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Self-assembled structures have numerous applications including drug delivery, solubilization, and food science. However, to date investigations into self-assembled structures have been largely limited to water, with some additives. This limits the types of assemblies that can form, as well as the accessible temperature range. Non-aqueous, polar solvents such as ionic liquids and deep eutectic solvents offer alternative self-assembly media that can overcome many of these challenges. These novel solvents can be designed to support specific types of assemblies or to remain stable under more extreme conditions. This review highlights recent advances in the field of self-assembly in polar non-aqueous solvents. Here we quantify the contribution of certain solvent properties such as nanostructure and solvent cohesion to lipid self-assembly. While this field is still relatively new, preliminary design rules are emerging, such as increasing hydrophobic regions leading to decreasing solvent cohesion, with a consequent reduction in lipid phase diversity. Ultimately, this review demonstrates the capacity for solvent control of lipid assemblies while also drawing attention to areas that need further work. With more systematic studies, solvents could be explicitly designed to achieve specific lipid assemblies for use in target applications, such as cargo delivery to particular cell types (e.g. cancerous), or triggered release under desired conditions (e.g. pH for release on wound infection).
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Proteins in ionic liquids (ILs) and deep eutectic solvents (DESs) have gained significant attention due to their potential applications in various fields, including biocatalysis, bioseparation, biomolecular delivery, and structural biology. Scattering approaches including dynamic light scattering (DLS) and small-angle X-ray and neutron scattering (SAXS and SANS) have been used to understand the solution behavior of proteins at the nanoscale and microscale. This review provides a thorough exploration of the application of these scattering techniques to elucidate protein properties in ILs and DESs. Specifically, the review begins with the theoretical foundations of the relevant scattering approaches and describes the essential solvent properties of ILs and DESs linked to scattering such as refractive index, scattering length density, ion-pairs, liquid nanostructure, solvent aggregation, and specific ion effects. Next, a detailed introduction is provided on protein properties such as type, concentration, size, flexibility and structure as observed through scattering methodologies. This is followed by a review of the literature on the use of scattering for proteins in ILs and DESs. It is highlighted that enhanced data analysis and modeling tools are necessary for assessing protein flexibility and structure, and for understanding protein hydration, aggregation and specific ion effects. It is also noted that complementary approaches are recommended for comprehensively understanding the behavior of proteins in solution due to the complex interplay of factors, including ion-binding, dynamic hydration, intermolecular interactions, and specific ion effects. Finally, the challenges and potential research directions for this field are proposed, including experimental design, data analysis approaches, and supporting methods to obtain fundamental understandings of complex protein behavior and protein systems in solution. We envisage that this review will support further studies of protein interface science, and in particular studies on solvent and ion effects on proteins.
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Líquidos Iónicos , Proteínas , Dispersión del Ángulo Pequeño , Líquidos Iónicos/química , Proteínas/química , Disolventes Eutécticos Profundos/química , Soluciones , Difracción de Neutrones , Difracción de Rayos X , Dispersión Dinámica de Luz , Solventes/químicaRESUMEN
Low-melting liquid metals are emerging as a new group of highly functional solvents due to their capability to dissolve and alloy various metals in their elemental state to form solutions as well as colloidal systems. Furthermore, these liquid metals can facilitate and catalyze multiple unique chemical reactions. Despite the intriguing science behind liquid metals and alloys, very little is known about their fundamental structures in the nanometric regime. To bridge this gap, this work employs small angle neutron scattering and molecular dynamics simulations, revealing that the most commonly used liquid metal solvents, EGaIn and Galinstan, are surprisingly structured with the formation of clusters ranging from 157 to 15.7 Å. Conversely, noneutectic liquid metal alloys of GaSn or GaIn at low solute concentrations of 1, 2, and 5 wt%, as well as pure Ga, do not exhibit these structures. Importantly, the eutectic alloys retain their structure even at elevated temperatures of 60 and 90 °C, highlighting that they are not just simple homogeneous fluids consisting of individual atoms. Understanding the complex soft structure of liquid alloys will assist in comprehending complex phenomena occurring within these fluids and contribute to deriving reaction mechanisms in the realm of synthesis and liquid metal-based catalysis.
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The most widely used method of platelet cryopreservation requires the addition of dimethyl sulfoxide (DMSO; Me2SO) as a cryoprotective agent (CPA) and pre-freeze removal of Me2SO before freezing to mitigate toxicity. However, alternative CPAs such as deep eutectic solvents (DES), which are less toxic could simplify this process. The aim of this study was to determine the effectiveness of a Proline-Glycerol (Prol-Gly 1:3) DES as a platelet CPA. Platelets were cryopreserved at -80 °C using 10 % Prol-Gly 1:3 (DES; n = 6), or in the absence of a cryoprotectant (no CPA; n = 6). Platelets were also cryopreserved according to the gold-standard blood-banking method using 5.5 % Me2SO (n = 6), with centrifugation and pre-freeze removal of the excess Me2SO. Platelet quality was assessed by flow cytometry and thromboelastography (TEG). Post-thaw recovery was similar between the three groups. The abundance of labile platelet glycoproteins GPIbα and GPVI were highest in the DES group, however, markers of activation (CD62P and annexin-V) were also higher in this group. In terms of function, the strength of the clot (maximum amplitude; TEG) and extent of clot retraction was better with DES platelets compared to no CPA, but lower than Me2SO platelets. DES provides a cryoprotective advantage to platelets when compared to no CPA. Importantly, when compared to Me2SO platelets, most quality parameters were similar in DES platelets. The major advantage with using a DES is biocompatibility, therefore it does not need to be removed prior to transfusion. This greatly simplifies the freezing and thawing process, avoiding the toxic effects of Me2SO.
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Plaquetas , Conservación de la Sangre , Criopreservación , Crioprotectores , Dimetilsulfóxido , Solventes , Crioprotectores/farmacología , Criopreservación/métodos , Humanos , Plaquetas/efectos de los fármacos , Plaquetas/metabolismo , Dimetilsulfóxido/farmacología , Dimetilsulfóxido/química , Solventes/química , Conservación de la Sangre/métodos , Glicerol/farmacología , Glicerol/química , Tromboelastografía , Prolina/farmacología , Prolina/químicaRESUMEN
Gel-based wound dressings have gained popularity within the healthcare industry for the prevention and treatment of bacterial and fungal infections. Gels based on deep eutectic solvents (DESs), known as eutectogels, provide a promising alternative to hydrogels as they are non-volatile and highly tunable and can solubilize therapeutic agents, including those insoluble in hydrogels. A choline chloride:glycerol-cellulose eutectogel was loaded with numerous antimicrobial agents including silver nanoparticles, black phosphorus nanoflakes, and commercially available pharmaceuticals (octenidine dihydrochloride, tetracycline hydrochloride, and fluconazole). The eutectogels caused >97% growth reduction in Gram-positive methicillin-resistant Staphylococcus aureus and Gram-negative Pseudomonas aeruginosa bacteria and the fungal species Candida albicans.
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Antiinfecciosos , Nanopartículas del Metal , Staphylococcus aureus Resistente a Meticilina , Solventes , Disolventes Eutécticos Profundos , Plata/farmacología , Antiinfecciosos/farmacología , HidrogelesRESUMEN
Lamellar unit cell reconstruction from neutron and X-ray diffraction data provides information about the disposition and position of molecules and molecular segments with respect to the bilayer. When supplemented with the judicious use of molecular deuteration, the technique probes the molecular interactions and conformations within the bilayer membrane and the water layer which constitute the crystallographic unit cell. The perspective is model independent, and potentially, with a higher degree of resolution than is available with other techniques. In the case of neutron diffraction the measurement consists of carefully normalised diffracted intensity under conditions of contrast variation of the water layer. The subsequent Fourier reconstruction of the unit cell is made using the phase information from variation of peak intensities with contrast. Although the phase problem is not as easily solved for the corresponding X-ray measurements, an intuitive approach can often suffice. Here we discuss the two complimentary techniques as probes of scattering length density profiles of a bilayer, and how such a perspective provides information about the location and orientation of molecules within or between lipid bilayers. Within the basic paradigm of lamellar phases this method has provided, for example, detailed insights into the location and interaction of cryoprotectants and stress proteins, of the mechanisms of actions of viral proteins, antimicrobial compounds and drugs, and the underlying structure of the stratum corneum. In this paper we review these techniques and provide examples of the systems that have been examined. We finish with a future outlook on the use of these techniques to improve our understanding of the interactions of membranes with biomolecules.
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Ionic liquids (ILs) are a diverse class of solvents which can be selected for task-specific properties, making them attractive alternatives to traditional solvents. To tailor ILs for specific biological applications, it is necessary to understand the structure-property relationships of ILs and their interactions with cells. Here, a selection of carboxylate anion-based ILs were investigated as cryoprotectants, which are compounds added to cells before freezing to mitigate lethal freezing damage. The cytotoxicity, cell permeability, thermal behavior, and cryoprotective efficacy of the ILs were assessed with two model mammalian cell lines. We found that the biophysical interactions, including permeability of the ILs, were influenced by considering the IL pair together, rather than as single species acting independently. All of the ILs tested had high cytotoxicity, but ethylammonium acetate demonstrated good cryoprotective efficacy for both cell types tested. These results demonstrate that despite toxicity, ILs may be suitable for certain biological applications. It also demonstrates that more research is required to understand the contribution of ion pairs to structure-property relationships and that knowing the behavior of a single ionic species will not necessarily predict its behavior as part of an IL.
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Líquidos Iónicos , Animales , Líquidos Iónicos/toxicidad , Solventes , Aniones , Iones , Criopreservación , MamíferosRESUMEN
Antimicrobial resistance is a pressing global health issue, with millions of lives at risk by 2050, necessitating the development of alternatives with broad-spectrum activity against pathogenic microbes. Antimicrobial peptides provide a promising solution by combating microbes, modulating immunity, and reducing resistance development through membrane and intracellular targeting. PuroA, a synthetic peptide derived from the tryptophan-rich domain of puroindoline A, exhibits potent antimicrobial activity against various pathogens, while the rationally designed P1 peptide demonstrates enhanced antimicrobial activity with its specific composition. This paper investigates the concentration-dependent effects of these cationic peptides on distinct types of vesicles representing strong-negative bacterial cell membranes (S-vesicles), weak-negative bacterial cell membranes (W-vesicles), and mammalian cell membranes (M-vesicles). To investigate the interactions between the peptides and vesicles, small-angle neutron scattering experiments were conducted. The cationic peptides, PuroA and P1, interact with S-vesicles through electrostatic interactions, leading to distinct effects. PuroA accumulates on the vesicle surface, increasing Rcore and Rtotal, aligning with the carpet model. P1 disrupts the vesicle structure at higher concentrations, consistent with the detergent model. Neither peptide significantly affects W-vesicles, emphasizing the role of charge. In uncharged M-vesicles, both peptides decrease Rcore and Rtotal and increase tshell, indicating peptide insertion and altered bilayer properties. These findings provide valuable insights into peptide-membrane interactions and their impact on vesicle structures. Furthermore, the implications of these findings extend to the potential development of innovative antimicrobial agents and drug delivery systems that specifically target bacterial and mammalian membranes. This research contributes to the advancement of understanding peptide-membrane interactions and lays the foundation for the design of approaches for targeting membranes in various biomedical applications.
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Antiinfecciosos , Péptidos Catiónicos Antimicrobianos , Animales , Péptidos Catiónicos Antimicrobianos/farmacología , Péptidos Catiónicos Antimicrobianos/química , Membrana Celular/química , Antiinfecciosos/química , Bacterias/metabolismo , Mamíferos/metabolismoRESUMEN
BACKGROUND: Systemic lupus erythematosus (SLE) is a chronic autoimmune disease characterized by multisystem inflammation. Medical management of SLE is based on reducing inflammation and tissue damage in the affected organs; however, medications used to treat SLE have been found to contribute to additional organ damage. Therefore, finding new ways to predict and prevent flares that require an inpatient (IP) stay or emergency department (ED) visit is critical for reducing the clinical and economic burden in patients with SLE. OBJECTIVE: To identify risk factors of SLE flares requiring an IP/ED visit among a Medicaid-insured population with SLE. METHODS: This retrospective study included patients from the Merative MarketScan Medicaid database (2013-2019). To capture patients at all stages of their SLE journey, all SLE claims for a patient were captured, and the index date was randomly selected among those claims that were at least 12 months after the first evidence of SLE. Patients were required to be continuously enrolled 1-year pre-index (year 1) and post-index (year 2). Demographics, clinical characteristics, and health care use and costs were measured in year 1, and flares requiring an IP/ED visit were identified in year 2 using the Garris algorithm. Multivariable logistic regression and classification and regression tree (CART) modeling were used to identify year 1 predictors and combination of factors, respectively, associated with flares-related IP/ED visits. RESULTS: Of the 8,083 patients included in the study, 37.6% of patients (n = 3,039) had a flare. Logistic regression identified ED visits in year 1 as one of the strongest predictors of flares-related IP/ED visits in year 2 (odds ratio = 2.19 [95% CI = 1.93-2.49]). SLE treatment progression to biologics (0.54 [0.42-0.70]) was the strongest predictor of decreased odds. Other strong predictors included other neurological disorders (1.63 [1.43-1.87]), Black race (1.49 [1.32-1.68]), chronic kidney disease/renal failure (1.35 [1.10-1.66]), and opioid use (1.30 [1.17-1.45]). CART modeling identified patients with an ED visit, an IP admission, and a diagnosis of Elixhauser Comorbidity Index-defined other neurological disorders in year 1 as having the highest probability of a flare-related IP/ED visit in year 2 (probability = 0.708), whereas patients without an ED visit had the lowest probability (probability = 0.185). CONCLUSIONS: Patients with the highest risk of a flare that required an IP/ED visit were those with a prior ED visit, IP admission, and other neurological disorders. Modeling also identified patients with prior opioid use, Black patients, and patients without SLE medications as subgroups with a high risk of a flare requiring an IP/ED visit.
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Lupus Eritematoso Sistémico , Enfermedades del Sistema Nervioso , Humanos , Estados Unidos , Estudios Retrospectivos , Pacientes Internos , Medicaid , Analgésicos Opioides/uso terapéutico , Lupus Eritematoso Sistémico/tratamiento farmacológico , Lupus Eritematoso Sistémico/epidemiología , Inflamación , Servicio de Urgencia en HospitalRESUMEN
Green fluorescent protein (GFP) and its variants are widely used in medical and biological research, especially acting as indicators of protein structural integrity, protein-protein interactions and as biosensors. This study employs superfolder GFP (sfGFP) to investigate the impact of varying alkyl chain length of 1-Cn-3-methylimidazolium chloride ionic liquid (IL) series ([Cnmim]Cl, n = 2, 4, 6, 8, 10, 12) on the protein fluorescence, structure, hydration, aggregation dynamics and crystallization behaviour. The results revealed a concentration-dependent decrease in the sfGFP chromophore fluorescence, particularly in long alkyl chain ILs ([C10mim]Cl and [C12mim]Cl). Tryptophan (Trp) fluorescence showed the quenching rate increased with longer alkyl chains indicating a nonpolar interaction between Trp57 and the alkyl chain. Secondary structural changes were observed at the high IL concentration of 1.5 M in [C10mim]Cl and [C12mim]Cl. Small-angle X-ray scattering (SAXS) indicated relatively stable protein sizes, but with IL aggregates present in [C10mim]Cl and [C12mim]Cl solutions. Dynamic light scattering (DLS) data showed increased protein size and aggregation with longer alkyl chain ILs. Notably, ILs and salts, excluding [C2mim]Cl, promoted sfGFP crystallization. This study emphasizes the influence of the cation alkyl chain length and concentration on protein stability and aggregation, providing insights into utilizing IL solvents for protein stabilization and crystallization purposes.
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Líquidos Iónicos , Proteínas Fluorescentes Verdes/genética , Líquidos Iónicos/química , Cristalización , Dispersión del Ángulo Pequeño , Difracción de Rayos XRESUMEN
BACKGROUND: Systemic lupus erythematosus (SLE) is a chronic autoimmune disease affecting as many as 322,000 people in the United States. Because of heterogeneity in both disease course and clinical manifestations, it is critical to identify a prevalent SLE population that includes patients with moderate or severe disease. Additionally, differences in the clinical and economic burden of SLE may exist across payer channels, yet to date this has not been reported in any previous studies. OBJECTIVE: To characterize the clinical and economic burden of SLE across disease severity and payer channels. METHODS: This retrospective study included patients from Merative MarketScan Commercial, Medicare Supplemental, and Medicaid databases from 2013 to 2020 (Commercial/Medicare) or 2013 to 2019 (Medicaid), with at least 1 inpatient or at least 2 outpatient SLE claims and no invalid steroid claims. The index date was a random SLE claim with at least 12 months of disease history. Patients were continuously enrolled 1 year pre-index (baseline) and 1 year post-index and classified with mild, moderate, or severe disease using a published algorithm. Baseline demographics, clinical characteristics, flares, and utilization/costs were compared across disease severity. RESULTS: 22,385 Commercial, 2,035 Medicare, and 8,083 Medicaid patients had SLE. Most Medicaid patients (51.1%) had severe disease. Comorbidity scores increased with disease severity (P < 0.001). 30.7% of Commercial, 34.1% Medicare, and 51.3% Medicaid patients had opioids, which increased with disease severity (P < 0.001). All-cause costs ranged from 1.8- to 2.3-fold for moderate vs mild and 4.2- to 6.5-fold for severe vs mild. Outpatient medical costs accounted for the highest proportion of all-cause costs, except Medicaid patients with severe disease, for whom inpatient costs were highest. Mean (SD) SLE-related annual costs were $23,030 (43,304) vs $1,738 (4,427) in severe vs mild for Commercial, $12,264 (31,896) vs $2,024 (4,998) for Medicare, and $7,572 (27,719) vs $787 (3,797) for Medicaid (P < 0.001). For patients with severe disease in Medicaid, 16.5% and 60.1% had inpatient and emergency department (ED) visits, respectively, vs 10.3% and 26.5% Commercial vs 10.6% and 24.6% Medicare. Mean [SD] flares per year in the baseline period increased from 2.5 [1.7] in mild to 4.6 [1.9] in severe for Commercial, 3.2 [1.9] to 5.0 [2.1] for Medicare, and 2.0 [1.6] to 4.5 [2.0] for Medicaid. CONCLUSIONS: Patients with severe SLE experienced more comorbidities, flares, and utilization/costs. Outpatient costs were the largest driver of all-cause costs for Commercial and Medicare (and Medicaid for mild to moderate SLE). Medicaid beneficiaries had the highest rate of severe SLE, highest use of ED and inpatient services, and highest oral corticosteroid and opioid use but the lowest utilization of disease-modifying treatments. Results demonstrate an unmet need in SLE treatment, especially among patients with moderate to severe disease or Medicaid coverage. DISCLOSURES: This study was funded by AstraZeneca. Drs Wu and Bryant are current employees of AstraZeneca and may own stock and/or options. At the time of the study, Ms Perry and Mr Tkacz were employed by IBM Watson Health, which received funding from AstraZeneca to conduct this study.
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Lupus Eritematoso Sistémico , Medicare , Humanos , Anciano , Estados Unidos , Estudios Retrospectivos , Lupus Eritematoso Sistémico/tratamiento farmacológico , Medicaid , Pacientes InternosRESUMEN
Deep eutectic solvents (DES) are tailorable non-aqueous solvents with promising properties for a range of applications, from industrial dissolution of plant products to biomedicine. They are mixtures of hydrogen bond donors and acceptors with low melting points that can be tailored to specific applications, and many support the self-assembly of amphiphilic molecules into lyotropic liquid crystal phases. Self-assembled lipid structures have potential for numerous applications, including drug delivery. These ordered structures can act as carriers, slow-release vehicles, or microreactors. Lipid self-assembly in non-aqueous solvents, such as deep eutectic solvents, is important for applications at extreme temperatures, or involving water-insoluble or water sensitive components. However, lipid self-assembly in these solvents remains largely unexplored. In this paper, we have examined the self-assembly of phytantriol, a non-ionic lipid, at 10 and 30 wt% in the deep eutectic solvent choline chloride:urea, with and without water. Self-assembly was assessed using small angle X-ray scattering and cross polarised optical microscopy at temperatures from 25-66 °C. We found that pure choline chloride:urea supports a Pn3m cubic phase similar to that formed in water. However, mixtures of the DES with water resulted in phytantriol forming an inverse hexagonal phase and influenced the phase transition temperatures. These results demonstrate that choline chloride:urea can support diverse phase behaviour, and also provides a mechanism for tailoring the phase for particular applications simply by controlling the amount of water in the solvent. In the future this could lead to methods of triggered release of drugs and biomolecules by the simple addition of water which could be critical for drug delivery applications.
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Colina , Urea , Urea/química , Colina/química , Disolventes Eutécticos Profundos , Solventes/química , Alcoholes Grasos/químicaRESUMEN
Metal-organic frameworks (MOFs) are currently under progressive development as a tool for non-viral biomolecule delivery. Biomolecules such as proteins, lipids, carbohydrates, and nucleic acids can be encapsulated in MOFs for therapeutic purposes. The favorable physicochemical properties of MOFs make them an attractive choice for delivering a wide range of biomolecules including nucleic acids. Herein, a green fluorescence protein (GFP)-expressing plasmid DNA (pDNA) is used as a representative of a biomolecule to encapsulate within a Zn-based metal-organic framework (MOF) called a zeolitic imidazolate framework (ZIF). The synthesized biocomposites are coated with positively charged amino acids (AA) to understand the effect of surface functionalization on the delivery of pDNA to prostate cancer (PC-3) cells. FTIR and zeta potential confirm the successful preparation of positively charged amino acid-functionalized derivatives of pDNA@ZIF (i.e., pDNA@ZIFAA). Moreover, XRD and SEM data show that the functionalized derivates retain the pristine crystallinity and morphology of pDNA@ZIF. The coated biocomposites provide enhanced uptake of genetic material by PC-3 human prostate cancer cells. The AA-modulated fine-tuning of the surface charge of biocomposites results in better interaction with the cell membrane and enhances cellular uptake. These results suggest that pDNA@ZIFAA can be a promising alternative tool for non-viral gene delivery.
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Estructuras Metalorgánicas , Neoplasias de la Próstata , Zeolitas , Humanos , Masculino , Aminoácidos/genética , Zeolitas/química , ADN/química , Plásmidos/genética , Compuestos Orgánicos/química , Estructuras Metalorgánicas/química , Neoplasias de la Próstata/genéticaRESUMEN
We report a new composite material consisting of silver nanoparticles decorated with three-dimensional molecular organic cages based on light-absorbing porphyrins. The porphyrin cages serve to both stabilize the particles and allow diffusion and trapping of small molecules close to the metallic surface. Combining these two photoactive components results in a Fano-resonant interaction between the porphyrin Soret band and the nanoparticle-localised surface-plasmon resonance. Time-resolved spectroscopy revealed the silver nanoparticles transfer up to 37 % of their excited-state energy to the stabilising layer of porphyrin cages. These unusual photophysics cause a 2-fold current increase in photoelectrochemical water-splitting measurements. The composite structure provides a compelling proof of concept for advanced photosensitiser systems with intrinsic porosity for photocatalytic and sensing applications.
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The maintenance of plasma membrane structure is vital for the viability of cells. Disruption of this structure can lead to cell death. One important example is the macroscopic phase separation observed during dehydration associated with desiccation and freezing, often leading to loss of permeability and cell death. It has previously been shown that the hybrid lipid 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) can act as a line-active component in ternary lipid systems, inhibiting macroscopic phase separation and stabilising membrane microdomains in lipid vesicles [1]. The domain size is found to decrease with increasing POPC concentration until complete mixing is observed. However, no such studies have been carried out at reduced hydration. To examine if this phase separation is unique to vesicles in excess water, we have conducted studies on several binary and ternary model membrane systems at both reduced hydration ("powder" type samples and oriented membrane stacks) and in excess water (supported lipid bilayers) at 0.2 mol fraction POPC, in the range where microdomain stabilisation is reported. Differential scanning calorimetry (DSC) and Fourier transform infrared spectroscopy (FTIR) are used to map phase transition temperatures, with X-ray and neutron scattering providing details of the changes in lipid packing and phase information within these boundaries. Atomic force microscopy (AFM) is used to image bilayers on a substrate in excess water. In all cases, macroscopic phase separation was observed rather than microdomain formation at this molar ratio. Thus POPC does not stabilise microdomains under these conditions, regardless of the type of model membrane, hydration or temperature. Thus we conclude that the driving force for separation under these conditions overcomes any linactant effects of the hybrid lipid.
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Membrana Dobles de Lípidos , Fosfatidilcolinas , Fosfatidilcolinas/química , Membrana Dobles de Lípidos/química , Transición de Fase , AguaRESUMEN
Differential Dynamic Microscopy (DDM) is a relatively new technique which measures the dynamics of suspended particles using a dynamic light scattering formalism. Videos are recorded using standard light microscopy at moderate frame rates, and fluctuations in pixel intensity are measured as a function of time. As only pixel intensity is analysed, it is not necessary to resolve individual particles. This allows for low magnifications and wide fields of view, and therefore dynamics can be measured on tens of thousands of scattering objects, providing robust statistics. A decade ago the technique was successfully applied to measure bacterial motility. Since then, it has been applied to a range of motile systems, but has not yet reached the wider biological community. This perspective reviews the work done so far, and provides the basic background to enable the broader application of this promising technique.
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Microscopía , Dispersión Dinámica de Luz , Microscopía/métodosRESUMEN
Nanomaterials have the potential to transform biological and biomedical research, with applications ranging from drug delivery and diagnostics to targeted interference of specific biological processes. Most existing research is aimed at developing nanomaterials for specific tasks such as enhanced biocellular internalization. However, fundamental aspects of the interactions between nanomaterials and biological systems, in particular, membranes, remain poorly understood. In this study, we provide detailed insights into the molecular mechanisms governing the interaction and evolution of one of the most common synthetic nanomaterials in contact with model phospholipid membranes. Using a combination of atomic force microscopy (AFM) and molecular dynamics (MD) simulations, we elucidate the precise mechanisms by which citrate-capped 5 nm gold nanoparticles (AuNPs) interact with supported lipid bilayers (SLBs) of pure fluid (DOPC) and pure gel-phase (DPPC) phospholipids. On fluid-phase DOPC membranes, the AuNPs adsorb and are progressively internalized as the citrate capping of the NPs is displaced by the surrounding lipids. AuNPs also interact with gel-phase DPPC membranes where they partially embed into the outer leaflet, locally disturbing the lipid organization. In both systems, the AuNPs cause holistic perturbations throughout the bilayers. AFM shows that the lateral diffusion of the particles is several orders of magnitude smaller than that of the lipid molecules, which creates some temporary scarring of the membrane surface. Our results reveal how functionalized AuNPs interact with differing biological membranes with mechanisms that could also have implications for cooperative membrane effects with other molecules.
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Oro , Nanopartículas del Metal , Membrana Dobles de Lípidos , Ácido Cítrico , Fosfolípidos , Microscopía de Fuerza AtómicaRESUMEN
Cryopreservation has facilitated numerous breakthroughs including assisted reproductive technology, stem cell therapies, and species preservation. Successful cryopreservation requires the addition of cryoprotective agents to protect against freezing damage and dehydration. For decades, cryopreservation has largely relied on the same two primary agents: dimethylsulfoxide and glycerol. However, both of these are toxic which limits their use for cells destined for clinical applications. Furthermore, these two agents are ineffective for hundreds of cell types, and organ and tissue preservation has not been achieved. The research presented here shows that deep eutectic solvents can be used as cryoprotectants. Six deep eutectic solvents were explored for their cryoprotective capacity towards mammalian cells. The solvents were tested for their thermal properties, including glass transitions, toxicity, and permeability into mammalian cells. A deep eutectic solvent made from proline and glycerol was an effective cryoprotective agent for all four cell types tested, even with extended incubation prior to freezing. This deep eutectic solvent was more effective and less toxic than its individual components, highlighting the importance of multi-component systems. Cells were characterised post-thawing using atomic force microscopy and confocal microscopy. Molecular dynamics simulations support the biophysical parameters obtained by experimentation. This is one of the first times that this class of solvents has been systematically tested for cryopreservation of mammalian cells and as such this research opens the way for the development of potentially thousands of new cryoprotective agents that can be tailored to specific cell types. The demonstrated capacity of cells to be incubated with the deep eutectic solvent at 37 °C for hours prior to freezing without significant loss of viability is a major step toward the storage of organs and tissues.
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Crioprotectores , Disolventes Eutécticos Profundos , Animales , Criopreservación , Crioprotectores/farmacología , Glicerol/farmacología , Mamíferos , SolventesRESUMEN
Ultrasmall metal nanoclusters (NCs) are employed in an array of diagnostic and therapeutic applications due to their tunable photoluminescence, high biocompatibility, polyvalent effect, ease of modification, and photothermal stability. However, gold nanoclusters' (AuNCs') intrinsically antimicrobial properties remain poorly explored and are not well understood. Here, we share an insight into the antimicrobial action of atomically precise AuNCs based on their ability to passively translocate across the bacterial membrane. Functionalized by a hydrophilic modified-bidentate sulfobetaine zwitterionic molecule (AuNC-ZwBuEt) or a more hydrophobic monodentate-thiolate, mercaptohexanoic acid (AuNC-MHA) molecule, 2 nm AuNCs were lethal to both Gram-negative Pseudomonas aeruginosa and Gram-positive Staphylococcus aureus bacteria. The bactericidal efficiency was found to be bacterial strain-, time-, and concentration-dependent. The direct visualizations of the translocation of AuNCs and AuNC-cell and subcellular interactions were investigated using cryo-soft X-ray nano-tomography, transmission electron microscopy (TEM), and scanning TEM energy-dispersive spectroscopy analyses. AuNC-MHA were identified in the bacterial cytoplasm within 30 min, without evidence of the loss of membrane integrity. It is proposed that the bactericidal effect of AuNCs is attributed to their size, which allows for efficient energy-independent translocation across the cell membrane. The internalization of both AuNCs caused massive internal damage to the cells, including collapsed subcellular structures and altered cell morphology, leading to the eventual loss of cellular integrity.
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Antiinfecciosos , Nanopartículas del Metal , Antibacterianos/farmacología , Bacterias , Oro/química , Oro/farmacología , Nanopartículas del Metal/química , Pseudomonas aeruginosa , Staphylococcus aureusRESUMEN
Intensity-correlation measurements allow access to nanostructural information on a range of ordered and disordered materials beyond traditional pair-correlation methods. In real space, this information can be expressed in terms of a pair-angle distribution function (PADF) which encodes three- and four-body distances and angles. To date, correlation-based techniques have not been applied to the analysis of microstructural effects, such as preferred orientation, which are typically investigated by texture analysis. Preferred orientation is regarded as a potential source of error in intensity-correlation experiments and complicates interpretation of the results. Here, the theory of preferred orientation in intensity-correlation techniques is developed, connecting it to the established theory of texture analysis. The preferred-orientation effect is found to scale with the number of crystalline domains in the beam, surpassing the nanostructural signal when the number of domains becomes large. Experimental demonstrations are presented of the orientation-dominant and nanostructure-dominant cases using PADF analysis. The results show that even minor deviations from uniform orientation produce the strongest angular correlation signals when the number of crystalline domains in the beam is large.