Application of 2-hydroxypropyl-β-cyclodextrin in the assay of acyl-CoA: cholesterol acyltransferase and neutral and acid cholesterol ester hydrolases

The utility of 2-hydroxypropyl-β-cyclodextrin for increasing the sensitivity of assays for the microsomal acyl-CoA:cholesterol acyltransferase, and the acid lysosomal and the neutral microsomal and cytosolic cholesterol ester hydrolase activity was studied in rat hepatocytes. Enzyme assays, at optimal concentrations of cyclodextrin, were validated by assessing: (i) linearity of product formation with incubation time and protein amount, and saturation with substrate, and (ii) the effect of treatments of cells or of subcellular fractions on enzyme activities. Delivery of cholesterol dissolved in 2-hydroxypropyl-β-cyclodextrin to the acyl-CoA:cholesterol acyltransferase assay mixture raised the enzyme activity more than 8-fold and was twice that measured when cholesterol was added in Triton WR-1339. 2-Hydroxypropyl-β-cyclodextrin itself was partially effective, apparently by making endogenous cholesterol more accesible to the enzyme. Inclusion of 2-hydroxypropyl-β-cyclodextrin in cholesterol ester hydrolase assays using standard micellar substrates doubled the activity estimated in lysosome and microsome preparations and enhanced the cytosolic cholesterol esterase activity by about 50%. Differences in the catalytic activity of acyl-CoA:cholesterol acyltransferase and cholesterol ester hydrolases caused by treatment of hepatocytes with compound 58-035 or 25-hydroxycholesterol, or of subcellular fractions with NaF, were maintained when enzymes were assayed with cyclodextrin. The results indicate that 2-hydroxypropyl-β-cyclodextrin is a suitable vehicle for delivering cholesterol to acyl-CoA:cholesterol acyltransferase and enhances the sensitivity of standard assays of the enzymes governing the intrahepatic hydrolysis of cholesteryl esters.     

Identification of Potential Antiviral Inhibitors from Hydroxychloroquine and 1,2,4,5-Tetraoxanes Analogues and Investigation of the Mechanism of Action in SARS-CoV-2

This study aimed to identify potential inhibitors and investigate the mechanism of action on SARS-CoV-2 ACE2 receptors using a molecular modeling study and theoretical determination of biological activity. Hydroxychloroquine was used as a pivot structure and antimalarial analogues of 1,2,4,5 tetraoxanes were used for the construction and evaluation of pharmacophoric models. The pharmacophore-based virtual screening was performed on the Molport^® database (~7.9 million compounds) and obtained 313 structures. Additionally, a pharmacokinetic study was developed, obtaining 174 structures with 99% confidence for human intestinal absorption and penetration into the blood–brain barrier (BBB); posteriorly, a study of toxicological properties was realized. Toxicological predictions showed that the selected molecules do not present a risk of hepatotoxicity, carcinogenicity, mutagenicity, and skin irritation. Only 54 structures were selected for molecular docking studies, and five structures showed binding affinity (ΔG) values satisfactory for ACE2 receptors (PDB 6M0J), in which the molecule MolPort-007-913-111 had the best ΔG value of −8.540 Kcal/mol, followed by MolPort-002-693-933 with ΔG = −8.440 Kcal/mol. Theoretical determination of biological activity was realized for 54 structures, and five molecules showed potential protease inhibitors. Additionally, we investigated the Mpro receptor (6M0K) for the five structures via molecular docking, and we confirmed the possible interaction with the target. In parallel, we selected the TopsHits 9 with antiviral potential that evaluated synthetic accessibility for future synthesis studies and in vivo and in vitro tests.     

Mesothelin Expression Is Not Associated with the Presence of Cancer Stem Cell Markers SOX2 and ALDH1 in Ovarian Cancer

Mesothelin (MSLN) overexpression (OE) is a frequent finding in ovarian carcinomas and increases cell survival and tumor aggressiveness. Since cancer stem cells (CSCs) contribute to pathogenesis, chemoresistance and malignant behavior in ovarian cancer (OC), we hypothesized that MSLN expression could be creating a favorable environment that nurtures CSCs. In this study, we analyzed the expression of MSLN and CSC markers SOX2 and ALDH1 by immunohistochemistry (IHC) in different model systems: primary high-grade serous carcinomas (HGSCs) and OC cell lines, including cell lines that were genetically engineered for MSLN expression by either CRISPR-Cas9-mediated knockout (Δ) or lentivirus-mediated OE. Cell lines, wild type and genetically engineered, were evaluated in 2D and 3D culture conditions and xenografted in nude mice. We observed that MSLN was widely expressed in HGSC, and restricted expression was observed in OC cell lines. In contrast, SOX2 and ALDH1 expression was limited in all tissue and cell models. Most importantly, the expression of CSC markers was independent of MSLN expression, and manipulation of MSLN expression did not affect CSC markers. In conclusion, MSLN expression is not involved in driving the CSC phenotype.     

Silk fibroin membranes with self-assembled globular structures for controlled drug release

In this paper, we have evaluated the incorporation of a drug model and its release from silk fibroin (SF) membranes, analyzing the morphological, chemical, barrier, and biological properties. SF self-assembled into stable globular structures, encapsulating the drug, when diclofenac sodium (DS) was incorporated into SF solution prior to membranes preparation. The membranes showed biostatic action and prevented microorganism permeation. Kinetic studies indicate that DS was released in 120 min, with Fickian diffusion as the main mechanism of release. Results of this paper emphasize the potential of SF in wound healing, with good barrier and biological properties. © 2019 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 2020 , 137, 48763.     

The new Bucharest: Two decades of restructuring

The Romanian capital became well-known under communism for major construction and redevelopment projects. Bucharest’s post-communist reconstruction has seen a major sectoral change from industry to commerce, backed by numerous building projects for housing, shops, offices and hotels. The urban landscape has changed in many parts of the downtown and outer suburban areas, especially with the intrusion of tall buildings into areas of pre-communist development. The approach has been quite permissive towards private enterprise, absent a clear vision of a 21st century city. Although development has not been excessively chaotic, there is a clear case for more effective regulation as well as greater attention to infrastructure in privately-managed utilities and in the public transport system.     

Metabolite composition of chestnut (Castanea sativa Mill.) upon cooking: Proximate analysis,fibre, organic acids and phenolics

The aim of this research was to study the processing effects (roasting and boiling) on primary and secondary metabolite composition of fruits from the following chestnut (Castanea sativa Mill.) cultivars (cvs.) of three Protected Designation of Origin (PDO) areas in the Trás-os-Montes e Alto Douro region (Portugal): PDO Terra Fria (cvs. Aveleira, Boaventura, Côta, Lamela and Trigueira), PDO Padrela (cvs. Judia, Lada, Longal and Negra) and PDO Soutos da Lapa (cvs. Longal and Martaínha). The cooking processes significantly (p < 0.0001) affected primary and secondary metabolite composition of the chestnuts. Roasted chestnuts had higher protein contents, insoluble and total dietary fibre and lower fat contents whilst boiled chestnuts had lower protein, but higher fat contents. Cooking increased citric acid contents, especially in roasted chestnuts. On the other hand, raw chestnuts had higher malic acid contents than cooked chestnuts. Moreover, roasted chestnuts had significantly higher gallic acid and total phenolics contents, and boiled chestnuts had higher gallic and ellagic acids contents, when compared to raw chestnuts. The present data confirms that cooked chestnuts are a good source of organic acids and phenolics and have low fat contents, properties that are associated with positive health benefits.     

Phase Diagrams of the LnAlO3–CaTiO3 (Ln = Nd, Sm) Systems and Polymorphism of CaTiO3

The phase equilibria in the NdAlO₃–CaTiO₃ and SmAlO₃–CaTiO₃ systems are investigated and their phase diagrams are constructed for the first time. The polymorphism of calcium titanate is studied.     

A Bayesian stochastic programming approach to an employee scheduling problem

Bayesian forecasting models provide distributional estimates for random parameters, and relative to classical schemes, have the advantage that they can rapidly capture changes in nonstationary systems using limited historical data. Unlike deterministic optimization, stochastic programs explicitly incorporate distributions for random parameters in the model formulation, and thus have the advantage that the resulting solutions more fully hedge against future contingencies. In this paper, we exploit the strengths of Bayesian prediction and stochastic programming in a rolling-horizon approach that can be applied to solve real-world problems. We illustrate the methodology on an employee production scheduling problem with uncertain up-times of manufacturing equipment and uncertain production rates. Computational results indicate the value of our approach.     

Low firing temperature ceramic for microcircuit substrates

Methods for fabricating substrates based on low-temperature (firing temperature below 1000°C) ceramic, fired together with a deposited metal coating, are examined. Materials which are promising for use as low-temperature simultaneously fired ceramic (LSC) in the systems B₂O₃-CaO-SiO₂ and B₂O₃-CaO-Al₃O₃ are proposed.     

Specific [3H]8-OH-DPAT binding in brain regions of rats genetically predisposed to various defense behavior strategies

BACKGROUND AND PURPOSE: We examined the 20-or-more-year survival and functional levels of 148 stroke survivors and 148 age- and sex-matched control subjects from the Framingham Study Cohort, whom we originally studied in 1972-1974 to ascertain the survival and disability status of stroke survivors compared with that of controls. METHODS: This long-term evaluation was done with use of data from the 1993-1995 Framingham Study Cohort Examination 23 on the 10 stroke survivors and 20 control subjects still living to identify and compare the host characteristics and functional status of each group. The survival curves for both stroke survivors and controls were derived from the ongoing Framingham Study database. RESULTS: Twenty-plus-year stroke survivors experienced a greater mortality than age- and sex-matched controls (92.5% and 81%, respectively). The slopes of the two survival curves were essentially the same. Functional status (eg, walking and independence in activities of daily living) of stroke survivors, however, compared very favorably with that of the control subjects. Stroke survivors were more likely to be female and to have a number of comorbidities, including elevated blood pressures, greater use of medications, less use of alcohol, and less depressive symptomology. CONCLUSIONS: In the Framingham cohort, 20-plus-year stroke survivors showed greater mortality than age- and sex-matched control subjects; functionally, however, the groups were very similar and in general quite independent.