НОВЫЙ ПОДХОД К ХРОМАТОГРАФИЧЕСКОМУ ОПРЕДЕЛЕНИЮ РАСТВОРИМОСТИ КВЕРЦЕТИНА В ВОДЕ

Significant discrepancies of literature data on the quercetin (Q) water solubility (the variations exceed 5000 times!)  at the ambient temperature prompted the elaboration of a new approach on its determination. The new approach is based not on the single measurements, but on the revealing and com­paring the two dependencies of solubility. First of them is the dependence of the solubility on pH of water solutions S(Q) = a×рН + b, followed by S(Q) interpolation on рН = 7, which corresponds to the pure water. The second one is the dependence of the solubility on acetonitrile content in acetonit­rile-water mixtures, log S(Q) = a×[CH3CN] + b, followed by log S(Q) extrapolation on the zero concentration of organic solvent, which corresponds to the pure water as well. The analysis of the solutions was fulfilled using the reversed phase high performance liquid chromatography (RP HPLC). The results obtained independently with various methods correspond well with each other: their joint average value for the few samples was 7.1 ± 1.6 mg L-1 (pS = 4.63). Since the results were obtained for quercetin samples that contained different amounts of crystalline and hydrate water, the final sub-routine of analytical procedure was their extrapolation on the zero-water content in the quercetin samples. The necessity of the preliminary control of both the water content and the organic im­pu­rities in the samples using elemental and HPLC analysis was discussed. The samples of quercetin, as well as other plant extractive substances, could contain impurities that distort the results of its water solubility.Keywords: Quercetin, water solubility, pH-dependence, dependence on concentration of organic constituent, extrapolation, HPLCDOI: http://dx.doi.org/10.15826/analitika.2019.23.3.013(Russian)1Igor G. Zenkevich, 1,2Dmitrii A. Olisov1, 2Roman V. Shafigulin, 2Andzhela V. Bulanova 1St. Petersburg State University, Institute for Chemistry, Universitetskii prosp., 26,St. Petersburg 198504, Russian Federation2Samara University; Moscow road, 34, Samara 443086, Russian FederationЗначительный разброс литературных данных по растворимости кверцетина (Q) в воде при комнатной температуре (более чем в 5000 раз!) заставляет пред­ло­жить новый подход к определению этой характеристики. Он основан не на еди­ничных определениях, а на выявлении и со­поставлении двух зависи­мостей рас­т­воримости кверцетина. Первая из них – зависимость растворимости от рН водных раство­ров, S(Q) = a×рН + b, с последующей интер­поляцией S(Q) на вели­чину рН = 7, соответствующую чистой воде. Вторая – зависимость растворимости от содержания ацетонитрила в водно-ацето­нит­ри­льных раство­рах, lg S(Q) = a×[CH3CN] + b, с экстраполяцией lg S(Q) на нулевую кон­цен­трацию органи­че­с­кого растворителя, также соответствующую чистой воде. Для анализа растворов использован метод обращенно-фазовой ВЭЖХ. Результаты, независимо определенные разными методами, согласуются между собой: их общее среднее S(Q) для не­скольких образцов равно 7.1 ± 1.6 мг/л (pS = 4.63). Поскольку полученные результаты относятся к образцам Q, содержащим различные количества кристаллизационной и гидратной воды, то заключительной стадией определений является их экст­ра­поляция на нулевое содержание воды в образцах. Обсуждается необходимость предварительного контроля содержания воды и органических примесей в образцах по данным элементного и ВЭЖХ ана­лиза. Образцы кверцетина, как и других экстрактивных веществ растений, могут содержать примеси, искажающие результаты определения растворимости.Ключевые слова: Кверцетин, растворимость в воде, зависимость от рН, зависимость от концентра­ции органического компонента раствора, экстрапо­ля­ция, высоко­эф­фек­тивная жидкост­ная хроматографияDOI: http://dx.doi.org/10.15826/analitika.2019.23.3.01

Mechanism of filopodia initiation by reorganization of a dendritic network

Afilopodium protrudes by elongation of bundled actin filaments in its core. However, the mechanism of filopodia initiation remains unknown. Using live-cell imaging with GFP-tagged proteins and correlative electron microscopy, we performed a kinetic-structural analysis of filopodial initiation in B16F1 melanoma cells. Filopodial bundles arose not by a specific nucleation event, but by reorganization of the lamellipodial dendritic network analogous to fusion of established filopodia but occurring at the level of individual filaments. Subsets of independently nucleated lamellipodial filaments elongated and gradually associated with each other at their barbed ends, leading to formation of cone-shaped structures that we term Λ-precursors. An early marker of initiation was the gradual coalescence of GFP-vasodilator-stimulated phosphoprotein (GFP-VASP) fluorescence at the leading edge into discrete foci. The GFP-VASP foci were associated with Λ-precursors, whereas Arp2/3 was not. Subsequent recruitment of fascin to the clustered barbed ends of Λ-precursors initiated filament bundling and completed formation of the nascent filopodium. We propose a convergent elongation model of filopodia initiation, stipulating that filaments within the lamellipodial dendritic network acquire privileged status by binding a set of molecules (including VASP) to their barbed ends, which protect them from capping and mediate association of barbed ends with each other

Response to : The GPRC5A frameshift variant c.183del is not associated with increased breast cancer risk in BRCA1 mutation carriers

Non peer reviewe

DIFFERENTIAL DIAGNOSIS OF RADICULOPATHY IN LYME BORRELIOSISAND DYSTROPHIC LESIONS OF THE VERTEBRAL COLUMN

Subjects and methods. Thirty patients, including 9 (30.0%) men and 21 (70.0%) women, with signs of radiculopathy (RP) in late-stages Lyme borreliosis (LB) were examined. A control group comprised 30 patients with vertebrogenic RP in the presence of dystrophic changes in the vertebral column. Results. 56.7% of the patients with LB were observed to have its primary chronic course in the absence of the acute period of LB. The latter with the signs of RP showed a topic association between the pain location and the tick bite site in 43.3% of the patients. A gradual disease development was more frequently (63.3%) observed in LB while the periods of remission and exacerbation were more typical (56.7%) in vertebrogenic RP. In the patients with LB, pain syndrome depended on posture and physical exercise less frequently (30.0%) than in those with vertebrogenic RP (96.7%). Bilateral pain irradiation was more characteristic of RP in LB than in dystrophic lesions of the vertebral column. The symptoms of tonic muscle tension and limited movement volume in the afflicted part of the vertebral column were significantly less common in the patients with LB than in those with vertebrogenic RP. LB was marked by a concomitance of radicular and polyneuritic disorders (83.3%) and vertebrogenic RP was characterized by a preponderance of the radicular-type of sensitivity disorder (100%). Systemic inflammatory syndrome and polysystemacy of manifestations were more characteristic of LB. The benefits of nonsteriodal antiinflammatory drugs in LB patients with RP were significantly worse than in those with vertebrogenic RP; regression of symptoms in LB was seen only after a course of specific antibiotic therapy

Orphan G protein-coupled receptor GPRC5A modulates integrin β1-mediated epithelial cell adhesion

G-Protein Coupled Receptor (GPCR), Class C, Group 5, Member A (GPRC5A) has been implicated in several malignancies. The underlying mechanisms, however, remain poorly understood. Using a panel of human cell lines, we demonstrate that CRISPR/Cas9-mediated knockout and RNAi-mediated depletion of GPRC5A impairs cell adhesion to integrin substrates: collagens I and IV, fibronectin, as well as to extracellular matrix proteins derived from the Engelbreth-Holm-Swarm (EHS) mouse sarcoma (Matrigel). Consistent with the phenotype, knock-out of GPRC5A correlated with a reduced integrin β1 (ITGB1) protein expression, impaired phosphorylation of the focal adhesion kinase (FAK), and lower activity of small GTPases RhoA and Rac1. Furthermore, we provide the first evidence for a direct interaction between GPRC5A and a receptor tyrosine kinase EphA2, an upstream regulator of FAK, although its contribution to the observed adhesion phenotype is unclear. Our findings reveal an unprecedented role for GPRC5A in regulation of the ITGB1-mediated cell adhesion and it's downstream signaling, thus indicating a potential novel role for GPRC5A in human epithelial cancers.</p

Primary carbonatite melt from deeply subducted oceanic crust

Partial melting in the Earth's mantle plays an important part in generating the geochemical and isotopic diversity observed in volcanic rocks at the surface. Identifying the composition of these primary melts in the mantle is crucial for establishing links between mantle geochemical 'reservoirs' and fundamental geodynamic processes. Mineral inclusions in natural diamonds have provided a unique window into such deep mantle processes. Here we provide experimental and geochemical evidence that silicate mineral inclusions in diamonds from Juina, Brazil, crystallized from primary and evolved carbonatite melts in the mantle transition zone and deep upper mantle. The incompatible trace element abundances calculated for a melt coexisting with a calcium-titanium-silicate perovskite inclusion indicate deep melting of carbonated oceanic crust, probably at transition-zone depths. Further to perovskite, calcic-majorite garnet inclusions record crystallization in the deep upper mantle from an evolved melt that closely resembles estimates of primitive carbonatite on the basis of volcanic rocks. Small-degree melts of subducted crust can be viewed as agents of chemical mass-transfer in the upper mantle and transition zone, leaving a chemical imprint of ocean crust that can possibly endure for billions of years.4 page(s

The Second Transmembrane Domain of P2X7 Contributes to Dilated Pore Formation

Activation of the purinergic receptor P2X7 leads to the cellular permeability of low molecular weight cations. To determine which domains of P2X7 are necessary for this permeability, we exchanged either the C-terminus or portions of the second transmembrane domain (TM2) with those in P2X1 or P2X4. Replacement of the C-terminus of P2X7 with either P2X1 or P2X4 prevented surface expression of the chimeric receptor. Similarly, chimeric P2X7 containing TM2 from P2X1 or P2X4 had reduced surface expression and no permeability to cationic dyes. Exchanging the N-terminal 10 residues or C-terminal 14 residues of the P2X7 TM2 with the corresponding region of P2X1 TM2 partially restored surface expression and limited pore permeability. To further probe TM2 structure, we replaced single residues in P2X7 TM2 with those in P2X1 or P2X4. We identified multiple substitutions that drastically changed pore permeability without altering surface expression. Three substitutions (Q332P, Y336T, and Y343L) individually reduced pore formation as indicated by decreased dye uptake and also reduced membrane blebbing in response to ATP exposure. Three others substitutions, V335T, S342G, and S342A each enhanced dye uptake, membrane blebbing and cell death. Our results demonstrate a critical role for the TM2 domain of P2X7 in receptor function, and provide a structural basis for differences between purinergic receptors. © 2013 Sun et al

Neuroinflammation, Mast Cells, and Glia: Dangerous Liaisons

The perspective of neuroinflammation as an epiphenomenon following neuron damage is being replaced by the awareness of glia and their importance in neural functions and disorders. Systemic inflammation generates signals that communicate with the brain and leads to changes in metabolism and behavior, with microglia assuming a pro-inflammatory phenotype. Identification of potential peripheral-to-central cellular links is thus a critical step in designing effective therapeutics. Mast cells may fulfill such a role. These resident immune cells are found close to and within peripheral nerves and in brain parenchyma/meninges, where they exercise a key role in orchestrating the inflammatory process from initiation through chronic activation. Mast cells and glia engage in crosstalk that contributes to accelerate disease progression; such interactions become exaggerated with aging and increased cell sensitivity to stress. Emerging evidence for oligodendrocytes, independent of myelin and support of axonal integrity, points to their having strong immune functions, innate immune receptor expression, and production/response to chemokines and cytokines that modulate immune responses in the central nervous system while engaging in crosstalk with microglia and astrocytes. In this review, we summarize the findings related to our understanding of the biology and cellular signaling mechanisms of neuroinflammation, with emphasis on mast cell-glia interactions