Search Results (465)

Gene transfection is a fundamental technique in the fields of biological research and therapeutic innovation. Due to their biocompatibility and membrane-mimetic properties, lipid vectors serve as essential tools in transfection. The successful delivery of genetic material into the cytoplasm is contingent upon the fusion of the vector and cellular membranes, which enables hydrophilic polynucleic acids to traverse the hydrophobic barriers of two intervening membranes. This review examines the critical role of membrane fusion in lipofection efficiency, with a particular focus on the molecular mechanisms that govern lipoplex–membrane interactions. This analysis will examine the key challenges inherent to the fusion process, from achieving initial membrane proximity to facilitating final content release through membrane remodeling. In contrast to viral vectors, which utilize specialized fusion proteins, lipid vectors necessitate a strategic formulation and environmental optimization to enhance their fusogenicity. This review discusses recent advances in vector design and fusion-promoting strategies, emphasizing their potential to improve gene delivery yield. It highlights the importance of understanding lipoplex–membrane fusion mechanisms for developing next-generation delivery systems and emphasizes the need for continued fundamental research to advance lipid-mediated transfection technology. Full article

In this study, molecular dynamics simulations were employed to analyze interactions between phospholipid membranes and human serum albumin (HSA) in the presence of mono- and divalent cations. Two types of membranes, composed of dipalmitoyl phosphatidylcholine (DPPC) and dipalmitoyl phosphatidylethanolamine (DPPE), were utilized. The results revealed that both systems exhibited high stability. The DPPE complexes displayed a greater affinity for albumin compared to DPPC. The high stability of the complexes was attributed to a high number of ionic contacts and hydrogen bonds. The presence of mono- and divalent metal cations significantly influenced the membrane’s capacity to bind proteins. However, these effects varied depending on the phospholipid composition of the bilayer. The studies confirmed the relatively low ability of DPPC to bind potassium ions, as previously observed by others. Consequently, the DPPC/HSA/K⁺ complex was found to be the least stable among the systems studied. While DPPC interactions were limited to HSA domains I and II, DPPE was able to interact with all domains of the protein. Both lipid bilayers exhibited substantial structural changes and characteristic curvature induced by interactions with HSA, which confirms the formation of relatively strong interactions capable of influencing the arrangement of the phospholipids. Full article

Acid adaptation in Escherichia coli can induce antimicrobial resistance (AMR), posing challenges to global public health. We investigated the effects of acid adaptation on antimicrobial susceptibility, gene expression, zeta potential, and the outer membrane (OM) properties of Escherichia coli NCCP 13719. The acid-adapted (AA) strain exhibited increased resistance to multiple antimicrobials, with minimum inhibitory concentrations for colistin and polymyxin B increasing eight- and two-fold, respectively. Transcriptomic analysis identified 2225 differentially expressed genes, including upregulated genes associated with resistance to cationic antimicrobial peptides such as arnCTE, marA, and tolC. The upregulation of the arn operon suggests modifications in lipid A of lipopolysaccharides (LPS), reducing the negative charge of the OM and decreasing polymyxin binding affinity. Zeta potential measurements indicated a shift toward a less negative surface charge in the AA strain, which is consistent with LPS modifications. The AA strain also showed decreased OM permeability, which correlated with increased resistance to antimicrobials that penetrate the OM. These mechanisms collectively diminish the efficacy of polymyxins and highlight the potential for environmental factors to drive antimicrobial resistance. In conclusion, the acid adaptation of E. coli NCCP 13719 enhances AMR through changes in gene expression and OM modifications, highlighting the need for careful control of acidic environments during the treatment of medical devices and wastewater from food processing to prevent the emergence of resistant strains. Full article

Background/Objectives: Standard-of-care influenza vaccines contain antigens that are typically derived from components of wild type (WT) influenza viruses. Often, these antigens elicit strain-specific immune responses and are susceptible to mismatch in seasons where antigenic drift is prevalent. Thanks to advances in viral surveillance and sequencing, influenza vaccine antigens can now be optimized using computationally derived methodologies and algorithms to enhance their immunogenicity. Methods: Mice and ferrets that had been previously exposed to historical H1N1 and H3N2 influenza viruses were vaccinated intramuscularly with bivalent mixtures of H1 and H3 recombinant hemagglutinin (rHA) proteins, which were generated using a computationally optimized broadly reactive antigen (COBRA) design methodology. The vaccine antigens were mixed with a cationic lipid nanoparticle adjuvant, Infectimune^®, which promotes both humoral and cellular immune responses. Results: Mice and ferrets vaccinated with Infectimune^® and COBRA rHAs elicited protective antibody titers against panels of H1N1 and H3N2 influenza viruses isolated over the past 10 years. These animals also had antibodies that neutralized numerous modern H1N1 and H3N2 influenza viruses in vitro. When challenged with the A/Victoria/2570/2019 H1N1 influenza virus, the COBRA rHA vaccinated animals had minimal weight loss, and no detectable virus was present in their respiratory tissues on day 3 post-infection. Conclusions: These results demonstrate that COBRA rHA vaccines formulated with Infectimune^® elicit protective antibody responses against influenza strains, which were isolated across periods of time when standard-of-care vaccines were frequently reformulated, thus reducing the need to update vaccines on a nearly annual basis. Full article

Lipid nanoparticles (LNPs) have shown promise as a delivery system for nucleic acid-based therapeutics, including DNA, siRNA, and mRNA vaccines. The immune system plays a critical role in the response to these nanocarriers, with innate immune cells initiating an early response and adaptive immune cells mediating a more specific reaction, sometimes leading to potential adverse effects. Recent studies have shown that the innate immune response to LNPs is mediated by Toll-like receptors (TLRs) and other pattern recognition receptors (PRRs), which recognize the lipid components of the nanoparticles. This recognition can trigger the activation of inflammatory pathways and the production of cytokines and chemokines, leading to potential adverse effects such as fever, inflammation, and pain at the injection site. On the other hand, the adaptive immune response to LNPs appears to be primarily directed against the protein encoded by the mRNA cargo, with little evidence of an ongoing adaptive immune response to the components of the LNP itself. Understanding the relationship between LNPs and the immune system is critical for the development of safe and effective nucleic acid-based delivery systems. In fact, targeting the immune system is essential to develop effective vaccines, as well as therapies against cancer or infections. There is a lack of research in the literature that has systematically studied the factors that influence the interaction between LNPs and the immune system and further research is needed to better elucidate the mechanisms underlying the immune response to LNPs. In this review, we discuss LNPs’ composition, physico-chemical properties, such as size, shape, and surface charge, and the protein corona formation which can affect the reactivity of the immune system, thus providing a guide for the research on new formulations that could gain a favorable efficacy/safety profile. Full article

Currently, lipid/polymer membranes are used in taste sensors to quantify food taste. This research aims to improve sweetness sensors by more selectively detecting uncharged sweetening substances, which have difficulty obtaining a potentiometric response. Lipid/polymer membranes with varying amounts of tetradodecylammonium bromide (TDAB) and 1,2,4-benzene tricarboxylic acid (trimellitic acid) were prepared. The carboxyl groups of trimellitic acid bind metal cations, and the sweetness intensity is estimated by measuring the potential change, as a sensor response, when these cations are complexed with sugars. This research showed that the potential of a sensor using the membrane with enough trimellitic acid in a sucrose solution remained constant, regardless of TDAB amounts, but the potential in the tasteless, so-called reference solution, depended on TDAB. By optimizing the content of TDAB and trimellitic acid, a sensor response of −100 mV was achieved, which is over 20% more sensitive than a previous sensor. This sensor also demonstrated increased selectivity to sweetness, with similar interference from other tastes (saltiness, sourness, umami, and bitterness) compared to previous sensors. As a result, the sensitivity to sweetness was successfully improved. This result contributes to the development of novel sensors, further reducing the burden on humans in quality control and product development. Full article

One of the major challenges in gene therapy is the efficient and safe introduction of nucleic acids into eukaryotic cells. This process requires overcoming various biological barriers and navigating complex pathways to reach target cells and achieve their biological function. To address this obstacle, numerous transfection methods have been developed, including physical techniques and the use of genetic vectors, both viral and non-viral. However, to date, no transfection method is 100% safe and efficient. Within the spectrum of non-viral genetic vectors, cationic liposomes formed by cationic lipids stand out for their ability to protect and deliver therapeutic NA. These liposomes offer greater biocompatibility and lower immunogenicity compared to viral vectors, although they still do not match the efficiency of viral delivery systems. Consequently, ongoing research focuses on synthesizing a wide variety of cationic lipids in the search for compounds that provide high transfection efficiency with minimal cytotoxicity. This study aimed to design and synthesize a novel cationic lipid (CholCadLys) derived from natural cellular molecules for transferring genetic material to eukaryotic cells. The lipid was synthesized using cholesteryl chloroformate for the hydrophobic region, cadaverine as a linker, and lysine for the polar region, connected by carbamate and amide bonds, respectively. Identification was confirmed through thin-layer chromatography, purification through preparative chromatography, and characterization via infrared spectroscopy and mass spectrometry. The synthesis yielded a 60% success rate, with stable nanoliposomes averaging 76 nm in diameter. Liposomes were formed using this CL and commercial neutral lipids, characterized by transmission electron microscopy and Nanoparticle Tracking Analysis. These liposomes, combined with plasmid DNA, formed lipoplexes used to transfect Hek-293 FT cells, achieving up to 40% transfection efficiency without cytotoxicity in the mixture of CholCadLys and CholCad. This novel CL demonstrates potential as an efficient, safe, and cost-effective gene transfer system, facilitating further development in gene therapy. Full article

In this study, we investigated the anti-fatigue effects of black ginseng ginsenosides using exercise performance tests, serum analyses, and gene expression profiling. No significant differences in dietary intake or body weight were observed between groups. The low-dose black ginseng (LBG) group showed no significant improvements in swimming and rotating rod tests. In contrast, the medium (MBG)- and high-dose (HBG) groups showed notable increases in swimming time and significant improvements in the rotating rod test. All treatment groups exhibited longer running times, particularly the HBG group. Serum analysis revealed increased muscle and hepatic glycogen, catalase, and lactate dehydrogenase levels in the MBG and HBG groups, whereas lactate, lipid peroxide, and superoxide dismutase levels were decreased. Additionally, gene expression analysis showed significant upregulation of key antioxidant and mitochondrial function genes, including those encoding cationic amino acid transporter 2, stearoyl-CoA desaturase-2, nuclear respiratory factor 1, nuclear factor erythroid 2-related factor 2, mitochondrial transcription factor A, cytochrome c oxidase II, and NADH–ubiquinone oxidoreductase core subunit 1, particularly in the HBG group, indicating enhanced antioxidant capacity and improved mitochondrial function. These findings suggested that black ginseng ginsenosides effectively mitigated fatigue. Full article

Background/Objectives: Hemophilia B is a hereditary bleeding disorder due to the production of liver malfunctional factor IX (FIX). Gene therapy with viral vectors offers a cure. However, applications are limited due to pre-existing antibodies, eligibility for children under 12 years of age, hepatotoxicity, and excessive costs. Lipid nanoparticles are a potential alternative owing to their biocompatibility, scalability, and non-immunogenicity. However, their therapeutic applications are still elusive due to the poor transfection efficiencies in delivering plasmid DNA into primary cells and target organs in vivo. To develop efficient liver-targeted lipid nanoparticles, we explored galactosylated lipids to target asialoglycoprotein receptors (ASGPRs) abundantly expressed on hepatocytes. Methods: We developed 12 novel liposomal formulations varying the galactose lipid Gal-LNC 5, cationic lipid MeOH16, DOPE, and cholesterol. We evaluated their physicochemical properties, toxicity profiles, and transfection efficiencies in hepatic cell lines. Among the formulations, Gal-LNC 5 could efficiently transfect the reporter plasmid eGFP in hepatic cell lines and specifically distribute into the liver in vivo. Toward developing functional factor IX, we cloned Padua mutant FIX-L in a CpG-free backbone to enhance the expression and duration. Results: We demonstrated superior expression of FIX with our galactosylated lipid nanoparticle system. Conclusions: The current research presents a specialized lipid nanoparticle system viz. Gal-LNC which is a specialized lipid nanoparticle system for liver-targeted gene therapy in hemophilia B patients that has potential for clinical use. The Gal-LNC successfully delivers a CpG-free Padua FIX gene to liver cells, producing therapeutically relevant levels of FIX protein. Among its benefits are the ideal qualities of stability, targeting the liver specifically, and maximizing efficiency of transfection. Optimization of liver-targeting lipid nanoparticle systems and function FIX plasmids will pave the way for novel lipid nanoparticle-based gene therapy products for hemophilia B and other monogenic liver disorders. Full article

Cationic vaccines of nanometric sizes can directly perform the delivery of antigen(s) and immunomodulator(s) to dendritic cells in the lymph nodes. The positively charged nanovaccines are taken up by antigen-presenting cells (APCs) of the lymphatic system often originating the cellular immunological defense required to fight intracellular microbial infections and the proliferation of cancers. Cationic molecules imparting the positive charges to nanovaccines exhibit a dose-dependent toxicity which needs to be systematically addressed. Against the coronavirus, mRNA cationic nanovaccines evolved rapidly. Nowadays cationic nanovaccines have been formulated against several infections with the advantage of cationic compounds granting protection of nucleic acids in vivo against biodegradation by nucleases. Up to the threshold concentration of cationic molecules for nanovaccine delivery, cationic nanovaccines perform well eliciting the desired Th 1 improved immune response in the absence of cytotoxicity. A second strategy in the literature involves dilution of cationic components in biocompatible polymeric matrixes. Polymeric nanoparticles incorporating cationic molecules at reduced concentrations for the cationic component often result in an absence of toxic effects. The progress in vaccinology against cancer involves in situ designs for cationic nanovaccines. The lysis of transformed cancer cells releases several tumoral antigens, which in the presence of cationic nanoadjuvants can be systemically presented for the prevention of metastatic cancer. In addition, these local cationic nanovaccines allow immunotherapeutic tumor treatment. Full article

Phospholipids and their metabolites play an important role in maintaining the membrane integrity and the metabolic functions of keratinocytes under physiological conditions and in the regeneration process after exposure to high-energy UVB radiation. Therefore, in the search for compounds with a protective and regenerative effect on keratinocyte phospholipids, the effectiveness of two antioxidant compounds has been tested: a stable derivative of ascorbic acid, 3-O-ethyl ascorbic acid (EAA) and cannabigerol (CBG), both of which are primarily located in the membrane structures of keratinocytes. In addition, this study has demonstrated that EAA and CBG, especially in a two-component combination, enhance the antioxidant properties of keratinocytes and reduce lipid peroxidation assessed at the level of MDA (malondialdehyde)/neuroprostanes. Moreover, by reducing the activity of enzymes that metabolise phospholipids, free PUFAs (polyunsaturated fatty acids) and endocannabinoids (PLA2; phospholipase A2, COX1/2; cyclooxygenases 1/2, LOX-5; lipoxygenase 5, FAAH; fatty acid amide hydrolase, MAGL; monoacylglycerol lipase), antioxidants have been found to regulate the levels of endocannabinoids (AEA; anandamide, 2-AG; 2-arachidonoylglycerol, PEA; palmitoylethanolamide) and eicosanoids (PGD2; prostaglandin D2, PGE2; prostaglandin E2, 15-d-PGJ2; 15-deoxy-Δ12,14-prostaglandin J2, 15-HETE; 15-hydroxyeicosatetraenoic acid), that are enhanced by UVB radiation. The metabolic effect of both groups of PUFA metabolites is mainly related to the activation of G protein-related receptors (CB1/2; cannabinoid receptor 1 and 2, PPARγ; peroxisome proliferator-activated receptor gamma, TRPV1; transient receptor potential cation channel subfamily V member 1), the expression of which is reduced under the influence of EAA, CBG, and especially the two-component combination. It promotes the regeneration of keratinocyte metabolism disrupted by UVB, particularly in relation to redox balance and inflammation. Full article

Analyses were conducted on extracts from the fruits of P. pyraster and P. ×myloslavensis. Extraction with 80% methanol was performed at room temperature. The total phenolic content was determined by spectrophotometry using the Folin–Ciocalteu reagent, with gallic acid as the reference standard. Phenolic compounds and organic acids were identified on a liquid chromatograph. The antioxidative activity of the extracts was tested in relation to linoleic acid incubation of the emulsions for 19 h based on the neutralization of the DPPH radical (2,2-diphenyl-1-picrylhydrazyl) and the ABTS cation radical (2,2′-azino-bis[3-ethylbenzothiazoline-6-sulfonic acid]) as well as by the ferric reducing antioxidant power (FRAP) assay. The analyses showed that the extract from P. pyraster fruits is characterized by a higher content of phenolic compounds and a higher antioxidative potential compared with that from P. ×myloslavensis. In extracts of both pear species, seven phenolic compounds and four organic acids were identified. The total fiber content in pears of P. pyraster and P. ×myloslavensis was determined at 36.45 g and 24.74 g/100 g d.m. of the pear fruits, of which most comprised the insoluble fraction (32.49 g and 20.86/100 g, respectively). The results of the conducted research are highly significant, as they confirm that pears contain many valuable nutrients and biologically active compounds, including antioxidants and dietary fiber. Adding pear extracts to food products may offer a way to boost their health benefits while also broadening the variety of items that have appealing sensory characteristics. Moreover, research has shown that fruit extracts can help to prolong the shelf life of food products by safeguarding them against lipid oxidation and the decline in their nutritional value. Full article

Background: The persistence of antibiotic resistance has incited a strong interest in the discovery of agents with novel antimicrobial mechanisms. The direct killing of multidrug-resistant bacteria by cationic antimicrobial peptides (AMPs) underscores their importance in the fight against infections associated with antibiotic resistance. Despite a vast body of AMP literature demonstrating a plurality in structural classes, AMP engineering has been largely skewed toward peptides with idealized amphipathic helices (H-amphipathic). In contrast to helical amphipathicity, we designed a series of peptides that display the amphipathic motifs in the primary structure. We previously developed a rational framework for designing AMP libraries of H-amphipathic peptides consisting of Arg, Trp, and Val (H-RWV, with a confirmed helicity up to 88% in the presence of membrane lipids) tested against the most common MDR organisms. Methods: In this study, we re-engineered one of the series of the H-RWV peptides (8, 10, 12, 14, and 16 residues in length) to display the amphipathicity in the primary structure by side-by-side (linear) alignment of the cationic and hydrophobic residues into the 2 separate linear amphipathic (L-amphipathic) motifs. We compared the 2 series of peptides for antibacterial activity, red blood cell (RBC) lysis, killing and membrane-perturbation properties. Results: The L-RWV peptides achieved the highest antibacterial activity at a minimum length of 12 residues (L-RWV12, minimum optimal length or MOL) with the lowest mean MIC of 3–4 µM, whereas the MOL for the H-RWV series was reached at 16 residues (H-RWV16). Overall, H-RWV16 displayed the lowest mean MIC at 2 µM but higher levels of RBC lysis (25–30%), while the L-RWV series displayed minor RBC lytic effects at the test concentrations. Interestingly, when the S. aureus strain SA719 was chosen because of its susceptibility to most of the peptides, none of the L-RWV peptides demonstrated a high level of membrane perturbation determined by propidium iodide incorporation measured by flow cytometry, with <50% PI incorporation for the L-RWV peptides. By contrast, most H-RWV peptides displayed almost up to 100% PI incorporation. The results suggest that membrane perturbation is not the primary killing mechanism of the L-amphipathic RWV peptides, in contrast to the H-RWV peptides. Conclusions: Taken together, the data indicate that both types of amphipathicity may provide different ideal pharmacological properties that deserve further investigation. Full article

Na⁺-K⁺ ATPase is an integral component of cardiac sarcolemma and consists of three major subunits, namely the α-subunit with three isoforms (α₁, α₂, and α₃), β-subunit with two isoforms (β₁ and β₂) and γ-subunit (phospholemman). This enzyme has been demonstrated to transport three Na and two K ions to generate a trans-membrane gradient, maintain cation homeostasis in cardiomyocytes and participate in regulating contractile force development. Na⁺-K⁺ ATPase serves as a receptor for both exogenous and endogenous cardiotonic glycosides and steroids, and a signal transducer for modifying myocardial metabolism as well as cellular survival and death. In addition, Na⁺-K⁺ ATPase is regulated by different hormones through the phosphorylation/dephosphorylation of phospholemman, which is tightly bound to this enzyme. The activity of Na⁺-K⁺ ATPase has been reported to be increased, unaltered and depressed in failing hearts depending upon the type and stage of heart failure as well as the association/disassociation of phospholemman and binding with endogenous cardiotonic steroids, namely endogenous ouabain and marinobufagenin. Increased Na⁺-K⁺ ATPase activity in association with a depressed level of intracellular Na⁺ in failing hearts is considered to decrease intracellular Ca²⁺ and serve as an adaptive mechanism for maintaining cardiac function. The slight to moderate depression of Na⁺-K⁺ ATPase by cardiac glycosides in association with an increased level of Na⁺ in cardiomyocytes is known to produce beneficial effects in failing hearts. On the other hand, markedly reduced Na⁺-K⁺ ATPase activity associated with an increased level of intracellular Na⁺ in failing hearts has been demonstrated to result in an intracellular Ca²⁺ overload, the occurrence of cardiac arrhythmias and depression in cardiac function during the development of heart failure. Furthermore, the status of Na⁺-K⁺ ATPase activity in heart failure is determined by changes in isoform subunits of the enzyme, the development of oxidative stress, intracellular Ca²⁺-overload, protease activation, the activity of inflammatory cytokines and sarcolemmal lipid composition. Evidence has been presented to show that marked alterations in myocardial cations cannot be explained exclusively on the basis of sarcolemma alterations, as other Ca²⁺ channels, cation transporters and exchangers may be involved in this event. A marked reduction in Na⁺-K⁺ ATPase activity due to a shift in its isoform subunits in association with intracellular Ca²⁺-overload, cardiac energy depletion, increased membrane permeability, Ca²⁺-handling abnormalities and damage to myocardial ultrastructure appear to be involved in the progression of heart failure. Full article

Background/Objectives: The development of polymer–lipid hybrid nanoparticles (PLNs) is a promising area of research, as it can help increase the stability of cationic lipid carriers. Hybrid PLNs are core–shell nanoparticle structures that combine the advantages of both polymer nanoparticles and liposomes, especially in terms of their physical stability and biocompatibility. Natural polymers such as polyhydroxyalkanoate (PHA) can be used as a matrix for the PLNs’ preparation. Methods: In this study, we first obtained stable cationic hybrid PLNs using a cationic liposome (CL) composed of a polycationic lipid 2X3 (1,26-bis(cholest-5-en-3β-yloxycarbonylamino)-7,11,16,20-tetraazahexacosane tetrahydrochloride), helper lipid DOPE (1,2-dioleoyl-sn-glycero-3-phosphoethanolamine), and the hydrophobic polymer mcl-PHA, which was produced by the soil bacterium Pseudomonas helmantisensis P1. Results: The new polymer-lipid carriers effectively encapsulated and delivered model mRNA-eGFP (enhanced green fluorescent protein mRNA) to BHK-21 cells. We then evaluated the role of mcl-PHA in increasing the stability of cationic PLNs in ionic solutions using dynamic light scattering data, electrophoretic mobility, and transmission electron microscopy techniques. Conclusions: The results showed that increasing the concentration of PBS (phosphate buffered saline) led to a decrease in the stability of the CLs. At high concentrations of PBS, the CLs aggregate. In contrast, the presence of isotonic PBS did not result in the aggregation of PLNs, and the particles remained stable for 120 h when stored at +4 °C. The obtained results show that PLNs hold promise for further in vivo studies on nucleic acid delivery. Full article