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In the present research, thermal investigation of amorphous solid dispersions (ASDs) was performed in order to compare their relaxation transitions. Two different active pharmaceutical ingredients (APIs), namely naproxen (NAP) and spironolactone (SPIR), were used as model drugs and combined with polyvinylpyrrolidone vinyl acetate (PVPVA64) copolymer to form stable ASDs. The reason for the API selection was their different interacting potentials based on the presence (in the case of NAP) and the lack of H-bond donor group (in the case of SPIR). Determination of both Tg and sub-Tg transitions suggested that ASDs containing NAP and PVPVA64 are 'interacting' systems with respect to the H-bonding. Besides, temperature-dependent Raman spectroscopic experiments confirmed that the naphthalene ring of the NAP has a significant role in the sub-Tg relaxations. In contrast, SPIR-PVPVA64 systems proved to be 'non-interacting' ASDs according to the MDSC, TSDC, and Raman measurements. This study highlights that the combination of the results of different thermoanalytical investigations with spectroscopic methods helps to understand the molecular background of the relaxations in ASDs and thus contributes to the conscious design of stable amorphous pharmaceuticals in the early stage of development. [ABSTRACT FROM AUTHOR]

Metal organic frameworks (MOFs) and particularly copper benzene-1,3,5-tricarboxylate (HKUST-1) are excellent materials for gas storage (e.g., CH4, N2, H2 adsorption) and gas separation. In this work, reduced graphene oxide (RGO)–HKUST-1 mechanical mixtures were studied in order to reveal the effect of RGO content on the pressure tolerance of the texture and heat conductivity. HKUST-1 was obtained by two different synthesis routes. Air-dried MOF and RGO were thoroughly mixed prior to the compression. Powder XRD and Raman spectroscopy were used to characterize the response of the crystal structure, while low-temperature nitrogen adsorption was used the follow the adsorption properties of the pellets. Finally, the "flash" heat pulse method was used to assess the thermal properties. The gas adsorption isotherms revealed that the adsorption capacity decreases when RGO is added. Based on Raman and XRD results, we found that the synthesis route has an effect on multiple scales. We experimentally confirmed that evaluation of the thermal diffusivity requires a model more complex than the simple Fourier equation, due to the inherent heterogeneous structure of the material. A good approximation of the Fourier coefficient of thermal diffusivity was obtained using the parameters of the Guyer–Krumhansl equation. The heat pulse experiments also revealed possible size-dependent behavior. [ABSTRACT FROM AUTHOR]

Purpose: In continuous manufacturing (CM), the material traceability and process dynamics can be investigated by residence time distribution (RTD). Many of the unit operations used in the pharma industry were characterized by dead time–dominated RTD. Even though feasible and proper feedback control is one of the many advantages of CM, its application is challenging in these cases. This study aims to develop a feedback control, implementing the RTD in a Smith predictor control structure in a continuous powder blender line. Methods: Continuous powder blending was investigated with near-infrared spectroscopy (NIR), and the blending was controlled through a volumetric feeder. A MATLAB GUI was developed to calculate and control the concentration of the API based on the chemometric evaluation of the spectra. The programmed GUI changed the feeding rate based on the proportional integral derivative (PID) and the Smith predictor, which implemented the RTD of the system. The control structures were compared even on a system with amplified dead time. Results: In this work, the control structure of the Smith control was devised by utilizing the RTD of the system. The Smith control was compared to a classic PI control structure on the normal system and on an increased dead time system. The Smith predictor was able to reduce the response time for various disturbances by up to 50%, and the dead time had a lower effect on the control. Conclusions: Implementing the RTD models in the control structure improved the process design and further expanded the wide range of applications of the RTD models. Both control structures were able to reduce the effect of disturbances on the system; however, the Smith predictor presented more reliable and faster control, with a wider space for control tuning. [ABSTRACT FROM AUTHOR]

Epithelial ion and fluid secretion determine the physiological functions of a broad range of organs, such as the lung, liver, or pancreas. The molecular mechanism of pancreatic ion secretion is challenging to investigate due to the limited access to functional human ductal epithelia. Patient-derived organoids may overcome these limitations, however direct accessibility of the apical membrane is not solved. In addition, due to the vectorial transport of ions and fluid the intraluminal pressure in the organoids is elevated, which may hinder the study of physiological processes. To overcome these, we developed an advanced culturing method for human pancreatic organoids based on the removal of the extracellular matrix that induced an apical-to-basal polarity switch also leading to reversed localization of proteins with polarized expression. The cells in the apical-out organoids had a cuboidal shape, whereas their resting intracellular Ca2+ concentration was more consistent compared to the cells in the apical-in organoids. Using this advanced model, we demonstrated the expression and function of two novel ion channels, the Ca2+ activated Cl− channel Anoctamin 1 (ANO1) and the epithelial Na+ channel (ENaC), which were not considered in ductal cells yet. Finally, we showed that the available functional assays, such as forskolin-induced swelling, or intracellular Cl− measurement have improved dynamic range when performed with apical-out organoids. Taken together our data suggest that polarity-switched human pancreatic ductal organoids are suitable models to expand our toolset in basic and translational research. [ABSTRACT FROM AUTHOR]

Industry 4.0 has started to transform the manufacturing industries by embracing digitalization, automation, and big data, aiming for interconnected systems, autonomous decisions, and smart factories. Machine learning techniques, such as artificial neural networks (ANN), have emerged as potent tools to address the related computational tasks. These advancements have also reached the pharmaceutical industry, where the Process Analytical Technology (PAT) initiative has already paved the way for the real-time analysis of the processes and the science- and risk-based flexible production. This paper aims to assess the potential of ANNs within the PAT concept to aid the modernization of pharmaceutical manufacturing. The current state of ANNs is systematically reviewed for the most common manufacturing steps of solid pharmaceutical products, and possible research gaps and future directions are identified. In this way, this review could aid the further development of machine learning techniques for pharmaceutical production and eventually contribute to the implementation of intelligent manufacturing lines with automated quality assurance. [ABSTRACT FROM AUTHOR]

Plant pathogenic fungi are responsible for enormous crop losses worldwide. Overcoming this problem is challenging as these fungi can be highly resistant to approved chemical fungicides. There is thus a need to develop and introduce fundamentally new plant and crop protection strategies for sustainable agricultural production. Highly stable extracellular antifungal proteins (AFPs) and their rationally designed peptide derivatives (PDs) constitute feasible options to meet this challenge. In the present study, their potential for topical application to protect plants and crops as combinatorial biofungicides is supported by the investigation of two Neosartorya (Aspergillus) fischeri AFPs (NFAP and NFAP2) and their γ-core PDs. Previously, the biofungicidal potential of NFAP, its rationally designed γ-core PD (γNFAP-opt), and NFAP2 was reported. Susceptibility tests in the present study extended the in vitro antifungal spectrum of NFAP2 and its γ-core PD (γNFAP2-opt) to Botrytis, Cladosporium, and Fusarium spp. Besides, in vitro additive or indifferent interactions, and synergism were observed when NFAP or NFAP2 was applied in combination with γNFAP-opt. Except for γNFAP2-opt, the investigated proteins and peptides did not show any toxicity to tomato plant leaves. The application of NFAP in combination with γNFAP-opt effectively inhibited conidial germination, biofilm formation, and hyphal extension of the necrotrophic mold Botrytis cinerea on tomato plant leaves. However, the same combination only partially impeded the B. cinerea-mediated decay of tomato fruits, but mitigated the symptoms. Our results highlight the feasibility of using the combination of AFP and PD as biofungicide for the fungal infection control in plants and crops. [ABSTRACT FROM AUTHOR]

The work aimed to develop the Absorption Driven Drug Formulation (ADDF) concept, which is a new approach in formulation development to ensure that the drug product meets the expected absorption rate. The concept is built on the solubility-permeability interplay and the rate of supersaturation as the driving force of absorption. This paper presents the first case study using the ADDF concept where not only dissolution and solubility but also permeation of the drug is considered in every step of the formulation development. For that reason, parallel artificial membrane permeability assay (PAMPA) was used for excipient selection, small volume dissolution-permeation apparatus was used for testing amorphous solid dispersions (ASDs), and large volume dissolution-permeation tests were carried out to characterize the final dosage forms. The API-excipient interaction studies on PAMPA indicated differences when different fillers or surfactants were studied. These differences were then confirmed with small volume dissolution-permeation assays where the addition of Tween 80 to the ASDs decreased the flux dramatically. Also, the early indication of sorbitol's advantage over mannitol by PAMPA has been confirmed in the investigation of the final dosage forms by large-scale dissolution-permeation tests. This difference between the fillers was observed in vivo as well. The presented case study demonstrated that the ADDF concept opens a new perspective in generic formulation development using fast and cost-effective flux-based screening methods in order to meet the bioequivalence criteria. [ABSTRACT FROM AUTHOR]

This study examined the concentrations of Al, Ca, Cd, Cr, Cu, Fe, K, Mg, Mn, Na, Ni, Pb, and Zn in the liver and kidney samples of golden jackals (n = 163) and red foxes (n = 64). Additionally, we studied how sex affected element concentration in both species, and in the case of golden jackals, how concentration levels were affected by age. Liver was the most important storage organ for Cu, Fe, Mn, and Zn, while kidney for Cd and Na in jackals and foxes. We found no sex-related differences in concentration of any of the selected elements in fox samples. In jackals, the mean concentrations of Al, Mn, and Zn were significantly higher in kidneys of males, while Zn and Cu had higher concentrations in female livers. Our results suggest bioaccumulation processes do not occur in mesocarnivores. Lead and Cd concentrations fell within ranges accepted as normal for canine species and were below toxic levels in the organs of the examined species. Concentrations of Cr, Cu, Fe, Mn, and Zn were far below the toxic levels in both organs of both species, with a slight sign of deficiency of Fe, Mn, and Zn in jackal and fox kidneys. In comparison with toxicity levels and results from other European study sites, the current study demonstrated that species living in the examined area of Somogy County, Hungary, are generally exposed to low levels of environmental contamination. [ABSTRACT FROM AUTHOR]

Detailed vibrational (IR, Raman, far-IR) and thermal (TGA, TG–MS, DSC) analysis has been performed on di[κ1O,κ2O-carbonatotetraamminecobalt(III)] sulfate trihydrate, ([Co(NH3)4CO3]2SO4·3H2O (1). Its isothermic heating at 100 °C leads to formation of [Co(NH3)4CO3]2SO4 (compound 2). UV and IR studies showed that the distorted octahedral arrangement around cis-O2CoN4 core in compound 1 does not change during dehydration, which explains the reversible water loss and ability of compound 2 to rehydrate into compound 1. Compound 2 decomposes at ~ 240 °C in inert atmosphere giving final decomposition products, which are two modifications of nanosized metallic cobalt (hcp-15 nm, fcc-250 nm) and CoO (55 nm). The redox reaction results in N2 as an ammonia oxidation product. The decomposition intermediate is a cobalt(II) compound, Co2O1,14+δ(SO4)0.86 (δ = the oxygen surplus due to the presence of 2.8% of Co(III) ion). The same reaction in air atmosphere resulted in Co2O1.25+δ(SO4)0.75 (δ = the oxygen surplus due to the presence of 5.3% of Co(III) ion (compound 3a). Compound 3a is oxidized in air at 793 °C into Co3O4. The compound 3a exhibits catalytic activity in photodegradation in Congo red. The photodegradation process follows pseudo-first-order kinetic (kapp = 1.0 and 7.0. at pH = 3.4 and 5.25, respectively). [ABSTRACT FROM AUTHOR]

We present facile methods to obtain purified sporopollenin exine capsules, and provide mass balances for classical and novel purification procedures. An ionic liquid, tetrabutyl phosphonium hydroxide turned out to be the most effective in removing the intine wall. The sporopollenin capsules were investigated by fluorescent microscopy, AFM, solid-state NMR and infrared Raman spectroscopy. The latter two methods showed that sunflower and rape exines have different proportions of O-aliphatic and aromatic constituents. Purified exine capsules were coated with functionalized fluorophores. The procedures presented in this paper could contribute to further spread of the applications of this hollow, and chemically highly resistant material. [ABSTRACT FROM AUTHOR]

Graphene oxide (GO) is not only an intermediate in top-down graphene preparation, but it is also acknowledged for its convenient application when the hydrophobicity of graphene is a drawback. Its preparation through the wet exfoliation of graphite inevitably leads to a product lacking several of the outstanding features of graphene. The destroyed graphene structure can be at least partially reconstructed by thermal and/or wet chemical reduction. Thermally reduced graphene oxide (TRGO) was obtained by mild heat treatment (300 °C) in argon, while chemically reduced graphene oxide (CRGO) was prepared by using a green reagent, ascorbic acid (AA). The parent GO and the two GO derivatives were compared by thermogravimetric analysis coupled with mass spectroscopy and Raman spectroscopy. Both reductive treatments resulted in enhanced thermal stability and higher C/O ratio. Although the chemical reduction with AA was more efficient in the elimination of the O functionalities, it also resulted in significant fragmentation of the graphene-like platelets. [ABSTRACT FROM AUTHOR]

Recent advancements in deep learning have revolutionized the way microscopy images of cells are processed. Deep learning network architectures have a large number of parameters, thus, in order to reach high accuracy, they require a massive amount of annotated data. A common way of improving accuracy builds on the artificial increase of the training set by using different augmentation techniques. A less common way relies on test-time augmentation (TTA) which yields transformed versions of the image for prediction and the results are merged. In this paper we describe how we have incorporated the test-time argumentation prediction method into two major segmentation approaches utilized in the single-cell analysis of microscopy images. These approaches are semantic segmentation based on the U-Net, and instance segmentation based on the Mask R-CNN models. Our findings show that even if only simple test-time augmentations (such as rotation or flipping and proper merging methods) are applied, TTA can significantly improve prediction accuracy. We have utilized images of tissue and cell cultures from the Data Science Bowl (DSB) 2018 nuclei segmentation competition and other sources. Additionally, boosting the highest-scoring method of the DSB with TTA, we could further improve prediction accuracy, and our method has reached an ever-best score at the DSB. [ABSTRACT FROM AUTHOR]

A simple synthetic method was developed to prepare 4[Agpy2ClO4]·[Agpy4]ClO4 in a low-temperature decomposition process of [Agpy4]ClO4. A detailed IR, Raman and far-IR study including factor group analysis has been performed, and the assignation of bands is given. The compound decomposes quickly with a multistep ligand loss process with the formation of [Agpy2]ClO4 and AgClO4 intermediates and AgCl as an end product around ~ 85, ~ 350 and 450 °C, respectively. During the first decomposition step, a small fraction of the ligands is lost in a redox reaction: perchlorate oxidizes the pyridine, forming carbon, carbon dioxide, water and NO, while it itself is reduced into AgCl. In the next step, when AgClO4 forms after complete ligand loss and reacts with the carbon formed in the degradation of pyridine at lower temperatures and produces NO, CO2 and H2O. This reaction becomes possible because the AgCl formed in the redox reactions makes a eutectic melt with AgClO4 in situ, which is a favorable medium for the carbon oxidation reaction. AgCl is known to reduce the temperature of decomposition of AgClO4, in which process forms AgCl as well as O2 and so is an autocatalytic process. The loss and degradation of pyridine ligand are endothermic; the redox reactions including carbon oxidation and AgClO4 decomposition into AgCl and O2 are exothermic. The amount of absorbed/evolved heats corresponding to these processes was determined by DSC both under N2 and O2 atmospheres. [ABSTRACT FROM AUTHOR]

Synthesis of ammonium hydroxodisulfitoferriate(III), (diammonium catena-{bis(μ2-sulfito-κO,κO)-μ2-hydroxo-κ2O}ferrate(III) monohydrate) (NH4)2[Fe(OH)(SO3)2]·H2O (compound 1) and its thermal behavior is reported. The compound is stable in air. Its thermal decomposition proceeds without the expected quasi-intramolecular oxidation of sulfite ion with ferric ions. The disproportionation reaction of the ammonium sulfite, formed from the evolved NH3, SO2 and H2O in the main decomposition stage of 1, results in the formation of ammonium sulfate and ammonium sulfide. The ammonium sulfide is unstable at the decomposition temperature of 1 (150 °C) and transforms into NH3 and H2S which immediately forms elementary sulfur by reaction with SO2. The formation and decomposition of other intermediate compounds like (NH4)2SnOx (n = 2, x = 3 and n = 3, x = 6) results in the same decomposition products (S, SO2 and NH3). Two basic iron sulfates, formed in different ratios during synthesizing experiments performed under N2 or in the presence of air, have been detected as solid intermediates which contain ammonium ions. The final decomposition product was proved to be α-Fe2O3 (mineral name hematite). [ABSTRACT FROM AUTHOR]

Several previous study results have already demonstrated that golden jackal and red fox may serve as biological indicators of trace elements and heavy metal concentrations in the various regions they inhabit. The aim of this study was to evaluate accumulation patterns of targeted elements (Al, Ca, Cr, Cu, Fe, Mg, Mn, Ni and Pb) in liver samples of red foxes and golden jackals collected during the same period in the southern part of Romania. The accumulation patterns of trace elements in the livers of sympatric golden jackal and red fox were practically the same. To date, separate studies of the species individually in different habitats have shown that either of the species can be used for ecotoxicological and biomonitoring studies. Moreover, in general gender related studies, no significant differences in the concentrations of the investigated elements were found in either jackals or foxes. Also, average metal concentrations in liver samples do not show significant differences between groups under and above 12 months of age. [ABSTRACT FROM AUTHOR]

We introduce a novel procedure that uses dynamic 3-D computer graphics as a diagnostic tool for assessing disease severity in schizophrenia patients, based on their reduced influence of top-down cognitive processes in interpreting bottom-up sensory input. Our procedure uses the hollow-mask illusion, in which the concave side of the mask is misperceived as convex, because familiarity with convex faces dominates sensory cues signaling a concave mask. It is known that schizophrenia patients resist this illusion and their resistance increases with illness severity. Our method uses virtual masks rendered with two competing textures: (a) realistic features that enhance the illusion; (b) random-dot visual noise that reduces the illusion. We control the relative weights of the two textures to obtain psychometric functions for controls and patients and assess illness severity. The primary novelty is the use of a rotating mask that is easy to implement on a wide variety of portable devices and avoids the use of elaborate stereoscopic devices that have been used in the past. Thus our method, which can also be used to assess the efficacy of treatments, provides clinicians the advantage to bring the test to the patient's own environment, instead of having to bring patients to the clinic. [ABSTRACT FROM AUTHOR]

Long-standing evidence indicates that human immunodeficiency virus type 1 (HIV-1) preferentially integrates into a subset of transcriptionally active genes of the host cell genome. However, the reason why the virus selects only certain genes among all transcriptionally active regions in a target cell remains largely unknown. Here we show that HIV-1 integration occurs in the outer shell of the nucleus in close correspondence with the nuclear pore. This region contains a series of cellular genes, which are preferentially targeted by the virus, and characterized by the presence of active transcription chromatin marks before viral infection. In contrast, the virus strongly disfavours the heterochromatic regions in the nuclear lamin-associated domains and other transcriptionally active regions located centrally in the nucleus. Functional viral integrase and the presence of the cellular Nup153 and LEDGF/p75 integration cofactors are indispensable for the peripheral integration of the virus. Once integrated at the nuclear pore, the HIV-1 DNA makes contact with various nucleoporins; this association takes part in the transcriptional regulation of the viral genome. These results indicate that nuclear topography is an essential determinant of the HIV-1 life cycle. [ABSTRACT FROM AUTHOR]

The inward rectifier K + current (I K1) plays an important role in terminal repolarization and stabilization of the resting potential in cardiac cells. Although I K1 was shown to be sensitive to changes in intracellular Ca 2+ concentration ([Ca 2+] i), the nature of this Ca 2+ sensitivity—in spite of its deep influence on action potential morphology—is controversial. Therefore, we aimed to investigate the effects of a nonadrenergic rise in [Ca 2+] i on the amplitude of I K1 in canine and human ventricular myocardium and its consequences on cardiac repolarization. I K1, defined as the current inhibited by 10 μM Ba 2+, was significantly increased in isolated canine myocytes following a steady rise in [Ca 2+] i. Enhanced I K1 was also observed when [Ca 2+] i was not buffered by ethylene glycol tetraacetic acid, and [Ca 2+] I transients were generated. This [Ca 2+] i-dependent augmentation of I K1 was largely attenuated after inhibition of CaMKII by 1 μM KN-93. Elevation of [Ca 2+] o in multicellular canine and human ventricular preparations resulted in shortening of action potentials and acceleration of terminal repolarization. High [Ca 2+] o enhanced the action potential lengthening effect of the Ba 2+-induced I K1 blockade and attenuated the prolongation of action potentials following a 0.3-μM dofetilide-induced I Kr blockade. Blockade of I Ks by 0.5 μM HMR-1556 had no significant effect on APD 90 in either 2 mM or 4 mM [Ca 2+] o. It is concluded that high [Ca 2+] i leads to augmentation of the Ba 2+-sensitive current in dogs and humans, regardless of the mechanism of the increase. This effect seems to be at least partially mediated by a CaMKII-dependent pathway and may provide an effective endogenous defense against cardiac arrhythmias induced by Ca 2+ overload. [ABSTRACT FROM AUTHOR]

An amendment to this paper has been published and can be accessed via a link at the top of the paper. [ABSTRACT FROM AUTHOR]

Proarrhythmia, the development of new or worse arrhythmias in response to drug therapy, is a major limitation to the development and use of new drugs. There are different types of drug-induced proarrhythmia, including long-QT syndrome (LQTS), short-QT syndrome and proarrhythmia related to Na+-channel blockade/conduction impairment. By far the most important form of proarrhythmia at present is drug-induced LQTS and its associated characteristic tachyarrhythmia, torsades de pointes (TdP). TdP arises when cellular action potentials (APs) are excessively prolonged, leading to arrhythmogenic afterdepolarizations, especially early afterdepolarizations (EADs), which trigger complex re-entry in a substrate involving increased transmural dispersion of repolarization. In vitro screening, increasingly involving high-throughput assays, is used to assess potential candidate molecules and eliminate potentially problematic structures at an early stage of development. The most commonly used screening assays assess drug block of the K+ current carried by human ether-à-go-go (hERG) subunits, corresponding to the rapid delayed-rectifier K+ channel, the overwhelmingly most common target of TdP-inducing drugs. In addition, the effects of drugs on AP duration or the in vivo equivalent, QT interval, are often assessed in animal models. Methods available for repolarization-related proarrhythmic risk assessment include in vitro (Langendorff-perfused rabbit or guinea pig hearts) and in vivo models (such as a-adrenoceptor-stimulated rabbits, rabbits with reduced repolarization reserve due to block of slow delayed-rectifier current, animals with chronic atrioventricular block or animals with cardiac remodelling caused by congestive heart failure). In silico modelling may be helpful for molecular design of non-hERG blocking candidates and for optimization of compound selection (at the molecular and pharmacological profile levels). Finally, clinical evaluation of effects on electrocardiographic intervals (particularly QT) and cardiac rhythm are often needed, both prior to drug approval and after successful introduction on the market (postmarketing surveillance). The successful avoidance of proarrhythmic complications is a shared responsibility of the innovative pharmaceutical industry, regulatory authorities, partners in the clinical drug development phase and practicing physicians. This paper reviews the principal forms of proarrhythmia and the methods that can be used to minimize the risk of proarrhythmia in drug development and clinical practice, with particular emphasis on the most common and problematic form, acquired LQTS. [ABSTRACT FROM AUTHOR]

SEA0400 is a recently developed inhibitor of the Na+/Ca2+ exchanger (NCX) shown to suppress both forward and reverse mode operation of NCX. Present experiments were designed to study the effect of partial blockade of NCX on Ca handling and contractility in Langendorff-perfused guinea pig hearts loaded with the fluorescent Ca-sensitive dye fura-2. Left ventricular pressure and intracellular calcium concentration ([Ca2+]i) were synchronously recorded before and after cumulative superfusion with 0.3 and 1 μM SEA0400. SEA0400 caused no significant change in the systolic and diastolic values of left ventricular pressure and [Ca2+]i. Accordingly, pulse pressure and amplitude of the [Ca2+]i transient also remained unchanged in the presence of SEA0400. SEA0400 had no influence either on the time required to reach peak values of pressure and [Ca2+]i or on half relaxation time. On the other hand, both 0.3 and 1 μM SEA0400 significantly increased the decay time constant of [Ca2+]i transients, obtained by fitting its descending limb between 30% and 90% of relaxation, from 127 ± 7 to 165 ± 7 and 177 ± 14 ms, respectively ( P < 0.05, n = 6). In contrast to the guinea pig hearts, rat hearts responded to SEA0400 treatment with increased [Ca2+]i transients and contractility. These interspecies differences observed in the effect of SEA0400 can be explained by the known differences in calcium handling between the two species. [ABSTRACT FROM AUTHOR]

Understanding the reaction of living cells in response to different extracellular stimuli, such as hyperosmotic stress, is of primordial importance. Mannitol, a cell-impermeable non-toxic alcohol, has been used successfully for reversible opening of the blood–brain barrier in hyperosmotic concentrations. In this study we analyzed the effect of hyperosmotic mannitol on the shape and surface structure of living cerebral endothelial cells by atomic force microscope imaging technique. Addition of clinically relevant concentrations of mannitol to the culture medium of the confluent cells induced a decrease of about 40% in the observed height of the cells. This change was consistent both at the nuclear and peripheral region of the cells. After mannitol treatment even a close examination of the contact surface between the cells did not reveal gap between them. We could observe the appearance of surface protrusions of about 100 nm. By force measurements the elasticity of the cells were estimated. While the Young’s modulus of the control cells appeared to be 8.04 ± 0.12 kPa, for the mannitol-treated cells it decreased to an estimated value of 0.93 ± 0.04 kPa which points to large structural changes inside the cell. [ABSTRACT FROM AUTHOR]

Exosomes are small extracellular vesicles (sEVs), playing a crucial role in the intercellular communication in physiological as well as pathological processes. Here, we aimed to study whether the melanoma-derived sEV-mediated communication could adapt to microenvironmental stresses. We compared B16F1 cell-derived sEVs released under normal and stress conditions, including cytostatic, heat and oxidative stress. The miRNome and proteome showed substantial differences across the sEV groups and bioinformatics analysis of the obtained data by the Ingenuity Pathway Analysis also revealed significant functional differences. The in silico predicted functional alterations of sEVs were validated by in vitro assays. For instance, melanoma-derived sEVs elicited by oxidative stress increased Ki-67 expression of mesenchymal stem cells (MSCs); cytostatic stress-resulted sEVs facilitated melanoma cell migration; all sEV groups supported microtissue generation of MSC-B16F1 co-cultures in a 3D tumour matrix model. Based on this study, we concluded that (i) molecular patterns of tumour-derived sEVs, dictated by the microenvironmental conditions, resulted in specific response patterns in the recipient cells; (ii) in silico analyses could be useful tools to predict different stress responses; (iii) alteration of the sEV-mediated communication of tumour cells might be a therapy-induced host response, with a potential influence on treatment efficacy. [ABSTRACT FROM AUTHOR]

As the process analytical technology (PAT) mindset is progressively introduced and adopted by the pharmaceutical companies, there is an increasing demand for effective and versatile real-time analyzers to address the quality assurance challenges of drug manufacturing. In the last decades, Raman spectroscopy has emerged as one of the most promising tools for non-destructive and fast characterization of the pharmaceutical processes. This review summarizes the achieved results of the real-time application of Raman spectroscopy in the field of the secondary manufacturing of pharmaceutical solid dosage forms, covering the most common secondary process steps of a tablet production line. In addition, the feasibility of Raman spectroscopy for real-time control is critically reviewed, and challenges and possible approaches to moving from real-time monitoring to process analytically controlled technologies (PACT) are discussed. [ABSTRACT FROM AUTHOR]