Study on the subgel-phase formation using an asymmetric phospholipid bilayer membrane by high-pressure fluorometry.

The myristoylpalmitoylphosphatidylcholine (MPPC) bilayer membrane shows a complicated temperature-pressure phase diagram. The large portion of the lamellar gel (L(ß)'), ripple gel (P(ß)'), and pressure-induced gel (L(ß)I) phases exist as metastable phases due to the extremely stable subgel (L(c)) phase. The stable L(c) phase enables us to examine the properties of the L(c) phase. The phases of the MPPC bilayers under atmospheric and high pressures were studied by small-angle neutron scattering (SANS) and fluorescence spectroscopy using a polarity-sensitive fluorescent probe Prodan. The SANS measurements clearly demonstrated the existence of the metastable L(ß)I phase with the smallest lamellar repeat distance. From a second-derivative analysis of the fluorescence data, the line shape for the L(c) phase under high pressure was characterized by a broad peak with a minimum of ca. 460 nm. The line shapes and the minimum intensity wavelength (λ″(min)) values changed with pressure, indicating that the L(c) phase has highly pressure-sensible structure. The λ″(min) values of the L(c) phase spectra were split into ca. 430 and 500 nm in the L(ß)I phase region, which corresponds to the formation of a interdigitated subgel L(c) (L(c)I) phase. Moreover, the phase transitions related to the L(c) phase were reversible transitions under high pressure. Taking into account the fluorescence behavior of Prodan for the L(c) phase, we concluded that the structure of the L(c) phase is highly probably a staggered structure, which can transform into the L(c)I phase easily.

Comparative analysis of viscosity of complex liquids and cytoplasm of mammalian cells at the nanoscale.

We present a scaling formula for size-dependent viscosity coefficients for proteins, polymers, and fluorescent dyes diffusing in complex liquids. The formula was used to analyze the mobilities of probes of different sizes in HeLa and Swiss 3T3 mammalian cells. This analysis unveils in the cytoplasm two length scales: (i) the correlation length ξ (approximately 5 nm in HeLa and 7 nm in Swiss 3T3 cells) and (ii) the limiting length scale that marks the crossover between nano- and macroscale viscosity (approximately 86 nm in HeLa and 30 nm in Swiss 3T3 cells). During motion, probes smaller than ξ experienced matrix viscosity: η(matrix) ≈ 2.0 mPa·s for HeLa and 0.88 mPa·s for Swiss 3T3 cells. Probes much larger than the limiting length scale experienced macroscopic viscosity, η(macro) ≈ 4.4 × 10(-2) and 2.4 × 10(-2) Pa·s for HeLa and Swiss 3T3 cells, respectively. Our results are persistent for the lengths scales from 0.14 nm to a few hundred nanometers.

Tumour-Related Parameters as a Prognostic Factor in Patients with Advanced Cervical Cancer: 20-Year Follow-Up of Diagnostic and Treatment Changes during Chemioradiotherapy.

Concurrent radiochemotherapy (RCHT) has been the standard treatment for locally advanced cervical cancer since 1999. During this 20-year period, both diagnostic and radiotherapy techniques have developed, such as positron emission tomography (PET) or brachytherapy (BT) planning. The aim of the study was to assess the relationships between prognostic factors and the results of treatment in patients with advanced cervical cancer independent of these changes. The analysis included 266 patients with stage IIB or IIIB FIGO 2009 cervical cancer divided into two groups: one including 147 patients diagnosed with physical examination and ultrasonography (USG) and treated with RCHT with 2D BT from 2001 to 2005; another including 119 patients with metastatic pelvic lymph node diagnosed with PET and treated from 2010 to 2016 with RCHT and 3D BT. The mean five-year overall survival (OS) rate was 59.2% in the first vs. 65.5% in the second group (p = 0.048). However, in both groups, stage IIB patients had a significantly higher 5-year OS rate, despite the presence of nodal metastases in group 2. In the first group it was 75.1% in IIB vs. 54.8% in IIIB (p = 0.040) 5-year OS and 77.5% vs. 55.8% (p = 0.034) in the second group. Important was also a significant association between the dose of BT and survival in group 2: 45.7% vs. 69.2% for dose <28 Gy and 28 Gy (p = 0.018). Evolution in the diagnosis and treatment of patients with cervical cancer had led to improvement in the survival of patients and precise treatment with an appropriate stage assessment. However local advance of the tumour is still the most important prognostic factor.

Synthesis and Photoluminescence Properties of Pr3+-Doped Ba0.5Ca0.5TixZr(1-x)O3 Perovskite Diphasic Ceramics Obtained by the Modified Pechini Method.

The Pr3+-doped solid solutions from (Ba,Ca)(Ti,Zr)O3 (BCTZO) system were successfully synthesized using an efficient and low-energy consuming route-the Pechini method combined with the sintering at relatively low temperature (1450 °C). The obtained materials were characterized by means of X-ray diffraction (XRD) and scanning electron microscopy (SEM). The dielectric properties were systematically studied. The Pr3+-doped BCTZO diphasic material generates intense and broad red photoluminescence (PL) emission at room temperature. The optical properties were significantly improved with the Ti4+ substitution by Zr4+ ions. As a result, the Pr3+-doped (Ba,Ca)(Ti,Zr)O3 ceramics is a promising candidate for environmentally friendly, multifunctional material by combining good dielectric and photoluminescent properties with prognosis for the manifestation of strong photoluminescent and mechanoluminescent effects.

Hydrothermal Aging of ATZ Composites Based on Zirconia Made of Powders with Different Yttria Content.

The presented work concerns the development and investigation of three different grades of ZrO2 materials containing Al2O3 particles (ATZ-Alumina Toughened Zirconia ceramics with 2.3-20 vol.% of alumina). The zirconia powders containing 3 mol.% of yttria were synthesized by a precipitation/calcination method and fabricated from two different zirconia powders with different yttria content. Then, the selected ATZ composites (ATZ-B, ATZ-10 and ATZ-20) were prepared by means of conventional mixing, compacting and sintering at 1450 °C for 1.5 h. The phase composition, microstructure, relative density and basic mechanical properties were determined. Uniform microstructures with relative densities over 99% of theoretical density, hardness values between 12.0-13.8 GPa, flexural strength up to 1 GPa and outstanding fracture toughness of 12.7 MPa⋅m1/2 were obtained. The aging susceptibility of alumina toughened zirconia materials, as a consequence of hydrothermal treatment, was investigated. The aim of this study was to determine the influence of LTD (low temperature degradation) on the tetragonal to monoclinic phase transitions and on the flexural strength of hydrothermally aged specimens. The results were compared to those obtained for commercially available tetragonal zirconia-based materials containing 3 mol.% of yttria. This research shows that ATZ composites that have excellent mechanical properties and sufficient hydrothermal aging resistance can be attained and later used in technical and biomedical applications.

Size and shape of micelles studied by means of SANS, PCS, and FCS.

The hexaethylene glycol monododecyl ether (C(12)E(6)) micelles at concentrations up to 10% have been studied in their isotropic phase (10-48 degrees C) by means of small angle neutron scattering (SANS) and photon correlation spectroscopy (PCS). The SANS data obtained at low temperatures could be unequivocally interpreted as a result of scattering from a suspension of compact globular micelles with the shape of a triaxial ellipsoid or a short end-capped elliptical rod. Different models have been applied to analyze the SANS data obtained at higher temperatures: (i) elongated rod-like micelles with purely sterical interactions, (ii) compact globular micelles with a weak attractive potential, and (iii) globular micelles influenced by the critical phenomena in the whole temperature range studied. The good quality of the experimental data indicated model (i) as the best fit for our data. The diffusion coefficients obtained from the PCS measurements have been compared to the diffusion coefficients calculated for the rod-like micelles--results of the SANS data analysis. A good agreement was achieved using the solvent viscosity, in agreement with the theoretical predictions for sterically interacting globular colloidal particles. Finally, the SANS results obtained at 24 degrees C were compared to the micelle self-diffusion coefficients previously measured by means of fluorescence correlation spectroscopy (FCS) at this temperature. The good agreement obtained after scaling the data with solution viscosity supports the validity of the generalized Stokes-Einstein relation in sterically interacting systems: the product of the colloidal particle self-diffusion coefficient and the macroscopic viscosity remains constant in a broad range of concentrations. It has been concluded that the FCS technique in combination with simple viscosity measurements might serve as a tool for estimating the micellar size and shape.

A Large Piezoelectric Strain Recorded in BCT Ceramics Obtained by a Modified Pechini Method.

There is a strong need in the industry to develop lead-free piezoelectrics for sensors and actuators. Although these materials have become an important component of many electronic devices, it is very important for the industry to decarbonise ceramic technology, especially through the introduction of modern sintering technologies. Among the many piezoelectric compounds available, Calcium Barium Titanate (BCT) have been widely investigated because of its similar performance to lead-containing Lead Titanate Zirconate (PZT). In this paper, a modified Pechini method for obtaining ceramic Ba0.9Ca0.1TiO3 nano-powders is described. Deviation from the established procedure resulted in the precipitation of the solution or obtaining of a low-quality (poorly crystallized) product with numerous impurities. The samples of BCT materials were examined to find their ideal microstructures and structures; these factors were confirmed by their outstanding X-ray diffraction spectra and high piezoelectric constant values that are comparable to commercial lead-containing materials.

Scaling form of viscosity at all length-scales in poly(ethylene glycol) solutions studied by fluorescence correlation spectroscopy and capillary electrophoresis.

We measured the viscosity of poly(ethylene glycol) (PEG 6000, 12,000, 20,000) in water using capillary electrophoresis and fluorescence correlation spectroscopy with nanoscopic probes of different diameters (from 1.7 to 114 nm). For a probe of diameter smaller than the radius of gyration of PEG (e.g. rhodamine B or lyzozyme) the measured nanoviscosity was orders of magnitude smaller than the macroviscosity. For sizes equal to (or larger than) the polymer radius of gyration, macroscopic value of viscosity was measured. A mathematical relation for macro and nanoviscosity was found as a function of PEG radius of gyration, R(g), correlation length in semi-dilute solution, xi, and probe size, R. For R < R(g), the nanoviscosity (normalized by water viscosity) is given by exp(b(R/xi)a), and for R > R(g), both nano and macroviscosity follow the same curve, exp(b(R/xi)a), where a and b are two constants close to unity. This mathematical relation was shown to equally well describe rhodamine (of size 1.7 nm) in PEG 20,000 and the macroviscosity of PEG 8,000,000, whose radius of gyration exceeds 200 nm. Additionally, for the smallest probes (rhodamine B and lysozyme) we have verified, using capillary electrophoresis and fluorescence correlation spectroscopy, that the Stokes-Einstein (SE) relation holds, providing that we use a size-dependent viscosity in the formula. The SE relation is correct even in PEG solutions of very high viscosity (three orders of magnitude larger than that of water).

Net charge and electrophoretic mobility of lysozyme charge ladders in solutions of nonionic surfactant.

We report on the electrophoretic mobility and on the thermal diffusion of lysozyme proteins dissolved in aqueous solutions of a nonionic surfactant (C12E6) at a wide range of concentrations of the surfactant (0-20% by weight). We want to estimate the influence of a dense network of elongated micelles of C12E6 on the effective charge of the proteins as observed in the capillary electrophoresis experiments. The possible mechanism leading to the change in the effective charge of protein could involve the deformation of the cloud of counterions around the protein when it squeezes through the narrow (of the order of a protein diameter) aqueous channels formed in the solution of elongated micelles. The combination of independent measurements of the electrophoretic mobility of a family of modified proteins (lysozyme charge ladder [Colton et al. J. Am. Chem. Soc. 1997, 119, 12701]), of the microviscosity of the solutions of surfactant (obtained via fluorescence correlation spectroscopy), and of the hydrodynamic radius of the proteins (photon correlation spectroscopy) allow us to conclude that the effective charge of the proteins is not affected by the presence of surfactant, even at high concentrations.

Spectroscopic studies of D-alpha-tocopherol concentration-induced transformation in egg phosphatidylcholine vesicles.

The effects of embedding up to 60 mol% of alpha-tocopherol (alpha-Toc) on the morphology and structure of the egg phosphatidylcholine (PC) membrane were studied using spectroscopic techniques. The resulting vesicles were subjected to turbidometric and dynamic light scattering measurements to evaluate their size distribution. The alpha-Toc intrinsic fluorescence and its quenching was used to estimate the tocopherol position in the membrane. Optical microscopy was used to visualize morphological changes in the vesicles during the inclusion of tocopherol into the 2 mg/ml PC membrane. The incorporation of up to 15 mol% of tocopherol molecules into PC vesicles is accompanied by a linear increase in the fluorescence intensity and the simultaneous formation of larger, multilamellar vesicles. Increasing the tocopherol concentration above 20 mol% induced structural and morphological changes leading to the disappearance of micrometer-sized vesicles and the formation of small unilamellar vesicles of size ranging from 30 to 120 nm, mixed micelles and non-lamellar structures.

Diffusion and viscosity in a crowded environment: from nano- to macroscale.

Although water is the chief component of living cells, food, and personal care products, the supramolecular components make their viscosity larger than that of water by several orders of magnitude. Using fluorescence correlation spectroscopy (FCS), photon correlation spectroscopy (PCS), NMR, and rheology data, we show how the viscosity changes from the value for water at the molecular scale to the large macroviscosity. We determined the viscosity experienced by nanoprobes (of sizes from 0.28 to 190 nm) in aqueous micellar solution of hexaethylene-glycol-monododecyl-ether (in a range of concentration from 0.1% w/w to 35% w/w) and identified a clear crossover at the length scale of 17 +/- 2 nm (slightly larger than persistence length of micelles) at which viscosity acquires its macroscopic value. The sharp dependence of the viscosity coefficients on the size of the probe in the nanoregime has important consequences for diffusion-limited reactions in crowded environments (e.g., living cells).

Movement of proteins in an environment crowded by surfactant micelles: anomalous versus normal diffusion.

Small proteins move in crowded cell compartments by anomalous diffusion. In many of them, e.g., the endoplasmic reticulum, the proteins move between lipid membranes in the aqueous lumen. Molecular crowding in vitro offers a systematic way to study anomalous and normal diffusion in a well controlled environment not accessible in vivo. We prepared a crowded environment in vitro consisting of hexaethylene glycol monododecyl ether (C(12)E(6)) nonionic surfactant and water and observed lysozyme diffusion between elongated micelles. We have fitted the data obtained in fluorescence correlation spectroscopy using an anomalous diffusion model and a two-component normal diffusion model. For a small concentration of surfactant (below 4 wt %) the data can be fitted by single-component normal diffusion. For larger concentrations the normal diffusion fit gave two components: one very slow and one fast. The amplitude of the slow component grows with C(12)E(6) concentration. The ratio of diffusion coefficients (slow to fast) is on the order of 0.1 for all concentrations of surfactant in the solution. The fast diffusion is due to free proteins while the slow one is due to the protein-micelle complexes. The protein-micelle interaction is weak since even in a highly concentrated solution (35% of C(12)E(6)) the amplitude of the slow mode is only 10%, despite the fact that the average distance between the micelles is the same as the size of the protein. The anomalous diffusion model gave the anomaly index (r(2)(t) approximately t(alpha)), alpha monotonically decreasing from alpha = 1 (at 4% surfactant) to alpha = 0.88 (at 37% surfactant). The fits for two-component normal diffusion and anomalous diffusion were of equally good quality, but the physical interpretation was only straightforward for the former.

Chain elongation of diacylphosphatidylcholine induces fully bilayer interdigitation under atmospheric pressure.

The phase transitions of dibehenoylphosphatidylcholine (C22PC) bilayer membrane were observed by differential scanning calorimetry under atmospheric pressure and light-transmittance measurements under high pressure. The constructed temperature-pressure phase diagram suggests that the gel phase at low temperatures is the interdigitated gel phase. To confirm the phase state, we performed small-angle neutron scattering and fluorescence measurements using a polarity-sensitive probe Prodan for the C22PC bilayer membrane under atmospheric pressure. The peaks obtained in both measurements clearly showed the characteristic patterns of the fully interdigitated gel phase. Taking into account of previous studies on the gel phase for long-chain PC bilayers under atmospheric pressure and our studies on the pressure-induced bilayer interdigitaion of diacyl-PCs, it turned out that the interdigitation of diacyl-PC bilayer membranes occurs when the carbon number of acyl chain reaches at least 22. The present study revealed that the interdigitation of PC bilayer membranes occurs not only by weakening the attractive force of polar head groups but also by strengthening the cohesive force of acyl chains. When dominating the force of acyl chains, the interdigitation can be induced even in a diacyl-PC bilayer membrane by only hydration under atmospheric pressure.

On the shape of bottle-brush macromolecules: systematic variation of architectural parameters.

We measured the form factor of bottle-brush macromolecules under good solvent conditions with small-angle neutron scattering and static light scattering. The systems under investigation are brushes, synthesized via the grafting-from route, built from a poly(alkyl methacrylate) backbone to which poly(n-butyl acrylate) side chains are densely grafted. The aim of our work is to study how the systematic variation of structural parameters such as the side chain length and backbone length change the conformation of the polymer brushes in solution. All spectra can be consistently described by a model, considering the bottle-brush polymers as flexible rods with internal density fluctuations. Parameters discussed are (1) the contour length per main chain monomer l(b), (2) the fractal dimension of the side chains Ds, as well as (3) the fractal dimension D, and (4) the Kuhn length lambdak of the overall brush. l(b)=0.253+/-0.008 nm is found to be independent of the side chain length and equal to the value found for the bare main chain, indicating a strongly stretched conformation for the backbone due to the presence of the side chains. The fractal dimension of the side chains is determined to be Ds=1.75+/-0.07 which is very close to the value of 10.588 approximately 1.70 expected for a three-dimensional self-avoiding random walk (3D-SAW) under good solvent conditions. On larger length scales the overall brush appears to be a 3D-SAW itself (D=1.64+/-0.08) with a Kuhn-step length of lambdak=70+/-4 nm. The value is independent of the side chain length and 46 times larger than the Kuhn length of the bare backbone (lambdak=1.8+/-0.2 nm). The ratio of Kuhn length to brush diameter lambda(k)d>or=20 determines whether lyotropic behavior can be expected or not. Since longer side chains do not lead to more persistent structures, lambda(k)d decreases from 8 to 4 with increasing side chain length and lyotropic behavior becomes unlikely.