-fold), with sustained enhancement at 4 h (2.2- and 4.69-fold, respectively). After 24 h, PE-BSA induced a markedly higher apoptotic response in 4T1 cells (32.5% early apoptosis and 0.8% late apoptosis/early necrosis) compared with free-PAZ (6.6% early apoptosis) and P-BSA (7.3% early apoptosis). Particles were stable. Conclusions: PE-BSA produced BSA particles with diffusion-governed release and enhanced 4T1 internalization, supporting albumin particles as a delivery platform to increase intracellular exposure of PAZ/ENZ in vitro. Full article

Nanomedicine has advanced rapidly as engineered nanoparticles have become increasingly capable of improving drug stability, targeting, controlled release, and biocompatibility. However, nanoparticle clinical utility relies on both delivery efficiency and how they are metabolized, retained, and cleared. This review examines the major biological pathways governing nanoparticle clearance and discusses how engineering parameters can be tuned to influence bioaccumulation, metabolism, excretion, and therapeutic performance with a wide range of available materials. This article is a narrative review of the recent and foundational literature on medically relevant nanoparticles, including lipid-based, polymeric, biopolymer, inorganic, polylactide, and bile-derived systems. All relevant translational, biochemical, chemical, and clinical literature from PubMed was searched from January 1971 to January 2026 to obtain a representative sample of work before information extraction. Nanoparticle clearance is governed by interconnected molecular and organ-level processes that vary according to composition, size, surface chemistry, and route of administration. Surface modifications with PEGylation, zwitterionic coatings, cholesterol, proteins, or responsive linkers can prolong circulation, alter immune recognition, and direct organ-specific handling. While rapid clearance remains desirable for many systemically acting drugs, prolonged intracellular or intratumoral retention may improve outcomes, particularly in boron neutron capture therapy and other activation-dependent treatments. Nanoparticle clearance should be regarded as a context-dependent design parameter rather than a universal limitation. Rational control of clearance kinetics may improve both safety and therapeutic effectiveness in next-generation engineered drug delivery systems. Full article

Nanomaterials are emerging versatile platforms for therapeutic delivery, as they offer precise control over drug, antioxidant, and genetic payload transport across biological barriers. Inorganic, organic, hybrid, and biomimetic systems are the major classes of nanomaterials, which all have different physicochemical properties such as size, surface charge, and surface functionalization. These properties collectively influence stability, biodistribution, cellular uptake, and release kinetics. Engineering strategies are increasingly using stimuli-responsive designs that are triggered by pH, reactive oxygen species (ROS), and intracellular redox gradients to perform spatially and temporally controlled delivery. Antioxidant and redox-modulating nanocarriers are of great importance as they overcome the limited bioavailability and nonspecific activity of conventional antioxidants by improving stability, targeting oxidative microenvironments, and allowing for regulated release. Improvements in lipid, polymeric, and inorganic nanoplatforms have also developed gene delivery applications, including siRNA, mRNA, and CRISPR/Cas systems, to provide better cytosolic release and precise therapeutics. When diagnostic imaging is integrated with therapy through theranostic nanoparticles, real-time monitoring and personalized intervention are possible. Safety, scalable manufacturing, and regulatory alignment are some challenges that show the need for standardization and translational procedures to utilize the potential of theranostic nanomedicine. Full article

Water-soluble fullerene derivatives such as fullerenol C₆₀(OH)₂₄ are promising candidates for nanomedicine applications, yet their effects on innate immune cells remain poorly characterized. We investigated the interaction of fullerenol with human neutrophils isolated from healthy donors, exposed to concentrations of 0.25–200 μg/mL over 24–72 h. Using multi-parameter flow cytometry, we assessed viability, apoptosis, phagocytic activity, and intracellular reactive oxygen species (ROS) production, complemented by cell-free DPPH radical scavenging assays. Fullerenol was taken up by neutrophils in a concentration- and time-dependent manner. No significant cytotoxicity was observed up to 100 μg/mL, while viability declined at 200 μg/mL. Phagocytosis of opsonized E. coli was preserved at lower concentrations, though a statistically significant negative correlation with fullerenol concentration was detected at higher doses. In cell-free assays, fullerenol scavenged DPPH radicals with an EC₅₀ of 48.90 ± 10.02 μg/mL, exhibiting slower kinetics than Trolox or ascorbic acid. Critically, fullerenol suppressed intracellular ROS production by >33% at 50 μg/mL following PMA stimulation of neutrophils. These findings demonstrate that fullerenol C₆₀(OH)₂₄ combines potent intracellular antioxidant activity with a favorable neutrophil safety profile, supporting its potential application in oxidative stress-related conditions. Full article

For decades, the pharmacological identity of Panax ginseng has been primarily attributed to triterpenoid saponins known as ginsenosides. However, accumulating evidence indicates that ginseng also contains a structurally distinct lipid–protein complex, termed gintonin, enriched in lysophosphatidic acid (LPA) species. Unlike ginsenosides, which predominantly exert modulatory effects on membrane dynamics and intracellular kinase pathways, gintonin directly activates LPA G protein-coupled receptors (GPCRs), thereby inducing rapid phospholipase C (PLC) activation and intracellular Ca²⁺ mobilization. Biochemical analyses have identified major LPA species within the gintonin fraction, including C16:0, C18:0, and C18:1, stabilized within a proteinaceous matrix that may influence receptor engagement kinetics. Pharmacological studies demonstrate that gintonin preferentially activates LPA₁ and LPA₃ receptor subtypes, triggering downstream signaling cascades involving MAPK, PI3K/Akt, and Rho pathways. These receptor-mediated effects occur on a rapid temporal scale, distinguishing gintonin from the slower transcriptional and kinase-modulating actions of ginsenosides. In this review, we synthesize current evidence regarding the chemical architecture, receptor pharmacology, and signaling dynamics of gintonin and propose a dual signaling framework in which steroid-like saponins and lipid GPCR ligands represent complementary molecular axes within P. ginseng. Recognition of this layered signaling organization refines the molecular understanding of ginseng biology and highlights gintonin as a unique plant-derived GPCR ligand system. Full article