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Nowadays, modern pharmaceutical investigations are directed toward the design and production of drug delivery systems for achieving prolonged and controlled drug delivery. In this respect, the use of interpenetrating polymer networks (IPNs) is an opportunity in the preparation of polymer drug delivery systems with desired characteristics. This paper describes the synthesis and characterization of novel poly(2-(dimethylamino) ethyl methacrylate) (PDMAEMA) and polyacrylamide (PAAm)-based IPNs with different compositions and their application as diclofenac sodium delivery systems. The prepared IPNs were shown to possess phase-separated structures at the nano level, as revealed by SEM and TM-DSC. The IPNs’ composition was shown to determine the swelling behavior of these novel materials, and the inclusion of the charged IPN component (PDMAEMA) has changed the water molecules type diffusion from Fickian to non-Fickian, as revealed by the swelling kinetics study. Loading efficiency of diclofenac sodium and diclofenac sodium content in the polymer network was evaluated, and in vitro drug release experiments were carried out in order to estimate the ability of the obtained IPNs to control the release of the water-soluble drug.

Natural alkaloid galantamine is widely used for the treatment of Alzheimer’s disease and various other memory impairments. Galantamine hydrobromide (GH) is available as fast-release tablets, extended-release capsules, and oral solution. However, oral delivery of GH can cause side effects, such as nausea, vomiting, and gastrointestinal disturbance. Transdermal delivery is one possible way to avoid such unwanted effects and is also patient friendly, especially for unconscious patients, patients with swallowing difficulties, and patients with mental disturbances. In this work, thermoresponsive copolymer networks based on poly(N-isopropylacrylamide) were studied as new platforms for potential transdermal GH delivery. Series of networks of varied structure and composition were synthesized and investigated by applying swelling kinetics and different spectroscopic and thermal methods. The network films were drug loaded in GH aqueous solution and main characteristics of the obtained GH delivery systems were evaluated. The GH release kinetic profiles in BS at 37 °C were established and discussed.

SUMMARYLong considered as cytotoxic reagents, reactive oxygen species (ROS) at the right concentration promote cell proliferation in cell culture, stem cells and model organisms. However, how ROS signaling is coordinated with cell cycle progression and integrated into the cell cycle control machinery on the molecular level remains unsolved. Here, we report oscillations of mitochondrial ROS during the cell cycle that target cyclin-dependent kinase 2 (CDK2). Chemical and metabolic interference with ROS production decrease T-loop phosphorylation on CDK2, impeding its full activation and thus efficient DNA replication. ROS regulate CDK2 activity through oxidation of a conserved cysteine residue in close proximity to the T-loop, which prevents binding of the T-loop phosphatase KAP. Together our data reveal how ROS couple mitochondrial metabolism to DNA replication and cell cycle progression, and provide a solution to the longstanding conundrum of how KAP activity towards CDKs can be cell cycle-regulated.

Natural alkaloid galantamine is widely used for the treatment of mild to moderate Alzheimer’s dementia. Galantamine hydrobromide (GH) is available as fast-release tablets, extended-release capsules, and oral solutions. However, its oral delivery can cause some unwanted side effects, such as gastrointestinal disturbances, nausea, and vomiting. Intranasal administration is one possible way to avoid such unwanted effects. In this work, chitosan-based nanoparticles (NPs) were studied as potential GH delivery vehicles for nasal application. The NPs were synthesized via ionic gelation and studied using dynamic light scattering (DLS) as well as by spectroscopic and thermal methods. The GH-loaded chitosan–alginate complex particles were also prepared as a way to modify the release of GH. The high loading efficiency of the GH was confirmed for both types of particles, at 67% for the GH-loaded chitosan NPs and 70% for the complex chitosan/alginate GH-loaded particles. The mean particle size of the GH-loaded chitosan NPs was about 240 nm, while the sodium alginate coated chitosan particles loaded with GH were expectedly bigger, with a mean particle size of ~286 nm. GH release profiles in PBS at 37 °C were obtained for both types of NPs, and it was found that the GH-loaded chitosan NPs allowed the prolonged release of the incorporated drug for a period of 8 h, while the complex GH-loaded chitosan/alginate NPs released the incorporated GH faster. The stability of the prepared GH-loaded NPs was also demonstrated after 1 year of storage at 5 °C ± 3 °C.

Reactive oxygen species (ROS) at the right concentration promote cell proliferation in cell culture, stem cells, and model organisms. However, the mystery of how ROS signaling is coordinated with cell cycle progression and integrated into the cell cycle control machinery on the molecular level remains unsolved. Here, we report increasing levels of mitochondrial ROS during the cell cycle in human cell lines that target cyclin-dependent kinase 2 (CDK2). Chemical and metabolic interferences with ROS production decrease T-loop phosphorylation on CDK2 and so impede its full activation and thus its efficient DNA replication. ROS regulate CDK2 activity through the oxidation of a conserved cysteine residue near the T-loop, which prevents the binding of the T-loop phosphatase KAP. Together, our data reveal how mitochondrial metabolism is coupled with DNA replication and cell cycle progression via ROS, thereby demonstrating how KAP activity toward CDKs can be cell cycle regulated.

Nowadays, the modern pharmaceutical investigations are directed toward obtaining of new polymer micro- and nano-sized drug delivery carriers. In this respect, the use of hydrogel carriers based on polyzwitterions (PZIs) is an opportunity in the preparation of polymer drug delivery systems with desired characteristics. This paper describes the synthesis and characterization of micro-structured p(VA-co- DMAPS) systems with different compositions in situ-loaded with Ibuprofen by emulsifier-free emulsion copolymerization (EEC) in water. The mean size of the prepared microparticles was measured by SEM and particles have been visualized by AFM. The inclusion of Ibuprofen in the polyzwitterionic copolymer microgel systems was established by using DSC. In vitro, drug release experiments were carried out in order to estimate the ability of the obtained microgels to modify the release of water-insoluble Ibuprofen.

This paper describes the synthesis and characterization of cationic copolymers with different macromolecular architecture and drug delivery properties of the corresponding dexamethasone sodium phosphate (DSP)-loaded systems. Copolyelectrolytes comprising poly[2-(acryloyloxy)ethyl] trimethylammonium chloride (PAETMAC) and poly(ethylene glycol) blocks as well as a tri-arm star-shaped PAETMAC were synthesized using cerium(IV) ion-mediated polymerization method. The obtained copolyelectrolytes and corresponding ionic associates with DSP have been characterized by 1H NMR, Fourier Transform Infrared spectroscopy, and differential scanning calorimetry. The average diameter, size distribution, and ζ-potential of the copolymers and DSP–copolymer ionic associates were determined by dynamic light scattering, and particles were visualized by scanning electron microscopy and transmission electron microscopy. The biocompatibility and cytotoxicity of obtained copolymers were determined. In vitro drug release experiments were carried out to estimate the ability of the obtained nanoparticles for sustained release of DSP for a period of 24 h. © 2014 Wiley Periodicals, Inc. and the American Pharmacists Association J Pharm Sci 103:2406–2413, 2014

Protein crystallography, being one of the most established methods for structure determination of biomacromolecules, relies on the diffraction analysis using X-rays for 3D (micro-)crystals. Electron diffraction is only employed for single layer 2D nano-crystals. However, there is no established method for analysing multi-layered 3D nano-sized protein crystals. Electron diffraction may fill this important niche, but several problems have to be surmounted for this method to become mainstream. Our group aims tackling some of the bottlenecks, and although our work is still very much in progress, we can report advances in some important areas. Here we summarise improvements in (i) the induction of growth of (nano-)crystals, (ii) electron diffraction data collection using the Medipix quantum area detector, (iii) unit cell determination using single, non- oriented diffraction patterns, (iv) integration of diffraction data and (v) phasing of electron diffraction data.

Electron diffractograms are lattice images of crystalline samples taken in transmission electron microscopy for molecular structure determination studies. Electron diffraction is a technique widely used in material science and recently it is gaining significance also in life science for studying 2D and 3D organic crystals. However, often the images suffer from strong background noise, masking the data points. Moreover, they suffer also from the strong center beam exposure or a big beam- stop which covers a lot of useful information. This paper presents a user-friendly peak search program in which an autocorrelation algorithm is utilized creatively to intensify the signal and to center the image in the particular regular lattice. An adaptive background removal algorithm is designed to remove the central beam and to reduce the background noise. The latter algorithm can be used for a wide range of applications, such as 2D spectral analysis in physics, NMR Analysis, stars recognition of aerospace photographs.

The typical resolution of three-dimensional reconstruction by cryo-EM single particle analysis is now being pushed up to and beyond the nanometer scale. Correction of the contrast transfer function (CTF) of electron microscopic images is essential for achieving such a high resolution. Various correction methods exist and are employed in popular reconstruction software packages. Here, we present a novel approximation method that corrects the amplitude modulation introduced by the contrast transfer function by convoluting the images with a piecewise continuous function. Our new approach can easily be implemented and incorporated into other packages. The implemented method yielded higher resolution reconstructions with data sets from both highly symmetric and asymmetric structures. It is an efficient alternative correction method that allows quick convergence of the 3D reconstruction and has a high tolerance for noisy images, thus easing a bottleneck in practical reconstruction of macromolecules.

Relative to the number of elastic scattering events, X-rays are three orders of magnitude more damaging to organic molecules than electrons, so for structure determination of 3D nano-crystals of proteins and other damage-prone molecules, electron diffraction should be an attractive alternative. For 2D protein crystals, electron diffraction is the only option. For 3D protein crystals, limitations in collecting diffraction data of sufficient quality and dynamical scattering have so far frustrated the promise electrons hold in this respect. Here we report that these problems could, to a certain extent, be overcome by combining modern flash-freezing techniques, low dose diffraction techniques (microdiffraction) and precession of the electron beam. Our procedures, specifically aimed at gathering high-resolution, 3D reciprocal space data, allowed electron diffraction up to 2.1 A resolution of 3D protein nano-crystals (lysozyme, see figure 1) and even beyond 1 A resolution for nano-crystals of complex organic pharmaceuticals. The resulting diffraction patterns of non-oriented crystals could be indexed.

The drive for elucidation of important macromolecular structures to high resolution in their 3D native or near-native state places continuously higher demands on the quality of the experimental data. For instance, recording of diffraction patterns good enough for structural studies from cryo-preserved bio-macromolecules at low dose conditions remains challenging and highly desirable. The emergence of hybrid pixel detectors opens up new possibilities for direct electron detection and superior detector performance. Here, we report on the characteristics of the Medipix2 detector in diffraction studies, with a special focus on the reliability of the intensities acquired in very low dose conditions. Diffraction data recorded on a Medipix2 detector were assessed in refinement analysis. R-factors lower than 10% were obtained from data recorded at electron dose of 0.05 el/A2. The reproducibility of the data was also shown to be high, given the correlation coefficient of the intensities being higher than 0.9970. The contrast that could be achieved at very low dose conditions was at least an order of magnitude better than that of image plates, based on a direct comparison.