Search Results (499)

Background/Objectives: Nanoparticles (NPs) were previously explored as enhancers in electroporation due to their potential to locally amplify electric fields near cell membranes, with gold nanoparticles (AuNPs) in particular showing promise in improving membrane permeability and gene electrotransfer (GET). In this study, we systematically investigated the influence of NP properties—including size, shape, surface functionalization, and material—on electroporation efficacy. Methods: A combined approach using theoretical modeling and experimental validation was employed, encompassing numerical simulations, membrane permeabilization assays, transmission electron microscopy, and GET efficiency measurements. Results: Numerical results revealed that the presence of NPs alters local electric field distributions, but the amplification is highly localized, regardless of NP conductivity or geometry. Experimentally, only two out of six tested NP types produced a statistically significant, yet modest, increase in membrane permeability at one electric field intensity. Similarly, GET improvement was observed with only one NP type, with no dependence on concentration or functionalization. Conclusions: Overall, our findings demonstrate that NPs, under tested conditions, do not substantially enhance cell membrane permeability or GET efficacy. These conclusions are supported by both computational modeling and in vitro experiments. Full article

Pancreatic cancer is among the deadliest malignancies, with limited treatment options and poor prognosis. Novel strategies are therefore urgently needed. Sotorasib, a KRAS G12C-specific inhibitor, offers targeted treatment for a small subset of patients with this mutation. Electrochemotherapy (ECT), which enhances the cytotoxicity of chemotherapeutic agents through electroporation-induced membrane permeabilization, has shown promise in various tumor types, including deep-seated malignancies such as pancreatic cancer. Combining ECT with sotorasib may potentiate antitumor effects in KRAS G12C-mutated pancreatic cancer; however, preclinical data on such combinations are lacking. This proof-of-concept study evaluated the cytotoxic effects of ECT using bleomycin (BLM) or cisplatin (CDDP) in combination with sotorasib in KRAS G12C-mutated MIA PaCa-2 and KRAS G12D-mutated PANC-1 pancreatic cancer cell lines. ECT alone significantly reduced cell viability, particularly in MIA PaCa-2 cells, where electric pulses induced approximately 75% cell death. Combining ECT with sotorasib resulted in an additive effect on KRAS G12C-mutated MIA PaCa-2 cells, though no synergy was observed, likely due to the high intrinsic sensitivity to electric pulses. These results support the potential of combining physical and molecular therapies in a subset of pancreatic cancer patients and lay the groundwork for further in vivo studies to optimize treatment parameters and explore clinical translatability. Full article

Aggregation of tau protein is a hallmark feature of tauopathies such as Alzheimer’s disease. The microtubule-binding domain of tau plays a crucial role in the tau aggregation process. In this study, we investigated the dual effects of membrane interactions of tau_298–317, a fragment peptide from the microtubule-binding domain, on peptide-induced membrane disruption and membrane-mediated peptide self-assembly. Our results show that neither wild-type tau_298–317 nor its P301L or Ser305-phosphorylated mutants aggregate in the presence of zwitterionic POPC vesicles or cause lipid vesicle leakage, indicating weak peptide–membrane interactions. In contrast, tau_298–317 strongly interacts with negatively charged POPG liposomes, leading to a rapid transition of the peptide conformation from random coils to α-helical intermediate conformation upon membrane adsorption, which may further promote peptide self-association to form oligomers and β-sheet-rich fibrillar structures. Tau_298–317-induced rapid POPG membrane leakage indicates a synergistic process of the peptide self-assembly at the membrane interface and the aggregation-induced membrane disruption. Notably, phosphorylation at Ser305 disrupts favorable electrostatic interactions between the peptide and POPG membrane surface, thus preventing peptide aggregation and membrane leakage. In contrast, the P301L mutation significantly enhances membrane-mediated peptide aggregation and peptide-induced membrane disruption, likely due to alleviation of local conformational constraints and enhancement of local hydrophobicity, which facilitates fast conformational conversion to β-sheet structures. These findings provide mechanistic insights into the molecular mechanisms underlying membrane-mediated aggregation of crucial regions of tau and peptide-induced membrane damage, indicating potential strategies to prevent tau aggregation and membrane rupture by targeting critical electrostatic interactions between membranes and key local regions of tau. Full article

Dendrimers, 4-dimethylamino-1,8-naphthalimide (DAB) and its halogenated analog 3-bromo-4-dimethylamino-1,8-naphthalimide (DAB-Br), were evaluated on eukaryotic cells, human HFF-1 fibroblast cells, and five fungal species. Although both dendrimers have demonstrated antibacterial and antiviral potential, thus far, their effects on eukaryotic cells, particularly human and fungal cells, have not been investigated. For this purpose, their cytotoxicity, mechanisms of cellular entry, and antifungal activity were studied. Dynamic light scattering measurements revealed that both dendrimers exhibited positive surface charges (+28 to +35 mV), good colloidal stability, and nanoscale dimensions (117–234 nm), facilitating interactions with target cells. The MTT assay showed that DAB was more cytotoxic toward HFF-1 cells (IC₅₀ = 27 µg/mL) compared to DAB-Br (IC₅₀ = 68 µg/mL). In contrast, the resazurin-based antifungal assay demonstrated that DAB-Br had superior antifungal activity, achieving a lower minimum inhibitory concentration (0.148 µg/µL), compared to DAB (0.295 µg/µL). A trypan blue exclusion test revealed that both dendrimers entered cells through membrane permeabilization, either temporarily or permanently, depending on the concentration and exposure time. At concentrations above 30 µg/mL, irreversible permeabilization was observed within two hours of treatment, accompanied by a decrease in membrane lipid order, indicating altered membrane integrity and permeability. Conversely, at lower concentrations (7.5–15 µg/mL), dendrimers induced only temporary membrane permeabilization, with membranes remaining intact, suggesting a reversible interaction with the lipid bilayer. Conducting thorough and systematic research to fully explore their biological activities could provide valuable insight for future applications. Full article

Some Vibrio parahaemolyticus strains cause translucent post-larvae disease (Vp_TPD), leading to significant economic losses in shrimp farming. We aimed to identify whether a mixture of natural antimicrobials, AuraAqua (Aq), can protect white-leg shrimp (Penaeus vannamei) against the lethal effects of Vp_TPD and to understand its biological mode of action. Herein, we demonstrate that Aq, an antimicrobial mixture composed of a blend of organic acids, citrus, and olive extracts, suppressed Vp_TPD virulence at sub-inhibitory concentrations and conferred robust protection to shrimp. The minimum inhibitory and bactericidal concentrations against the Vp_TPD isolate were at 0.05% and 0.2%, respectively. At 0.05–0.1%, Aq reduced bacterial growth and downregulated six major virulence genes (vhvp-1, vhvp-2, vhvp-3, pirA_Vp, pirB_Vp, pirAB_Vp), while leaving metabolic ldh expression unaltered. Parallel in vitro assays revealed diminished adhesion of Vp_TPD to primary shrimp gut and hepatopancreas epithelial cells and a ≈50% reduction in infection-induced extracellular H₂O₂, indicating an antioxidant effect. The treatment also triggered a time-dependent surge in extracellular alkaline phosphatase (ALP) activity, consistent with membrane permeabilization. In vivo, a challenge of post-larvae with 10⁴ CFU/mL Vp_TPD resulted in 91% mortality after 45 h; co-treatment with 0.1% and 0.2% Aq reduced mortality to ≈12% and ≈6%, respectively, while 1% Aq achieved ≈98% survival. The clinical protection test confirmed that 0.1% Aq preserved high survival across four pathogen inocula (10¹–10⁴ CFU/mL). Conclusively, Aq destabilized the pathogen and therefore transcriptionally silenced multiple virulence determinants, translating into significant in-pond protection for controlling Vp_TPD for shrimp aquaculture. Full article

The study of the effects of 2.45 GHz electromagnetic fields on the health and safety of people and organisms as a whole is essential due to their widespread use in everyday life. It is known that they can cause thermal and non-thermal effects—at the molecular, cellular and organismal level. Yeast suspensions were treated with 2.45 GHz microwave radiation in the near-field of antenna at two distances (2 and 4 cm) and two time periods (20 and 60 min)—setups resembling the use of mobile devices. The release of UV-absorbing substances from the cells was studied as an indicator of membrane permeabilization, total intracellular antioxidant activity and reduced glutathione were determined, and a comet assay for damage to the DNA was performed. A correlation between reduced antioxidants and increased membrane permeability during EMF treatment was observed at a distance of 2 cm for 20 min, suggesting the presence of oxidative stress, while a similar effect was not observed with conventional heating. Slightly increased membrane permeability was observed after irradiation for 60 min at a distance of 4 cm, but this was not related to the antioxidant status of the cells. A trend towards increased DNA damage was observed under both conditions. Full article

Nowadays, healthcare systems face two global challenges: the rise of multidrug-resistant pathogens and the growing incidence of cancer. Due to their broad spectrum of activities, antimicrobial peptides emerged as potential alternatives against both threats. Our group previously described the antifungal activity of the α-helical peptide Cm-p5, a derivative of the natural peptide Cm-p1, isolated from the coastal mollusk Cenchritis muricatus; however, its anti-cancer properties remained unexplored. Analyses through calorimetry and molecular dynamics simulations suggest the relevance of phosphatidylserine for the attachment of Cm-p5 to cancer cell membranes. Cm-p5 exhibited cytotoxic activity in a dose-dependent manner against A375 melanoma cells, without toxicity against non-malignant cells or hemolytic activity. DAPI/PI and DiSC3(5) staining confirmed permeabilization, disruption, and depolarization of A375 cytoplasmic membranes by Cm-p5. Furthermore, Annexin V-FITC/PI assay revealed the induction of cellular death in melanoma cells, which can result from the cumulative membrane damage and oxidative stress due to the overproduction of reactive oxygen species (ROS). Moreover, after the treatment, the proliferation of A375 cells was dampened for several days, suggesting that Cm-p5 might inhibit the recurrence of melanomas. These findings highlight the multifunctional nature of Cm-p5 and its potential for treating malignant melanoma. Full article

Anti-infectives include molecules that target microbes in the context of infection but lack antimicrobial activity under conventional growth conditions. We previously described D66, a small molecule that kills the Gram-negative pathogen Salmonella enterica serovar Typhimurium (S. Typhimurium) within cultured macrophages and murine tissues, with low host toxicity. While D66 fails to inhibit bacterial growth in standard media, the compound is bacteriostatic and disrupts the cell membrane voltage gradient without lysis under growth conditions that permeabilize the outer membrane or reduce efflux pump activity. To gain insights into specific bacterial targets of D66, we pursued two genetic approaches. Selection for resistance to D66 revealed spontaneous point mutations that mapped within the gmhB gene, which encodes a protein involved in the biosynthesis of the lipopolysaccharide core molecule. E. coli and S. Typhimurium gmhB mutants exhibited increased resistance to antibiotics, indicating a more robust barrier to entry. Conversely, S. Typhimurium transposon insertions in genes involved in outer membrane permeability or efflux pump activity reduced fitness in the presence of D66. Together, these observations underscore the significance of the bacterial cell envelope in safeguarding Gram-negative bacteria from small molecules. Full article

As emerging contaminants increasingly detected in aquatic and terrestrial ecosystems, environmental estrogens (EEs) pose significant ecological risks to marine ecosystems, particularly affecting photosynthetic microorganisms occupying fundamental roles in marine food webs. This review summarizes the current knowledge on the toxicological effects of EEs in marine microalgae through a systematic analysis of dose-dependent physiological, biochemical, and molecular responses. Experimental evidence reveals a biphasic response pattern characterized by growth promotion and photosynthetic enhancement in microalgae under low-concentration EE exposure (0.1–10 μg/L), while marked inhibition of both growth and photosynthetic activity was observed at elevated EE concentrations (>50 μg/L). Notably, sustained EE exposure induces metabolic reprogramming, manifested through reduced protein and polysaccharide biosynthesis concurrent with accelerated lipid accumulation. Cellular stress responses include significant ultrastructural alterations such as chloroplast membrane disruption, cell wall thickening, and the formation of multicellular aggregates. The study further elucidates the concentration-dependent modulation of toxin metabolism, with sublethal doses stimulating intracellular microcystin synthesis (1.5–2.3-fold increase), while acute exposure triggers toxin release through membrane permeabilization. At molecular levels, transcriptomic analyses identify the up-regulation of heat shock proteins (HSP70/90) and the differential expression of genes governing cell cycle progression (cyclin-D), apoptotic pathways (caspase-3), photosynthetic electron transport (psbA), and oxidative stress responses (SOD, CAT). These findings demonstrate that EEs exert multilevel impacts on microalgal physiology through interference with fundamental metabolic processes, potentially disrupting marine primary productivity and biogeochemical cycles. The identified response mechanisms provide critical insights for environmental risk assessment and establish a conceptual framework for investigating estrogenic pollutant effects in aquatic ecosystems. Full article

Lysosomal dysfunction and endoplasmic reticulum (ER) stress play essential roles in cancer cell survival, growth, and stress adaptation. Among the various stressors in the tumor microenvironment, oxidative stress (OS) is a central driver that exacerbates both lysosomal and ER dysfunction. In healthy cells, the ER manages protein folding and redox balance, while lysosomes regulate autophagy and degradation. Cancer cells, however, are frequently exposed to elevated levels of reactive oxygen species (ROS), which disrupt protein folding in the ER and damage lysosomal membranes and enzymes, promoting dysfunction. Persistent OS activates the unfolded protein response (UPR) and contributes to lysosomal membrane permeabilization (LMP), leading to pro-survival autophagy or cell death depending on the context and on the modulation of pathways like PERK, IRE1, and ATF6. Cancer cells exploit these pathways by enhancing their tolerance to OS and shifting UPR signaling toward survival. Moreover, lysosomal impairment due to ROS accumulation compromises autophagy, resulting in the buildup of damaged organelles and further amplifying oxidative damage. This vicious cycle of ROS-induced ER stress and lysosomal dysfunction contributes to tumor progression, therapy resistance, and metabolic adaptation. Thus, targeting lysosomal and ER stress responses offers potential as cancer therapy, particularly in increasing oxidative stress and promoting apoptosis. This review explores the interconnected roles of lysosomal dysfunction, ER stress, and OS in cancer, focusing on the mechanisms driving their crosstalk and its implications for tumor progression and therapeutic resistance. Full article

Reactive oxygen species (ROS) are critical signalling molecules, but their overproduction leads to oxidative stress (OS), a common denominator in the pathogenesis of numerous non-communicable diseases (NCDs) and aging. General antioxidant therapies have largely been unsuccessful, highlighting the need for a deeper understanding of ROS amplification mechanisms to develop targeted interventions. This review proposes a unified, self-amplifying “vicious cycle” of inter-organelle crosstalk that drives pathological ROS elevation and cellular damage. We outline a pathway initiated by extracellular stressors that co-activate plasma membrane TRPM2 channels and NADPH oxidase-2. This synergy elevates cytoplasmic Ca²⁺, leading to lysosomal dysfunction and permeabilization, which in turn releases sequestered Zn²⁺. Mitochondrial uptake of this labile Zn²⁺ impairs electron transport chain function, particularly at Complex III, resulting in mitochondrial fragmentation, loss of membrane potential and a burst of mitochondrial ROS (mtROS). These mtROS diffuse to the nucleus, activating PARP-1 and generating ADPR, which further stimulates TRPM2, thereby perpetuating the cycle. This “circular domino effect” integrates signals generated across the plasma membrane (Ca²⁺), lysosomes (Zn²⁺), mitochondria (ROS) and nucleus (ADPR), leading to progressive organelle failure, cellular dysfunction, and ultimately cell death. Understanding and targeting specific nodes within this TRPM2-NOX2-Ca²⁺-Zn²⁺-mtROS-ADPR axis offers novel therapeutic avenues for NCDs by selectively disrupting pathological ROS amplification while preserving essential physiological redox signalling. Full article

Cryptosporidium spp. oocysts are highly resistant to conventional disinfection methods and have been associated with foodborne outbreaks linked to unpasteurized fruit and vegetable juices. This study aimed to evaluate the effectiveness of Pulsed Electric Fields (PEF) in permeabilizing Cryptosporidium oocysts in water, apple juice, and carrot juice. Oocysts were exposed to monopolar square-wave pulses (3 µs) at electric field strengths ranging from 15 to 35 kV/cm, with treatment times up to 180 µs, and application temperatures between 25 °C and 60 °C. Membrane permeabilization was assessed using propidium iodide uptake via fluorescence microscopy and flow cytometry. Results showed that oocyst permeabilization increased with electric field strength, treatment time, and temperature, with up to 90% permeabilization achieved at 35 kV/cm and 45 °C. Carrot juice treatments yielded higher permeabilization levels than apple juice, attributed to greater electrical conductivity and energy input. Temperatures below 60 °C alone had negligible effects, but synergistically enhanced PEF efficacy. These findings demonstrate that PEF, particularly when combined with mild heat, is a promising non-thermal technology for reducing Cryptosporidium viability in beverages, offering an effective alternative for improving the microbiological safety of minimally processed juices while preserving sensory and nutritional quality. Full article

Breast cancer remains a leading cause of cancer-related morbidity and mortality among women worldwide. Its treatment is complicated by molecular heterogeneity and the frequent development of multidrug resistance (MDR). Conventional drug delivery approaches are often limited by poor aqueous solubility, rapid systemic clearance, non-specific biodistribution, and off-target toxicity. This review will critically explore the possibility of surfactant-based drug delivery systems (DDSs) in addressing the constraints of standard breast cancer treatments. It focuses on the mechanisms by which surfactants promote solubility, facilitate cellular uptake, and overcome drug resistance, while also analyzing current therapeutic success and future directions. A thorough review of preclinical and clinical investigations was undertaken, focusing on important surfactant-based DDSs such as polymeric micelles, nanoemulsions, liposomes, and self-emulsifying systems (SEDDSs). Mechanistic insights into surfactant functions, such as membrane permeabilization and efflux pump inhibition, were studied alongside delivery systems incorporating ligands and co-loaded medicines. Pluronic^® micelles, TPGS-based systems, biosurfactant-stabilized nanoparticles, and lipid-based carrier surfactant platforms improve medication solubility, stability, and delivery. Genexol^® are examples of formulations demonstrating effective use and FDA translational potential. These systems now incorporate stimuli-responsive release mechanisms—such as pH, temperature, redox, immuno- and photodynamic treatment—artificial intelligence treatment design, and tailored treatment advancement, and responsive tailoring. Surfactant-enabled DDSs can improve breast cancer care. Innovative approaches for personalized oncology treatment are countered by the enduring challenges of toxicity, regulatory hurdles, and diminished scalability. Full article

Oleocanthal (OC), a secoiridoid phenolic compound exclusive to extra virgin olive oil (EVOO), has emerged as a promising nutraceutical with multifaceted anti-cancer properties. Despite its well-characterized anti-inflammatory and antioxidant effects, the mechanistic breadth and translational potential of OC in oncology remain underexplored and fragmented across the literature. This comprehensive review synthesizes and critically analyzes recent advances in the molecular, pharmacological, and translational landscape of OC’s anti-cancer activities, providing an integrative framework to bridge preclinical evidence with future clinical application. We delineate the pleiotropic mechanisms by which OC modulates cancer hallmarks, including lysosomal membrane permeabilization (LMP)-mediated apoptosis, the inhibition of key oncogenic signaling pathways (c-MET/STAT3, PAR-2/TNF-α, COX-2/mPGES-1), the suppression of epithelial-to-mesenchymal transition (EMT), angiogenesis, and metabolic reprogramming. Furthermore, this review uniquely highlights the emerging role of OC in modulating drug resistance mechanisms by downregulating efflux transporters and sensitizing tumors to chemotherapy, targeted therapies, and immunotherapies. We also examine OC’s bidirectional interaction with gut microbiota, underscoring its systemic immunometabolic effects. A major unmet need addressed by this review is the lack of consolidated knowledge regarding OC’s pharmacokinetic limitations and drug–drug interaction potential in the context of polypharmacy in oncology. We provide an in-depth analysis of OC’s poor bioavailability, extensive first-pass metabolism, and pharmacogenomic interactions, and systematically compile preclinical evidence on advanced delivery platforms—including nanocarriers, microneedle systems, and peptide–drug conjugates—designed to overcome these barriers. By critically evaluating the mechanistic, pharmacological, and translational dimensions of OC, this review advances the field beyond isolated mechanistic studies and offers a strategic blueprint for its integration into precision oncology. It also identifies key research gaps and outlines the future directions necessary to transition OC from a nutraceutical of dietary interest to a viable adjunctive therapeutic agent in cancer treatment. Full article

Antibiotics are indispensable in medical patient care, yet they may elicit off-target effects, particularly by affecting mitochondrial function. This study investigates three commonly used antibiotics, gentamicin, ciprofloxacin, and amoxicillin, for their direct effects on mitochondrial respiration and membrane potential. Using high-resolution respirometry, we show that gentamicin and ciprofloxacin markedly increase mitochondrial leak respiration in permeabilized human embryonic kidney cells, suggesting alterations in the mitochondrial inner membrane. This is supported by a gentamicin-induced decrease in mitochondrial membrane potential. Especially gentamicin, but also ciprofloxacin, dose- and time-dependently inhibit oxidative phosphorylation and the mitochondrial electron transfer capacity, pronouncedly in the NADH-linked but also in the succinate-linked pathway. Furthermore, gentamicin decreases Complex IV (CIV) activity in a time-dependent fashion. In contrast, amoxicillin has no significant effect on mitochondrial respiration. These findings emphasize the importance of evaluating the potential direct toxicity of antibiotics on mitochondria, as they are most critical off-target sites. High-resolution respirometry provides a powerful approach to characterize such effects early in the drug development process. Full article