Bilayer Disks Research Articles

Designing nanodiscs as versatile platforms for on-demand therapy.

Nowadays, there has been an increasing utilization of nanomedicines for disease treatment. Nanodiscs (NDs) have emerged as a novel platform technology that garners significant attention in biomedical research and drug discovery. NDs are nanoscale phospholipid bilayer discs capable of incorporating membrane proteins and lipids within a native-like environment. They are assembled using amphiphilic biomacromolecular materials, such as apolipoprotein A1 or membrane scaffold proteins (MSPs), peptides, and styrene-maleic acid polymers (SMAs). NDs possess well-defined sizes and shapes, offering a stable, homogeneous, and biologically relevant environment for studying membrane proteins and lipids. Their unique properties have made them highly desirable for diverse applications, including cancer immunotherapy, vaccine development, antibacterial and antiviral therapy, and treating Alzheimer's disease (AD) and diabetes-related conditions. This review discusses the classifications, advantages, and applications of NDs in disease therapy.

Thermodynamic and structural properties of lipid-photosensitizer conjugates mixed with phospholipids: Impact on the formation and stability of nano-assemblies

The photosensitizer Phenalenone (PN) was grafted with one or two lipid (C18) chains to form pure nano-assemblies or mixed lipid vesicles suitable for photodynamic therapy. Mixtures of PN-C18 conjugates with stearoyl-oleoyl phosphatidylcholine (SOPC) form vesicles that disintegrate into bilayer sheets as the concentration of PN-C18 conjugates increases. We hypothesized that PN-C18 conjugates control the thermodynamic and structural properties of the mixtures and induce the disintegration of vesicles due to PN π-π-interactions.Monolayers were analyzed by surface pressure and grazing incidence X-ray diffraction (GIXD) measurements, and vesicles by differential scanning calorimetry and cryo-TEM. The results showed that PN-triazole-C18 (1A) and PN-NH-C18 (1B) segregate from the phospholipid domains. PN-(C18)2 (conjugate 2) develops favorable interactions with SOPC and distearoyl-phosphatidylcholine (DSPC). GIXD demonstrates the contribution of SOPC to the structuring of conjugate 2 and the role of the major component in controlling the structural properties of DSPC-conjugate 2 mixtures. Above 10 mol% conjugate 2 in SOPC vesicles, the coexistence of domains with different molecule packing leads to conjugate segregation, vesicle deformation, and the formation of small bilayer discs stabilized by the inter-bilayer π-π stacking of PN molecules.

Fatigue behavior of multilayer ceramic structures in traditional and reverse layering designs.

This study evaluated the fatigue failure load (FFL) and the number of cycles for fatigue failure (CFF) of traditional (porcelain layer up) and reversed (zirconia layer up) designs of porcelain-veneered zirconia samples prepared with heat-pressing or file-splitting techniques. Zirconia discs were prepared and veneered with heat-pressed or machined feldspathic ceramic. The bilayer discs were bonded onto a dentin-analog according to the bilayer technique and sample design: traditional heat-pressing (T-HP), reversed heat-pressing (R-HP), traditional file-splitting with fusion ceramic (T-FC), reversed file-splitting with fusion ceramic R-FC), traditional file-splitting with resin cement (T-RC), and reversed file-splitting with resin cement (R-RC). The fatigue tests were performed using the stepwise approach at 20Hz, 10,000 cycles/step, step-size of 200 N starting at 600 N, and proceeding until failure detection or up to 2600 N if enduring. The failure modes (from radial and/or cone cracks) were analyzed in a stereomicroscope. The reversed design decreased the FFL and CFF of bilayers prepared with heat-pressing and file-splitting with fusion ceramic. The T-HP and T-FC reached the highest results, which were statistically similar between them. The bilayers prepared by the file-splitting with resin cement (T-RC and R-RC) were similar to the R-FC and R-HP groups regarding FFL and CFF. Almost all reverse layering samples failed by radial cracks. The reverse layering design did not improve the fatigue behavior of porcelain veneered zirconia samples. The three bilayer techniques behaved similarly when used in the reversed design.

Vortex arrangement in an ultrathin superconducting bilayer disc

A direct current superconducting quantum interference device (SQUID) fixed on a recently developed superconducting bilayer disc is effective for studying fractional magnetic quantum and fractional vortices. However, such devices cannot magnetically image fractional vortices. To overcome this limitation, we compared the bilayer SQUID signal with and without a pinhole to experimentally investigate the vortex arrangement. We measured the amount of flux passing through the SQUID ring via the SQUID signal phase shift, which reflected the vortex locations. We measured the voltages of a series of 100 SQUIDs in which the bilayer discs were present under each SQUID. When we applied a constant bias current of 5.5 μA and swept the external magnetic field, a voltage appeared when the amount of magnetic flux passing through the SQUID ring became close to half of the flux quantum. A sharp voltage peak was obtained for the bilayers with a pinhole, whereas these peaks were broader for the bilayers without a pinhole. We evaluated the amount of the flux originating from the vortices trapped in the bilayer discs and estimated the vortex locations from the peak position. When the amount of flux originating from the trapped vortices increased, the bilayer discs without a pinhole exhibited reduced amounts of flux originating from the trapped vortices by approximately 15% compared to the bilayer discs with a pinhole. When no pinhole was present, the vortex locations varied inside the SQUID ring. The results indicated that the vortices were located at the same position for all 100 bilayer discs with a pinhole; therefore, we concluded that the pinholes must trap the vortices.

SJLA_2024_A16: Unusual Complications of Open Preperitoneal Tension-Free Inguinal Hernia Repair

Background: The purpose of this study is to report two patients referred to our hospital after developing unusual complications attributed to prior open preperitoneal tension-free inguinal hernia repair. Methods: Case # 1 A 75-year-old male developed a left proximal thigh abscess requiring incision and drainage 5 years after open prosthetic, preperitoneal inguinal hernia repair with single-layer mesh. A pelvic computed tomography (CT) scan demonstrated an abscess in the left inguinal preperitoneal space as well as diverticulosis but no diverticulitis. A colonoscopy was negative for any additional findings. The patient underwent groin exploration, debridement of the inguinal canal and sinus tract, and mesh resection followed by JP drain placement. The inguinal defect was repaired utilizing the should ice technique. Intraoperative cultures revealed penicillin-resistant Staphylococcus aureus. The drain was removed on postoperative day nine, and his recovery remains uneventful without hernia recurrence or infectious complications at 11-month follow-up. Case # 2 An 82-year-old man developed progressive left lower extremity edema and disabling claudication 1 month after open prosthetic inguinal hernia repair with a bilayer disc mesh. Venous duplex imaging revealed compression of the left common femoral and external iliac veins. A CT angiogram demonstrated left external iliac and common femoral artery thrombosis and a 5.2 cm fluid collection consistent with hematoma. An inferior vena cava filter was placed. The common femoral artery was controlled after initial exploration revealed a common femoral artery pseudoaneurysm. The mesh was densely adherent to the external iliac vein and was thus not resected. The common femoral artery pseudoaneurysm was repaired and the profunda femoris artery over sewn. An external iliac to superficial femoral artery 8 mm Polytetrafluoroethylene bypass graft was placed. The patient was discharged on systemic anticoagulation without claudication on postoperative day thirteen. Results: Laparoscopic inguinal hernia repair and novel, open techniques and mesh devices have popularized the placement of mesh in the preperitoneal space. Acute, severe vascular complications and latent infectious complications can occur after open, preperitoneal tension-free inguinal hernia repair and may require remedial surgery. CT scans and angiogram pictures are available. Discussion: Laparoscopic inguinal hernia repair and novel, open techniques and mesh devices have popularized the placement of mesh in the preperitoneal space. Acute, severe vascular complications and latent infectious complications can occur after open, preperitoneal tension-free inguinal hernia repair and may require remedial surgery. CT scans and angiogram pictures are available. Conclusion: Laparoscopic inguinal hernia repair and novel, open techniques and mesh devices have popularized the placement of mesh in the preperitoneal space. Acute, severe vascular complications and latent infectious complications can occur after open, preperitoneal tension-free inguinal hernia repair and may require remedial surgery. CT scans and angiogram pictures are available.

α-Synuclein Interaction with Lipid Bilayer Discs.

α-Synuclein (aSyn) is a 140 residue long protein present in presynaptic termini of nerve cells. The protein is associated with Parkinson’s disease, in which case it has been found to self-assemble into long amyloid fibrils forming intracellular inclusions that are also rich in lipids. Furthermore, its synaptic function is proposed to involve interaction with lipid membranes, and hence, it is of interest to understand aSyn–lipid membrane interactions in detail. In this paper we report on the interaction of aSyn with model membranes in the form of lipid bilayer discs. Using a combination of cryogenic transmission electron microscopy and small-angle neutron scattering, we show that circular discs undergo a significant shape transition after the adsorption of aSyn. When aSyn self-assembles into fibrils, aSyn molecules desorb from the bilayer discs, allowing them to recover to their original shape. Interestingly, the desorption process has an all-or-none character, resulting in a binary coexistence of circular bilayer discs with no adsorbed aSyn and deformed bilayer discs having a maximum amount of adsorbed protein. The observed coexistence is consistent with the recent finding of cooperative aSyn adsorption to anionic lipid bilayers.

Observation of multiple fractional quanta in a superconducting bilayer disk with a pinhole

The magnetic-flux quantum (Φ0) cannot be generally divided into smaller ones in a usual superconductor. If a subdividing path is present, the science and technology of the superconductor drastically change. In previous studies, we experimentally presented a new, easy way to subdivide Φ0 into smaller quanta (Φf) using an ultrathin Nb bilayer with a through-pinhole (H. Ishizu, H. Yamamori, S. Arisawa, K. Tokiwa, Y. Tanaka, Physica C 595 (2022) 1354029.). However, because of an experimental technical problem, we could not use a higher field to generate more than one Φf. It was not clear if multiples of Φf could be trapped in a pinhole (in a usual superconductor, multiples of Φ0 are trapped in a pinhole). In this study, we observed multiple fractional phase shifts with a direct-current superconducting interference device (SQUID) placed on an ultrathin Nb bilayer disk containing a through-pinhole using an improved modification of an experimental procedure. The phase shift for the SQUID with the bilayer disk is an integer multiple of a basic fractional phase shift divided by 2π. However, we did not observe the fractional phase shift for a SQUID with a single-layer disk. These observations indicate that multiple fractional flux quanta are trapped in the bilayer disk. According to the vortex-molecule model, if the vortex molecule is larger than the bilayer disk, such quanta can be trapped. This is one example of altering a fundamental (universal) physical constant. The SQUID can more sensitively detect Φf than Φ0, rendering the design of novel superconducting electronics possible.

Structure of AlNiGd nanocomposites with enhanced ductility produced by high pressure torsion processing

The fully consolidated bilayer (consisting of two amorphous Al86Ni9Gd5 melt-spun ribbons) disks with a diameter of 5 mm were produced using unconstrained high pressure torsion (HPT) processing subjecting for 2–5 turns at applied pressures of 2 and 4 GPa. The structure and mechanical properties of the disks as well as those of the as-prepared and heated up to 479 K ribbon samples were studied by X-ray diffraction analysis, transmission and scanning electron microscopy, three-point bending tests and measurements of microhardness. It was established that the heat treatment and consolidation resulted in formation of nanocomposite structures consisting of Al nanocrystals 21 and 6 nm in size, respectively, with enhanced microhardness. It was shown that the heat treatment of the ribbon led to a loss of the plastic strain (from 2 to 0.4%) while the consolidated disks retained the same level of plasticity as that in the as-cast amorphous ribbon. The possible reason of the enhanced plasticity of the nanophase composites obtained by deformation-induced crystallization was discussed.

An optimized protocol for on-column nanodisc formation

For membrane proteins, a bilayer environment has proven to be of fundamental importance for the isolation and stabilization of individual subunits and macromolecular complexes. For that reason, membrane protein reconstitution has long represented a biochemical challenge in the pursuit of biochemical, structural and functional studies. Protein-lipid nanodiscs offer a membrane-mimicking environment that provides many advantages for structural and functional studies of integral membrane proteins. In its original conception (Sliger’s lab), nanodisc formation requires that target membrane proteins of interest are purified in detergent and reconstituted into a bilayer disc, encapsulated by an amphipathic membrane scaffold protein (MSP).

Vortex Arrangement in an Ultrathin Superconducting Bilayer Disc

Vortex Arrangement in an Ultrathin Superconducting Bilayer Disc

Observation of Multiple Fractional Quanta in a Superconducting Bilayer Disk with a Pinhole

Observation of Multiple Fractional Quanta in a Superconducting Bilayer Disk with a Pinhole

Effective method of forming and detecting a fractional magnetic flux quantum

We developed a minimal method of forming and detecting a fractional magnetic flux quantum in a two-component superconductor fabricated using an ultra-thin niobium bilayer that sandwiches a Josephson barrier. The fractional quantum created by cooling a bilayer disk in the magnetic field survived after turning off the magnetic field. The survived quantum was confirmed by the change in the critical current of a superconducting quantum interference device (SQUID) placed on the bilayer disk. By replacing a flux qubit with the SQUID, we can use the fractional flux quantum as the flux bias for the flux qubit. This method enables the investigation of fractional quantization and the related topology.

Inverse Design of Axisymmetric Shapes in Glassy Nematic Bilayers

Abstract Programming desired shapes of stimuli-responsive nematic sheets is a challenging yet crucial problem for morphing applications. Here we propose a strategy for inverse design of axisymmetric shapes via circular bilayers composed of two glassy nematic layers with radially symmetric and orthogonal director fields. In this situation, an explicit link can be established between the director orientation and the bilayer deflection. The result provides a recipe for how to construct a director pattern which, upon stimulation, leads to a prescribed shape of revolution. For illustration, two examples concerning simply supported and loosely clamped bilayer discs are given.

Asymmetric opening of the homopentameric 5-HT3A serotonin receptor in lipid bilayers

Pentameric ligand-gated ion channels (pLGICs) of the Cys-loop receptor family are key players in fast signal transduction throughout the nervous system. They have been shown to be modulated by the lipid environment, however the underlying mechanism is not well understood. We report three structures of the Cys-loop 5-HT3A serotonin receptor (5HT3R) reconstituted into saposin-based lipid bilayer discs: a symmetric and an asymmetric apo state, and an asymmetric agonist-bound state. In comparison to previously published 5HT3R conformations in detergent, the lipid bilayer stabilises the receptor in a more tightly packed, ‘coupled’ state, involving a cluster of highly conserved residues. In consequence, the agonist-bound receptor conformation adopts a wide-open pore capable of conducting sodium ions in unbiased molecular dynamics (MD) simulations. Taken together, we provide a structural basis for the modulation of 5HT3R by the membrane environment, and a model for asymmetric activation of the receptor.

Assessment of Capability of Lithium Disilicate Glass Ceramics to Reproduce the A2 Shade and to Mask A4 Substrates

Aim: Capability of lithium disilicate glass ceramics to reproduce the A2 shade and to mask A4 substrates.Materials and Method: 32 lithium disilicate ceramic specimens were classified materials into 8 groupsof 4 each. Group I- III high translucency (HT) monolayer discs, group IV- VI had low translucency (LT)monolayer discs, group VII- VIII had medium translucency (MO) bilayer discs. A2 shade tab shade guide(VITA classical A1-D4) was used for CIE L*a*b* measurements of control. Color differences using thresholdswas as follows: for imperceptible (?E < 1.0), perceptible (1.0 < ?E < 3.3), and clinically unacceptable (?E> 3.3) differences.Results: ?E in group IV exhibited highest (9.92±0.21) and group III lowest (8.08± 0.25) value. When thecomplex was compared to the A4 substrate, significant difference among all groups (p < 0.05) was found.?E values when the complex was compared to the A2 shade tab, group III exhibited highest (4.82±0.16)and group V lowest (0.72±0.06) value. In groups I specimens had acceptable differences, group II hadperceptible differences, group III had perceptible differences, group IV had acceptable differences, group Vhad imperceptible differences, group VI and VII had acceptable differences and group VIII had imperceptibledifferences.Conclusion: Authors suggested to use LT ceramics with less thickness in the range of 0.5 mm- 0.7 mm tomask discoloration.

A thermodynamic study of F108 and F127 block copolymer interactions with liposomes at physiological temperature

The interactions of egg yolk phosphatidylcholine liposomes with F108 and F127 triblock copolymers, in the monomer state, were analyzed by isothermal titration calorimetry (ITC) at 37 °C. According to the results, the critical micelle concentration was determined to be 0.4 and 0.04 wt.% for F108 and F127, respectively, by surface tension at 37 °C. According to the results, liposomes/poloxamers were not favoured energetically, since endothermic interactions were observed. However, positive changes in entropy promoted a spontaneous process. F127 had a greater partition coefficient (51.97 ± 1.77 × 104), stronger affinity, than F108 (8.19 ± 0.37 × 104) towards the vesicle lipid bilayer due to its larger hydrophobic block. After the ITC experiments, an increased vesicle size (within about 1–3 nm average) by dynamic light scattering and the formation of bilayer discs by electron microscopy (EM) was observed at low copolymer concentrations (0.57 mol% of F108 and 1.01 mol% of F127). The EM and ITC results confirmed the intimate association of the copolymers with the membrane instead of being simply absorbed onto the bilayer surface. Our results indicate that the temperature of the system (37 °C), the copolymer concentration and hydrophobic chain length are important factors for the interaction of poloxamers with lipid bilayers and the stability of liposomes.

Magnetic Phase Diagram of a Superconducting Bilayer Disk: Effects of Geometrical Modification

We study the equilibrium states of a superconducting bilayer disk, which has been attracting attention as a base of a novel fractional flux quantum state. Our analysis is based on the Ginzburg–Land...

Fatigue behavior and colorimetric differences of a porcelain-veneered zirconia: effect of quantity and position of specimens during firing

To evaluate the influence of quantity and positioning of veneered zirconia specimens during firing of porcelain on their fatigue performance and colorimetric differences. Bilayer discs (Ø=15 mm) were made, following ISO 6872 guidelines, using a Y-TZP core (yttria-stabilized tetragonal zirconia polycrystal ceramic; VITA In-Ceram YZ) and a feldspathic veneering material (VITA VM9), being both layers with 0.7 mm thickness. Y-TZP discs were sintered, the veneering material was applied over it, and the bilayer specimens were fired according to two factors (n=20): 'quantity' (1 or 5 samples per firing cycle; G1 and G5 groups respectively) and 'positioning' of the specimens inside the furnace (center or periphery of the refractory tray; G5C and G5P groups, respectively). The CIEL*a*b* parameters were recorded with a spectrophotometer and the color difference (ΔE 00 ) and translucency (TP 00 ) were calculated using CIEDE2000 equations. The step-stress fatigue test was performed with the veneer facing down (region of tensile stress concentration), 10 Hz frequency, initial tension of 20 MPa for 5,000 cycles, followed by steps of 10,000 cycles using a step size of 10 MPa, up to 100 MPa; data from strength and number of cycles for failure were recorded for statistical analysis. Unacceptable color differences (ΔE 00 >1.8) were observed comparing G5C vs. G1 (quantity) and G5C vs. G5P (positioning), meanwhile translucency parameters were not affected. Besides, only the 'quantity' factor influenced the fatigue performance (G1>G5C). None of the tested specimens survived beyond 90N and/or 75000 cycles. The quantity and position of the specimens during firing influence the final color of porcelain-veneered zirconia, and firing one specimen per cycle improved the fatigue performance of the bilayer system.

Effect of gangliosides on structure and integrity of polyethylene glycol (PEG)-stabilized liposomes

The in vivo efficacy and tolerance of polyethylene glycol (PEG)-decorated drug nanocarriers, such as liposomes, is compromised bytheir tendency to induce the generation of PEG-specific immunoglobulin M (IgM) antibodies. Recently, a number of independent studies have reported on an attenuated anti-PEG immune response upon incorporation of gangliosides in the membrane of PEGylated liposomes.In the present study we investigate the effect of gangliosides on the self-assembled structures found in lipid dispersions based on hydrogenated egg phosphatidylcholine (HEPC), cholesterol and 1,2-distearoyl-sn-glycero-3-phosphoethanolamine-N-[methoxy(polyethyleneglycol)-2000] (DSPE-PEG(2000)). Results from cryo-transmission electron microscopy (cryo-TEM) and dynamic light scattering (DLS) investigations show that gangliosides promote structural transitions from liposomes to bilayer disks. In case of samples comprising 5 mol% PEG-conjugated lipids (PEG-lipid), inclusion of 2.5 mol% ganglioside (porcine ganglioside extract) results in the presence of a small but significant amount of disks. With increasing ganglioside content the population of disks grows at the expense of the liposomes. Comparative investigations using isolated ganglioside components reveal that disialoganglioside GD1a is more potent than monosialoganglioside GM1 in promoting disk formation. Experiments involving liposome encapsulated carboxyfluorescein confirm that the ganglioside-induced structural transformations have a detrimental effect on the total entrapped aqueous volume of the samples. The reported coexistence of liposomes and bilayer disks may if overlooked have important implications for the therapeutic efficacy and immunogenicity of ganglioside-supplemented liposomal formulations.

Tumor-Targeted Delivery of the p53-Activating Peptide VIP116 with PEG-Stabilized Lipodisks.

Stapled peptides targeting the interaction between p53 and its negative regulators MDM2 and MDM4 have exhibited great potential as anti-cancer drugs, albeit with room for improvement in formulation and tumor specificity. Lipid bilayer disks (lipodisks) have emerged as promising drug nanocarriers and can by attachment of targeting moieties be directed selectively towards tumor cells. Tumor-targeted delivery of stapled peptides by use of lipodisks may therefore increase the uptake in the tumors and limit toxicity in healthy tissue. Here, we utilized epidermal growth factor receptor (EGFR)-targeted lipodisks to deliver p53-activating stapled peptide VIP116 to EGFR-expressing tumor cells. We demonstrate that VIP116 can be stably formulated in lipodisks (maximum peptide/lipid molar ratio 0.11). In vitro cell studies verify specific binding of EGF-decorated lipodisks to tumor cells and confirm that targeted delivery of VIP116 significantly decreases tumor cell viability.