Dipalmitoylphosphatidylglycerol Research Articles

Electrochemical Biosensors Composed of Polyethylenimine (PEI) and Graphene Derivatives for Rapid Detection of Alzheimer’s Disease

Graphene materials can be used as nanofillers in polymers to take advantage of graphene's suitable properties. Graphene-PEI composites are materials that combine the unique properties of graphene and PEI. These composites are made by combining graphene with PEI through physical or chemical method. The resulting material is characterized by enhanced mechanical properties, improved electrical conductivity and increased stability. By incorporating graphene into biosensor platforms, it can improve overall’s performance, including sensitivity, specificity and response. In this work, the detection of anti-βA40 biomarker was tested. Beta-amyloid peptide (βA1-40) was incorporated into liposomes of Dipalmitoyl Phosphatidylglycerol (DPPG) and was immobilized using Layer-by-Layer self-assembling technique on PEI_graphene derivatives composites. To obtain these composites it was utilized graphene oxide (GO) from Hummers’ method, it’s thermally reduced form (rGO) and also electrochemically exfoliated graphene (EEG). MilliQ water was added to the PEI to obtain a concentration of 2mg/mL and stirred for 30 min at room temperature for complete dissolution of the PEI. Then EEG, GO and rGO were added to PEI solution and subjected to ultrasonication in a Hielscher UP400ST sonicator, using the H14 probe for 30 minutes to obtain a homogeneous and stable dispersion.Carbon microelectrodes modified with PEI-graphene derivatives composites and (DPPG +Aβ-40) were used as biosensors platforms to detect anti-Aβ40 related to Alzheimer's disease. The assembly of the biosensors is shown in Figure 1. Cyclic Voltammetry was performed in three cycles, at a potential of -0.6 V to 0.6 V and scan rate of 0.05 V s-1 to evaluate the sensory response. The electrochemical measurements were performed with saline phosphate buffer solution (PBS). The morphology of PEI and PEI_graphene oxide can be seen in Figure 2. The large surface area of graphene allows for increased binding sites for bioreceptors, which may favor the adsorption of Aβ-40 peptide, improving signal transduction and sensitivity of the biosensor. The wettability of the composites was obtained through measurements of the static contact angle using the Ramé-Hart Contact Angle Goniometer (model 100-25-A). Figure 3 presents the contact angle for a film layer of the composites. All PEI_graphene derivative composites showed hydrophilic behavior (θ < 90°), PEI_EEG composite stood out as less hydrophilic (θ = 68°) among all composites. Hydrophilicity is essential in biosensors because it affects the interactions between the biological recognition elements and the target analytes, as well as the overall stability and reliability of the biosensor. Figure 4 presents the calibration curves using the normalized area obtained from cyclic voltammetry of the biosensors manufactured with 2 bilayers (PEI_graphene derivative/DPPG+Aβ40) and anti-Aβ40 biomarker in concentrations ranging from 1 ng mL to 5 µg mL-1. Among the tested PEI_graphene derivatives composites, the ones with EEG and RGO stood out in relation to pure PEI. Table 1 presents the adjusted R-square of the data in Figure 4. All composites, except PEI_GO, showed excellent linear fit, adjusted R-square close to 1 in the detection range between 1 and 1000 ng mL-1. The PEI_graphene oxide composite can be used in biosensors to enable rapid and efficient transduction of the binding signal, leading to improved detection performance. Figure 1

Effect of Ropivacaine Hydrochloride Nanoliposomes on Delayed Cerebral Vasospasm After Subarachnoid Hemorrhage in Rabbits

This research was aimed to investigate the effect of ropivacaine hydrochloride (RP-HCl) nanoliposome (NLP) on delayed cerebral vasospasm (DCVS) after subarachnoid hemorrhage (SAH) in rabbits. RP-HCl liposomes were prepared with RP-HCl, chloroform-ether, soybean phospholipid, dipalmitoyl phosphatidyl glycerol, and L-lysine 5% glucose solution by secondary emulsification, and blank NLP (without RP-HCl) was constructed at the same time. Additionally, an SAH model was also established by secondary blood injection into the large occipital pool. Forty New Zealand rabbits were randomly assigned into sham operation group (the injected liquid was 0.9% sodium chloride injection), control (Ctrl) group (the injected liquid was autologous blood and sodium chloride injection in sequence), no-load group (the injected liquid was autologous blood and blank liposomes in sequence), drug-loaded group (the injected liquid was autologous blood and RP-HCl NLP in sequence), and conventional group (the injected liquid was sodium chloride injection and ropivacaine in sequence), with eight rabbits in each group. The effects of RP-HCl NLP were analyzed through the changes in mean blood flow velocity (Vm), peak systolic blood flow velocity (Vp), neuron-specific enolase (NSE), S100B, diameter and apoptosis of the basilar artery (BA), vascular endothelial cell (EC) (VEC) endothelin-1 (ET-1), and endothelial nitric oxide synthase (eNOS). Results: the average particle size of RP-HCl NLP was 17.73±3.22 μm, and the average encapsulation efficiency was 86.40%. Relative to those before modeling, the Vm, Vp, NSE, and S100B levels in the BA of the five groups were increased on the 1st, 5th, and 10th days after modeling (P &lt;0.05). In contrast to sham group, Vm, Vp, NSE, S100B, ET-1, and the apoptosis index were increased on the 1st, 5th, and 10th days after modeling in the other four groups, while the caliber and eNOS were decreased (P &lt;0.05). The Vm, Vp, NSE, S100B, ET-1, and apoptosis index on the 1st, 5th, and 10th days after modeling in Ctrl and no-load groups were increased versus drug-loaded group, while the pipe diameter and eNOS were decreased (P &lt;0.05). In conclusions, RP-HCl NLP can regulate the balance between ET-1 and eNOS, enhance neurological function, alleviate EC apoptosis, and reduce cerebral blood flow velocity to relieve DCVS after SAH.

Interspecies Variation Affects Islet Amyloid Polypeptide Membrane Binding.

The aggregation of islet amyloid polypeptide (IAPP) is associated with β-cell dysfunction in type 2 diabetes (T2D) in humans. One possible mechanism of toxicity is the interaction of IAPP oligomers with lipid membranes to disrupt the bilayer integrity and/or homeostasis of the cell. Amino acid sequence variations of IAPPs between species can greatly decrease their propensity for aggregation. For example, human IAPP is toxic to β-cells, but rat and pig IAPP are not. However, it is not clear how these differences affect membrane association. Using native mass spectrometry with lipid nanodiscs, we explored the differences in the association of human, rat, and pig IAPP with lipid bilayers. We discovered that human and rat IAPP bound nanodiscs with anionic dipalmitoyl-phosphatidylglycerol (DPPG) lipids, but pig IAPP did not. Furthermore, human and rat IAPP interacted differently with the membrane. Human IAPP show potential tetramer complexes, but rat IAPP associated with the membrane sequentially. Thus, overall IAPP-bilayer interactions are not necessarily related to disease, but small differences in oligomeric behavior at the membrane may instead play a role.

The interaction of tryptophan enantiomers with model membranes is modulated by polar head type and physical state of phospholipids

The mutual influence of chiral bioactive molecules and supramolecular assemblies is currently being studied in many research fields, including medical-pharmaceutical applications. Model membranes of phospholipids, such as the zwitterionic dipalmitoylphosphatidylcholine (DPPC) and the anionic dipalmitoylphosphatidylglycerol (DPPG), interact with a variety of chiral compounds that include amino acids. In this work, the interaction of tryptophan enantiomers, L-Trp and D-Trp, on DPPC and DPPG bilayers was investigated by using differential scanning calorimetry, attenuated total reflectance-Fourier transform infrared and spin-label electron spin resonance spectroscopies as well as molecular docking simulations. The results show that Trp enantiomers slightly perturb the bilayer thermotropic phase transitions. For both membranes, O atoms in the carbonyl groups have a propensity to act as acceptors of a (weak) hydrogen bond. The Trp chiral forms also promote formation of hydrogen bonds and/or hydration in the PO2– moiety of the phosphate group, especially for the DPPC bilayer. In contrast, they interact more closely with the glycerol group of DPPG polar head. Only for DPPC bilayers, both enantiomers increase the packing of the first hydrocarbon chain segments for temperatures through the gel state, whereas they do not affect the lipid chain order and mobility in the fluid state. The results are consistent with a Trp association in the upper region of the bilayers without permeation in the innermost hydrophobic region. The findings suggest that neutral and anionic lipid bilayers are differently sensitive to amino acid chirality.

Effect of partial O-methylation in dehydrodieugenol on its antitrypanosomal activity - correlation with the toxicity using cell membrane models

Biseugenol (1), a neolignan with antiprotozoal activity against Trypanosoma cruzi, was partially methylated, and the compound obtained – methyl biseugenol (2) – had its activity evaluated against the extracellular (trypomastigotes) and intracellular (amastigotes) forms of T. cruzi. It was observed that both compounds 1 and 2 exhibited similar effects against trypomastigotes (IC50 of 11.7 and 16.2 μM, respectively), whereas compound 2 displayed higher activity against amastigotes (IC50 = 8.2 μM) in comparison with biseugenol (IC50 = 15.4 μM). Additionally, reduced toxicity against NCTC cells for compound 2 was observed (CC50 > 200 μM), differently from compound 1 with CC50 = 58.0 μM. Aiming to understand better the molecular mechanism of the biological action of compound 2, the prodrug was incorporated into cellular membrane models constituted of Langmuir monolayers of the lipids dipalmitoylphosphatidylcholine (DPPC), dipalmitoylphosphatidylethanolamine (DPPE), dipalmitoylphosphatidylserine (DPPS), and dipalmitoylphosphatidylglycerol (DPPG). The lipid-drug interaction was inferred through tensiometry, surface potential, infrared spectroscopy (PM-IRRAS), and Brewster angle microscopy (BAM). The prodrug expanded DPPC and DPPG monolayers and condensed DPPE ones, as well as presented characteristic behaviors regarding the chemical structure of the lipid considering expansion-compression curves, surface potential-area isotherms, and stability of previously compressed monolayers to relevant-biological surface pressures. PM-IRRAS indicated a molecular disorder for DPPC and DPPS alkyl chains in the presence of the drug. BAM revealed the presence of domains in the DPPG and DPPE monolayers, which was probably induced by the prodrug. These data suggest, in general, that the lipid composition modulates the interaction of compound 2, whose results are expected to correlate to its trypanocidal activity, which involves the plasma membrane of T. cruzi as the primary target, i.e., the first barrier that the compound should encounter to interact with the microorganism.

Understanding the Effects of Cholesterol and Ethanol on the Stability and Morphology of Bicellar Mixtures

Elucidating the stability and morphology of bicellar mixtures is of great importance in the biomedical field. In this report, self-assembly cholesterol (CHOL)-containing bicellar mixtures were investigated, that were composed of long chain hydrogenated soybean phospholipids (HSPC), short chain 1, 2-diheptanoyl-sn-glycero-3-phosphocholine (DHPC) phospholipids, negatively charged dipalmitoyl phosphatidylglycerol (1, 2-dipalmitoyl-sn-glycero-3-phospho-(1'-rac-glycerol) (DPPG), and polyethylene glycol conjugated (1, 2-dimyristoyl-sn-glycero-3-phosphoethano-lamine-N-[methoxy (polyethylene glycol)-2000] (PEG2000-DSPE) lipid. Analytical techniques like; Dynamic Light Scattering (DLS), Small Angle X-Ray Scattering (SAXS), and Transmission Electron Microscopy (TEM) were used to investigate the stability and morphology changes of these bicellar mixtures. The effects of CHOL and ethanol on the morphology and stability of these bicellar mixtures under 50ºC were also studied. The transition happened from nanodiscs to nanovesicles when the CHOL molar ratio was 40% in pure water. However, adding ethanol could improve CHOL solubility when the ethanol weight ratio was 20 wt% (ethanol/lipid) that favored nanodiscs-nanovesicle transition. Additionally, the ammonium sulfate maybe another factor that could affect the morphology of these bicellar mixtures form high order d-spacing structure.

Cellular Localization, Aggregation, and Cytotoxicity of Bicelle-Quantum Dot Nanocomposites

Bicelles are discoidal lipid nanoparticles (LNPs) in which the planar bilayer and curved rim are, respectively, composed of long- and short-chain lipids. Bicellar LNPs have a hydrophobic core, allowing hydrophobic molecules and large molecular complexes such as quantum dots (QDs) to be encapsulated. In this study, CdSe/ZnS QDs were encapsulated in bicelles made of dipalmitoyl phosphatidylcholine, dihexanoyl phosphatidylcholine, dipalmitoyl phosphatidylglycerol, and distearoyl phosphatidylethanolamine conjugated with polyethylene glycerol amine 2000 to form a well-defined bicelle-QD nanocomplex (known as NANO2-QD or bicelle-QD). The bicelle-QD was then incubated with Hek293t cells and HeLa cells for different periods of time to determine changes in their cellular localization. Bicelle-QDs readily penetrated Hek293t cell membranes within 15 min of incubation, localized to the cytoplasm, and associated with mitochondria and intracellular vesicles. After 1 h, the bicelle-QDs enter the cell nucleus. Large aggregates form throughout the cell after 2 h and QDs are nearly absent from the nucleus by 4 h. Previous reports have demonstrated that CdSe/ZnS QDs can be toxic to cells, and we have found that encapsulating QDs in bicelles can attenuate but did not eliminate cytotoxicity. The present research outcome demonstrates the time-resolved pathway of bicelle-encapsulated QDs in Hek293t cells, morphological evolution in cells over time, and cytotoxicity of the bicelle-QDs, providing important insight into the potential application of the nanocomplex for cellular imaging.

How do antimicrobial peptides disrupt the lipopolysaccharide membrane leaflet of Gram-negative bacteria?

HypothesisIt is widely regarded that antimicrobial peptides (AMPs) kill bacteria by physically disrupting microbial membranes and causing cytoplasmic leakage, but it remains unclear how AMPs disrupt the outer membrane (OM) of Gram-negative bacteria (GNB) and then compromise the inner membrane. We hypothesise that different AMPs impose different structural disruptions, with direct implications to their antimicrobial efficacies. ExperimentsThe antimicrobial activities of three typical AMPs, including the designed short AMP, G3, and two natural AMPs, melittin and LL37, against E. coli and their haemolytic activities were studied. Lipopolysaccharide (LPS) and anionic di-palmitoyl phosphatidyl glycerol (DPPG) monolayer models were constructed to mimic the outer membrane and inner membrane leaflets of Gram-negative bacteria. The binding and penetration of AMPs to the model lipid monolayers were systematically studied by neutron reflection via multiple H/D contrast variations. FindingG3 has relatively high antimicrobial activity, low cytotoxicity, and high proteolytic stability, whilst melittin has significant haemolysis and LL37 has weaker antimicrobial activity. G3 could rapidly lyse LPS and DPPG monolayers within 10–20 min. In contrast, melittin was highly active against the LPS membrane, but the dynamic process lasted up to 80 min, with excessive stacking in the OM. LL37 caused rather weak destruction to LPS and DPPG monolayers, leading to massive adsorption on the membrane surface without penetrating the lipid tail region. These findings demonstrate that the rationally designed AMP G3 was well optimised to impose most effective destruction to bacterial membranes, consistent with its highest bactericidal activity. These different interfacial structural features associated with AMP binding shed light on the future development of active and biocompatible AMPs for infection and wound treatments.

Pulmonary Administration of the Liposome-Based Adjuvant CAF01: Effect of Surface Charge on Mucosal Adjuvant Function

Mucosal surfaces of the lungs represent a major site of entry for airborne pathogens, and pulmonary administration of vaccines is an attractive strategy to induce protective mucosal immunity in the airways. Recently, we demonstrated the potential of pulmonary vaccination with the tuberculosis subunit antigen H56 adjuvanted with the cationic liposomal adjuvant formulation CAF01, which consists of the cationic lipid dimethyldioctadecylammonium (DDA) bromide and the synthetic cord factor trehalose-6,6'-dibehenate. However, the cationic charge of DDA represents a major safety challenge. Hence, replacing DDA with a safer zwitterionic or anionic phospholipid is an attractive approach to improve vaccine safety, but the effect of liposomal surface charge on the induction of mucosal immunity after airway immunization is poorly understood. Here, we investigated the effect of surface charge by replacing the cationic DDA component of CAF01 with zwitterionic dipalmitoylphosphatidylcholine (DPPC) or anionic dipalmitoylphosphatidylglycerol (DPPG), and we show that charge modification enhances antigen-specific pulmonary T-cell responses against co-formulated H56. We systematically replaced DDA with either DPPC or DPPG and found that these modifications resulted in colloidally stable liposomes that have similar size and morphology to unmodified CAF01. DPPC- or DPPG-modified CAF01 displayed surface charge-dependent protein adsorption and induced slightly higher follicular helper T cells and germinal center B cells in the lung-draining lymph nodes than unmodified CAF01. In addition, modified CAF01 induced significantly higher levels of H56-specific Th17 cells and polyfunctional CD4+ T cells in the lungs, as compared to unmodified CAF01. However, the strong H56-specific humoral responses induced by CAF01 in the lungs and spleen were not influenced by surface charge. Hence, these results provide insights into the importance of surface charge for liposomal adjuvant function and can also guide the design of safe pulmonary subunit vaccines against other mucosal pathogens.

Ruscogenin interacts with DPPC and DPPG model membranes and increases the membrane fluidity: FTIR and DSC studies

Ruscogenin, a kind of steroid saponin, has been shown to have significant anti-oxidant, anti-inflammatory, and anti-thrombotic characteristics. Furthermore, it has the potential to be employed as a medicinal medication to treat a variety of acute and chronic disorders. The interaction of a drug molecule with cell membranes can help to elucidate its system-wide protective and therapeutic effects, and it's also important for its pharmacological activity. The molecular mechanism by which ruscogenin affects membrane architecture is still a mystery. Ruscogenin's interaction with zwitterionic dipalmitoyl phosphatidylcholine (DPPC) and anionic dipalmitoyl phosphatidylglycerol (DPPG) multilamellar vesicles (MLVs) was studied utilizing two non-invasive approaches, including: Fourier Transform Infrared (FTIR) spectroscopy and Differential Scanning Calorimetry. Ruscogenin caused considerable alterations in the phase transition profile, order, dynamics and hydration state of head groups and glycerol backbone of DPPC and DPPG MLVs at all concentrations. The DSC results indicated that the presence of ruscogenin decreased the main phase transition temperature (Tm) and enthalpy (ΔH) values of both membranes and increased half height width of the main transition (ΔT1/2). The FTIR results demonstrated that all concentrations (1, 3, 6, 9, 15, 24 and 30 mol percent) of ruscogenin disordered the DPPC MLVs both in the gel and liquid crystalline phases while it increased the order of DPPG MLVs in the liquid crystalline phase. Moreover, ruscogenin caused an increase in the dynamics of DPPC and DPPG MLVs in both phases. Additionally, it enhanced the hydration of the head groups of lipids and the surrounding water molecules implying ruscogenin to interact strongly with both zwitterionic and charged model membranes.

Incorporation of dehydrodieugenol, a neolignan isolated from Nectandra leucantha (Lauraceae), in lipid Langmuir monolayers as biomembrane models

Dehydrodieugenol, a neolignan isolated from the Brazilian plant Nectandra leucantha (Lauraceae) with reported antiprotozoal and anticancer activity, was incorporated in Langmuir monolayers of selected lipids as cell membrane models, aiming to comprehend its action mechanism at the molecular level. The interaction of this compound with the lipids dipalmitoylphosphatidylcholine (DPPC), dipalmitoylphosphatidylethanolamine (DPPE), dipalmitoylphosphatidylserine (DPPS), and dipalmitoylphosphatidylglycerol (DPPG) was inferred through tensiometry, infrared spectroscopy, and Brewster angle microscopy. The interactions had different effects depending on the chemical nature of the lipid polar head, with expansion for DPPC monolayers, condensation for DPPE, and expansion (at low surface pressures) followed by the overlap of the isotherms (at high surface pressure values) for DPPS and DPPG. Effects caused by dehydrodieugenol in the negatively charged lipids were distinctive, which was also reflected in the hysteresis assays, surface potential-area isotherms, and rheological measurements. Infrared spectroscopy indicated that the drug interaction with the monolayer affects not only the polar groups, but also the acyl lipid chains for all lipids. These results pointed to the fact that the interaction of the drug with lipid monolayers at the air-water interface is modulated by the lipid composition, mainly considering the polar head of the lipids, as well as the hydrophobicity of the lipids and the drug. As negatively charged lipids pointed to distinctive interaction, we believe this can be related to the antiprotozoal and anticancer properties of the compound.

The influence of a biomimetic pulmonary surfactant modification on the in vivo fate of nanoparticles in the lung

Direct biomimetic modification of nanoparticles (NPs) with endogenous surfactants is helpful to improve the biocompatibility of NPs and avoid damage to the physiological function of the lung. Therefore, the objective of this study is to investigate the influence of biomimetic endogenous pulmonary surfactant phospholipid modification on the in vivo fate of NPs after lung delivery. Here, two neutral phospholipids (dipalmitoylphosphatidylcholine (DPPC), dipalmitoylphosphatidylamine (DPPE)) and two negatively charged phospholipids (dipalmitoylphosphatidylglycerol (DPPG), dipalmitoylphosphatidylserine (DPPS)) were selected to modify paclitaxel (PTX)-loaded PLGA NPs with different molar ratio. DPPC, DPPE, and DPPG improved mucoadhesion, in contrast, DPPS improved the mucus permeability of the NPs. Neutral DPPC and DPPE reduced, but negatively charged DPPS and DPPG increased the uptake by alveolar macrophages, all types of phospholipid increased the uptake by lung epithelial cells and increased PTX retention in the whole lung. Whereas, DPPC, DPPE, and DPPG promoted PTX retention in bronchoalveolar lavage fluid (BALF), while DPPS promoted PTX absorption in the lung tissue. Only DPPS-PLGA (1:1) NPs remarkably increased PTX systemic exposure. A good correlation between PTX percentage in the supernatant of BALF and PTX concentration in plasma was established, implying PTX entered the system circulation mainly in molecular form. Phospholipid modification had no effect on extrapulmonary organ distribution of PTX. Taken together, our study disclosed that different phospholipid modification can endow the NPs mucoadhesive or mucus penetration and cellular uptake properties, with tunable retention in BALF and absorption in the lung tissue, providing the scientific background for translational nanocarrier design for inhalation as required. Statement of significanceInhaled nanomedicines will inevitably interact with pulmonary surfactant and form “surfactant corona”. However, the contribution of individual pulmonary surfactant phospholipid on the in vivo fate of nanomedicines is still unclear. In this regard, the most abundant pulmonary surfactant phospholipid dipalmitoylphosphatidylcholine and dipalmitoylphosphatidylamine, and dipalmitoylphosphatidylglycerol and dipalmitoylphosphatidylserine were selected to modify the paclitaxel loaded PLGA nanoparticles and the effect of these pulmonary surfactant phospholipids on their in vivo fate was investigated. It was demonstrated that different phospholipid modification can endow the nanoparticles mucoadhesive or mucus penetration properties, with tunable retention in bronchoalveolar lavage fluid, alveolar macrophages uptake and absorption in the lung tissue. The present study provided a comprehensive understanding for the role of pulmonary surfactant phospholipid on inhaled nanomedicines.

Alzheimer's disease diagnosis based on detection of autoantibodies against Aβ using Aβ40 peptide in liposomes

BackgroundAlzheimer’s disease (AD) is the most common form of dementia and affect more than 50 million people worldwide. Thus, there is a high demand by non-invasive methods for an early diagnosis. This work explores the AD diagnostic using the amyloid beta 1–40 (Aβ40) peptide encapsulated into dipalmitoyl phosphatidyl glycerol (DPPG) liposomes and immobilized on polyethylene imine previously deposited on screen-printed carbon electrodes to detect autoantibodies against Aβ40, a potential biomarker found in plasma samples. MethodsThe immunosensor assembly was accompanied by atomic force microscopy (AFM) images that showed globular aggregates from 20 to 200 nm corresponding liposomes and by cyclic voltammetry (CV) through increase of the voltammogram area each material deposited. After building the immunosensor, when it was exposed to antibody anti-Aβ40, there was an increase in film roughness of approximately 9 nm, indicating the formation of the immunocomplex. ResultsIn the detection by CV, the presence of specific antibody, in the range of 0.1 to 10 μg/ml, resulted in an increase in the voltammograms area and current in 0.45 V reaching 3.2 µA.V and 5.7 μA, respectively, in comparison with the control system, which remained almost unchanged from 0.1 μg/ml. In patient samples, both cerebrospinal fluid (CSF) and plasma, was possible separated among positive and negative samples for AD using CV profile and area, with a difference of 0.1 μA.V from the upper error bar of healthy samples for CSF sample and 0.6 μA.V for plasma sample. ConclusionsThese results showed the feasibility of the method employed for the non-invasive diagnostic of Alzheimer’s disease detecting natural autoantibodies that circulate in plasma through a simple and easy-to-interpret method.

Changes Experienced by Low-Concentration Lipid Bicelles as a Function of Temperature.

Differential scanning calorimetry (DSC) of dipalmitoyl phosphatidylcholine (DPPC), dihexanoyl phosphatidylcholine, and dipalmitoyl phosphatidylglycerol bicelles reveals two endothermic peaks. Based on analysis of small angle neutron scattering and small angle X-ray scattering data, the two DSC peaks are associated with the melting of DPPC and a change in bicellar morphology─namely, either bicelle-to-spherical vesicle or oblate-to-spherical vesicle. The reversibility of the two structural transformations was examined by DSC and found to be consistent with the corresponding small angle scattering data. However, the peak that is not associated with the melting of DPPC does not correspond to any structural transformation for bicelles containing distearoyl phosphatidylethanolamine conjugated with polyethylene glycol. Based on complementary experimental data, we conclude that membrane flexibility, lipid miscibility, and differential solubility between the long- and short-chain lipids in water are important parameters controlling the reversibility of morphologies experienced by the bicelles.

Penicillin-binding proteins (PBPs) determine antibiotic action in Langmuir monolayers as nanoarchitectonics mimetic membranes of methicillin-resistant Staphylococcus aureus

The membrane of methicillin-resistant Staphylococcus aureus (MRSA) contains penicillin-binding proteins (PBPs) in the phospholipidic bilayer, with the protein PBP2a being linked with the resistance mechanism. In this work we confirm the role of PBP2a with molecular-level information obtained with Langmuir monolayers as cell membrane models. The MRSA cell membrane was mimicked with a mixed monolayer of dipalmitoyl phosphatidyl glycerol (DPPG) and cardiolipin (CL), also incorporating PBP2a. The surface pressure-area isotherms and the Brewster angle microscopy (BAM) images for these mixed monolayers were significantly affected by the antibiotic meropenem, which is PBP2a inhibitor. The meropenem effects were associated with the presence of PBP2a, as they were absent in the Langmuir monolayers without PBP2a. The relevance of PBP2a was confirmed with results where the antibiotic methicillin, known to be unsuitable to kill MRSA, had the same effects on mixed DPPG/CL and DPPG/CL-PBP2a monolayers since it prevented PBP2a from incorporating in the monolayer. The biological implication of the findings presented here is that a successful antibiotic against MRSA should be able to interact with PBP2a, but in the membrane.

Restriction‐In‐Motion of Surface Ligands Enhances Photoluminescence of Quantum Dots—Experiment and Theory

AbstractThe relationship between emission and ligand restriction of a series of ZnSe/ZnS quantum dots (QDs) encapsulated in nanoparticles is investigated systematically via experiments and quantum theory. The QDs have a ZnSe core and a ZnS shell, capped with hydrophobic ligands (triotylphosphine oxide/hexadecylamine), allowing them to be entrapped in a model biomembrane, bicelle, made of zwitterionic dipalmitoyl and dihexanoyl phosphatidylcholines and charged dipalmitoyl phosphatidylglycerol. Enhanced photoluminescence is observed upon encapsulation, depending on the QD‐to‐lipid ratio. Transmission electron microscopy and small‐angle X‐ray scattering confirm that QDs are preferably situated at the rim of bicellar discs. A simplified quantum dissipation heat‐bath theory is proposed to correlate the enhancement with slower nonradiative processes caused by the restriction‐in‐motion (RIM) of the surface ligands. However, Förster resonance energy transfer due to QD aggregation counteracts the effect.

A Simple and Low Cost Approach to Fabricate a Screen-Printed Electrode and Its Application for Alzheimer’s Disease Detection

Alzheimer's Disease (AD) is a progressive neurodegenerative disorder characterized by cognitive and memory deterioration, which requires an early diagnosis for effectiveness of treatment. Immunosensors based on AD biomarkers stand out as a fast, inexpensive and practical alternative for an early detection before it’s first symptoms. The development of Screen-Printed Electrodes (SPEs) meets a growing demand in the market for applications such as signal transducers in biosensor devices. The screen-printing technology stands out due to its simplicity of elaboration, the possibility of mass production of disposable electrodes and ease in miniaturization for use in portable devices. The objective of this work is to develop a simple and low cost method to manufacture a Screen Printed Carbon Electrode (SPCE) system to detect AD. The microelectrodes were obtained from carbon paste on two substrates: one from High Density Polyethylene (HDPE), and another one from Polyvinyl Chloride (PVC), shown Figure 1. The electrochemical response to adsorption of self-assembled polymer and lipid films was studied by cyclic voltammetry. The HDPE designed in Figure 3, offered a less capacitive and sensitive response to the assembly of the films compared to PVC. The immunosensor for AD diagnosis based on the detection of the IgG anti-Aβ1-40 autoantibody was built on SPCEs using layer by layer technique (LBL), as shown in Figure 4. The first bilayer consists of pure or functionalized Polyethyleneimine (PEI), and the second of liposomes of Dipalmitoyl Phosphatidyl Glycerol (DPPG) immobilizing the Aβ1-40 peptide, repeating this process in the second bilayer. In order to improve their sensory response, PEI functionalized with graphene produced via electrochemical exfoliation of graphite (EEG) was used, in the proportions of 0.6%, 1.2%, 2.4% and 10% (w / w).Cyclic voltammetry was used to evaluate the electrochemical behavior of the SPCE in three cycles, at potentials from -0.6 V to 0.6 V with a support electrolyte containing PBS pH 7.4, Figure 5. Electrochemical Impedance Spectroscopy (EIS) measurements, Figure 6, were evaluated, with frequencies ranging from 0.1 MHz to 0.1 Hz, amplitude 10mV, for pure and modified SPCEs with PEI and their EEG composites. For detection, autoantibody solutions, diluted in PBS, were prepared in concentrations ranging from 1ng / ml to 10 µg / ml. There was an evident decrease in capacitance in the samples of PEI with EEG, as seen in the voltammograms. After the functionalization of PEI, the capacitive currents of the electrodes clearly decreased, possibly due to an increase in the conductivity of the electrodes. On the other hand, the EIS curves for the electrodes with PEI functionalized, despite appearing to have larger modules than those in which it is not functionalized, the inclination of impedance spectra at low frequency presents minor slopes. This is possibly due to the higher permeation of charges in the interstices of the PEI, as observed in the microscopies in Figure 7, associating the capacitive process of these functionalized PEI electrodes capable of carrying more charge than the purely capacitive electrodes with bilayers composed only of PEI. This gave them a significantly higher degree of conductivity. After functionalization with EEG, the graphene particles appear to be dispersed in a disordered way on the polymer, being a possible cause of charge dispersion in pores, increasing their contact area, and justifying greater impedance values. PEI bands, as described in Figure 8, refer to asymmetric stretches of the C-N group at 1045, 1119, 1310 cm-1, 1598 and 1651 cm-1. Last band may be associated both to symmetrical angular deformation in plane for the -NH2 to amides (possibly provided by reaction between EEG carbonyl and PEI amines) and PEI existing amines It was noticed a change in the band at 1651 cm-1 in PEI composites with 0.6%, 2.4% and 10% assigned to amide formation, conferring a suitable dispersion. However, for others EEG contents, amide bands may be masked by amine groups of PEI.Sensorial measurements were better performed at HDPE subtract modified with PEI functionalized with EEG until 1.2% of content. There was an exponential growth trend in the normalized area of the voltammograms, Figure 9, as the Aβ1-40 autoantibody concentrations increased, distinguishing its variation. However, after 1μg / ml the curve stabilized showing the limit of detection (LOD). Figure 1

Chitosan effects on monolayers of zwitterionic, anionic and a natural lipid extract from E. coli at physiological pH

Langmuir monolayers are used to simulate the biological membrane environment, acting as a mimetic system of the outer or the inner membrane leaflet. Herein, we analyze the interaction of membrane models with a partially N-acetylated chitosan (Ch35%) possessing a quasi-ideal random pattern of acetylation, full water solubility up to pH ≈ 8.5 and unusually high weight average molecular weight. Lipid monolayers containing dipalmitoyl phosphatidyl choline (DPPC), dipalmitoyl phosphatidyl ethalonamine (DPPE), dipalmitoyl phosphatidyl glycerol (DPPG) or E. coli total lipid extract were spread onto subphases buffered at pH 4.5 or 7.4. The incorporation of Ch35% chitosan caused monolayer expansion and a general trend of decreasing monolayer rigidity with Ch35% concentration. Due to its relatively high content of N-acetylglucosamine (GlcNAc) units, Ch35% interactions with negatively charged monolayers and with E. coli extract were weaker than those involving zwitterionic monolayers or lipid rafts. While the smaller interaction with negatively charged lipids was unexpected, this finding can be attributed to the degree of acetylation (35%) which imparts a small number of charged groups for Ch35% to interact. Chitosan properties are therefore determinant for interactions with model cell membranes, which explains the variability in chitosan bactericide activity in the literature. This is the first study on the effects from chitosans on realistic models of bacterial membranes under physiological pH.

Composition, functionality and structural correlates of mixed lipid monolayers at air-water interface

Physicochemical properties of lipid monolayer depend on its composition where a blend of lipids exhibit superior behaviour than the individual component. Surface pressure ( π ) – area ( A ) isotherms of mixed monolayer (PC mix ) formed by dipalmitoylphosphatidylcholine (DPPC), palmitoleylphosphatidylcholine (POPC), soyphosphatidylcholine (SPC) and hydrogenated soy phosphatidylcholine (HSPC) in combination with 30 mole % cholesterol (Chol) were obtained by using Langmuir trough. Effects of dipalmitoylphosphatidyethanolamine (DPPE) on its mutual miscibility at the air-water interface with DPPC ​+ ​Chol, POPC ​+ ​Chol, HSPC ​+ ​Chol and SPC ​+ ​Chol were also investigated separately, followed by studying the effects of DPPE and dipalmitoylphosphatidylglycerol (DPPG) to PC mix ​+ ​Chol and PC mix ​+ ​DPPE ​+ ​Chol respectively. Lift-off area, minimum molecular area, excess molecular area, collapse pressure, Gibbs free energy of interfacial mixing, compressibility modulii values were evaluated by analyzing the isotherms. Deviations from the ideal mixing behaviours were dependent on the composition of the lipid blends. Surface dilatational rheology studies could assess monolayer elasticity, whereas the film morphologies were analysed by Brewster angle microscopic (BAM) studies. DPPC induced formation of condensed monolayer, whereas film rigidity was increased with the incorporation of DPPE and DPPG into mixed systems. Interactions among the lipid components of the investigated mixed systems were thoroughly discussed from the point of view of polar head and acyl chain saturation and obtained results follow the sequence: PC mix ​+ ​DPPE ​+ ​DPPG ​+ ​Chol ​> ​PC mix ​+ ​DPPE ​+ ​Chol ​> ​individual PC ​+ ​DPPE ​+ ​Chol which could be translated into the bilayer studies. Such investigation of mixed lipid is important for class of its own composition and combined results are expected to contribute in better understanding the interaction of selected lipid in proposed blend.

Periodic bilayer organization in the complexes of Beta-2 Glycoprotein I with anionic lipid membranes

β2 glycoprotein I (β2GPI) is a soluble protein that participates in blood coagulation, clearance of apoptotic bodies and generation of antigens in antiphospholipid syndrome among many other functions. We studied the aggregates formed by β2GPI with the anionic phospholipids palmitoyloleoylphosphatidyl glycerol, dimyristoylphosphatidyl glycerol, dipalmitoylphosphatidyl glycerol and cardiolipin using small angle X-ray scattering. The complexes obtained in a medium containing 0.01 M NaCl showed Bragg peaks up to the sixth order in a well-defined integer sequence indicating the presence of a lamellar stacking with a periodicity of 17.8 nm and with largely reduced membrane fluctuations. Modeling the complex signal allowed us to conclude that the coherence length was only two bilayers and that about 15% of the total surface was actually stacked. The space between bilayers allows accommodating an extended β2GPI molecule making a bridge between the interacting bilayers. The interactions between membranes mediated by β2GPI was favored when the membranes were in the liquid crystalline state.