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Recently revised brain tumor classification suggested a glioma treatment strategy that takes into consideration molecular variants in IDH1 and TP53 marker genes. While pathogenic variants of IDH1 and TP53 can be accompanied by chromosomal instability (CIN), the impact of IDH1 and TP53 mutations on genome stability remains unstudied. Elevated CIN might provide therapeutic targets, based on synergistic effects of chemotherapy with CIN-inducing drugs. Using an assay based on human artificial chromosomes, we investigated the impact of common glioma missense mutations in IDH1 and TP53 on chromosome transmission and demonstrated that IDH1R132H and TP53R248Q variants elevate CIN. We next found enhanced CIN levels and the sensitivity of IDH1 R132H/WT and TP53 R248Q/R248Q genotypes, introduced into U87 MG glioma cells by CRISPR/Cas9, to different drugs, including conventional temozolomide. It was found that U87 MG cells carrying IDH1 R132H/WT exhibit dramatic sensitivity to paclitaxel, which was independently confirmed on cell cultures derived from patients with naturally occurring IDH1 R132H/WT . Overall, our results suggest that the development of CIN-enhancing therapy for glioma tumors with the IDH1 R132H/WT genotype could be advantageous for adjuvant treatment., Competing Interests: None declared., (© The Author(s) 2025. Published by Oxford University Press on behalf of NAR Cancer.)

Albumin is a major protein in mammalian blood plasma or serum, where its concentration in healthy organisms is about 600 μM [...].

Data are accumulating on the hydrolytic activity of serum albumin towards esters and organophosphates. Previously, with the help of the technology of proton nuclear magnetic resonance ( 1 H NMR) spectroscopy, we observed the yield of acetate in the solution of bovine serum albumin and p -nitrophenyl acetate (NPA). Thus, we showed that albumin possesses true esterase activity towards NPA. Then, using the methods of molecular docking and molecular dynamics, we established site Sudlow I as the catalytic center of true esterase activity of albumin. In the present work, to expand our understanding of the molecular mechanisms of albumin pseudoesterase and true esterase activity, we investigated-in experiments in vitro and in silico-the interaction of anticoagulant warfarin (WRF, specific ligand of site Sudlow I) and benzodiazepine diazepam (DIA, specific ligand of site Sudlow II) with albumins of different species, and determined how the binding of WRF and DIA affects the hydrolysis of NPA by albumin. It was found that the characteristics of the binding modes of WRF in site Sudlow I and DIA in site Sudlow II of human (HSA), bovine (BSA), and rat (RSA) albumins have species differences, which are more pronounced for site Sudlow I compared to site Sudlow II, and less pronounced between HSA and RSA compared to BSA. WRF competitively inhibits true esterase activity of site Sudlow I towards NPA and does not affect the functioning of site Sudlow II. Diazepam can slow down true esterase activity of site Sudlow I in noncompetitive manner. It was concluded that site Sudlow I is more receptive to allosteric modulation compared to site Sudlow II.

The study of drugs of natural origin that increase endurance and/or accelerate recovery is an integral part of sports medicine and physiology. In this paper, decaffeinated green tea extract (GTE) and two ammonium salts-chloride (ACL) and carbonate (ACR)-were tested individually and in combination with GTE as stimulants of physical performance in a forced swimming rat experimental model. The determined parameters can be divided into seven blocks: functional (swimming duration); biochemistry of blood plasma; biochemistry of erythrocytes; hematology; immunology; gene expression of slow- and fast-twitch muscles (m. soleus, SOL, and m. extensor digitorum longus, EDL, respectively); and morphometric indicators of slow- and fast-twitch muscles. Regarding the negative control (intact animals), the maximum number of changes in all blocks of indicators was recorded in the GTE + ACR group, whose animals showed the maximum functional result and minimum lactate values on the last day of the experiment. Next, in terms of the number of changes, were the groups ACR, ACL, GTE + ACL, GTE and NaCl (positive control). In general, the number of identified adaptive changes was proportional to the functional state of the animals of the corresponding groups, in terms of the duration of the swimming load in the last four days of the experiment. However, not only the total number but also the qualitative composition of the identified changes is of interest. The results of a comparative analysis suggest that, in the model of forced swimming we developed, GTE promotes restoration of the body and moderate mobilization of the immune system, while small doses of ammonium salts, especially ammonium carbonate, contribute to an increase in physical performance, which is associated with satisfactory restoration of skeletal muscles and the entire body. The combined use of GTE with ammonium salts does not give a clearly positive effect.

Human serum albumin (HSA) is an endogenous inhibitor of angiotensin I-converting enzyme (ACE) and, thus, plays a key role in the renin-angiotensin-aldosterone system (RAAS). However, little is known about the mechanism of interaction between these proteins, and the structure of the HSA-ACE complex has not yet been obtained experimentally. The purpose of the presented work is to apply computer modeling methods to study the interaction of HSA with ACE in order to obtain preliminary details about the mechanism of their interaction. Ten possible HSA-ACE complexes were obtained by the procedure of macromolecular docking. Based on the number of steric and polar contacts between the proteins, three leading complexes were selected, the stabilities of which were then tested by molecular dynamics (MD) simulation. Based on the results of MD simulation, the two most probable conformations of the HSA-ACE complex were selected. The analysis of these conformations revealed that the processes of oxidation of the thiol group of Cys34 of HSA and the binding of albumin to ACE can reciprocally affect each other. Known point mutations in the albumin molecules Glu82Lys, Arg114Gly, Glu505Lys, Glu565Lys and Lys573Glu can also affect the interaction with ACE. According to the result of MD simulation, the known ACE mutations, albeit associated with various diseases, do not affect the HSA-ACE interaction. A comparative analysis was performed of the resulting HSA-ACE complexes with those obtained by AlphaFold 3 as well as with the crystal structure of the HSA and the neonatal Fc receptor (FcRn) complex. It was found that domains DI and DIII of albumin are involved in binding both ACE and FcRn. The obtained results of molecular modeling outline the direction for further study of the mechanisms of HSA-ACE interaction in vitro. Information about these mechanisms will help in the design and improvement of pharmacotherapy aimed at modulation of the physiological activity of ACE.

In mammals, glycated serum albumin (gSA) contributes to the pathogenesis of many metabolic diseases by activating the receptors (RAGE) for advanced glycation end products (AGEs). Many aspects of the gSA-RAGE interaction remain unknown. The purpose of the present paper was to study the interaction of glycated human albumin (gHSA) with RAGE using molecular modeling methods. Ten models of gHSA modified with different lysine residues to carboxymethyl-lysines were prepared. Complexes of gHSA-RAGE were obtained by the macromolecular docking method with subsequent molecular dynamics simulation (MD). According to the MD, the RAGE complexes with gHSA glycated at Lys233, Lys64, Lys525, Lys262 and Lys378 are the strongest. Three-dimensional models of the RAGE dimers with gHSA were proposed. Additional computational experiments showed that the binding of fatty acids (FAs) to HSA does not affect the ability of Lys525 (the most reactive lysine) to be glycated. In contrast, modification of Lys525 reduces the affinity of albumin for FA. The interspecies differences in the molecular structure of albumin that may affect the mechanism of the gSA-RAGE interaction were discussed. The obtained results will help us to learn more about the molecular basis for the involvement of serum albumin in the AGE/RAGE axis and improve the methodology for studying cellular signaling pathways involving RAGE.

The process of aging is accompanied by a dynamic restructuring of the immune response, a phenomenon known as immunosenescence. Further, damage to the endothelium can be both a cause and a consequence of many diseases, especially in elderly people. The purpose of this study was to carry out immunological and biochemical profiling of elderly people with acute ischemic stroke (AIS), chronic cerebral circulation insufficiency (CCCI), prediabetes or newly diagnosed type II diabetes mellitus (DM), and subcortical ischemic vascular dementia (SIVD). Socio-demographic, lifestyle, and cognitive data were obtained. Biochemical, hematological, and immunological analyses were carried out, and extracellular vesicles (EVs) with endothelial CD markers were assessed. The greatest number of significant deviations from conditionally healthy donors (HDs) of the same age were registered in the SIVD group, a total of 20, of which 12 were specific and six were non-specific but with maximal differences (as compared to the other three groups) from the HDs group. The non-specific deviations were for the MOCA (Montreal Cognitive Impairment Scale), the MMSE (Mini Mental State Examination) and life satisfaction self-assessment scores, a decrease of albumin levels, and ADAMTS13 (a Disintegrin and Metalloproteinase with a Thrombospondin Type 1 motif, member 13) activity, and an increase of the VWF (von Willebrand factor) level. Considering the significant changes in immunological parameters (mostly Th17-like cells) and endothelial CD markers (CD144 and CD34), vascular repair was impaired to the greatest extent in the DM group. The AIS patients showed 12 significant deviations from the HD controls, including three specific to this group. These were high NEFAs (non-esterified fatty acids) and CD31 and CD147 markers of EVs. The lowest number of deviations were registered in the CCCI group, nine in total. There were significant changes from the HD controls with no specifics to this group, and just one non-specific with a maximal difference from the control parameters, which was α1-AGP (alpha 1 acid glycoprotein, orosomucoid). Besides the DM patients, impairments of vascular repair were also registered in the CCCI and AIS patients, with a complete absence of such in patients with dementia (SIVD group). On the other hand, microvascular damage seemed to be maximal in the latter group, considering the biochemical indicators VWF and ADAMTS13. In the DM patients, a maximum immune response was registered, mainly with Th17-like cells. In the CCCI group, the reaction was not as pronounced compared to other groups of patients, which may indicate the initial stages and/or compensatory nature of organic changes (remodeling). At the same time, immunological and biochemical deviations in SIVD patients indicated a persistent remodeling in microvessels, chronic inflammation, and a significant decrease in the anabolic function of the liver and other tissues. The data obtained support two interrelated assumptions. Taking into account the primary biochemical factors that trigger the pathological processes associated with vascular pathology and related diseases, the first assumption is that purine degradation in skeletal muscle may be a major factor in the production of uric acid, followed by its production by non-muscle cells, the main of which are endothelial cells. Another assumption is that therapeutic factors that increase the levels of endothelial progenitor cells may have a therapeutic effect in reducing the risk of cerebrovascular disease and related neurodegenerative diseases.

During the initial diagnosis of urgent medical conditions, which include acute infectious diseases, it is important to assess the severity of the patient's clinical state as quickly as possible. Unlike individual biochemical or physiological indicators, derived indices make it possible to better characterize a complex syndrome as a set of symptoms, and therefore quickly take a set of adequate measures. Recently, we reported on novel diagnostic indices containing butyrylcholinesterase (BChE) activity, which is decreased in COVID-19 patients. Also, in these patients, the secretion of von Willebrand factor (vWF) increases, which leads to thrombosis in the microvascular bed. The objective of this study was the determination of the concentration and activity of vWF in patients with COVID-19, and the search for new diagnostic indices. One of the main objectives was to compare the prognostic values of some individual and newly derived indices. Patients with COVID-19 were retrospectively divided into two groups: survivors (n = 77) and deceased (n = 24). According to clinical symptoms and computed tomography (CT) results, the course of disease was predominantly moderate in severity. The first blood sample (first point) was taken upon admission to the hospital, the second sample (second point)-within 4-6 days after admission. Along with the standard spectrum of biochemical indicators, BChE activity (BChEa or BChEb for acetylthiocholin or butyrylthiocholin, respectively), malondialdehyde (MDA), and vWF analysis (its antigen level, AGFW, and its activity, ActWF) were determined and new diagnostic indices were derived. The pooled sensitivity, specificity, and area under the receiver operating curve (AUC), as well as Likelihood ratio (LR) and Odds ratio (OR) were calculated. The level of vWF antigen in the deceased group was 1.5-fold higher than the level in the group of survivors. Indices that include vWF antigen levels are superior to indices using vWF activity. It was found that the index [Urea] × [AGWF] × 1000/(BChEb × [ALB]) had the best discriminatory power to predict COVID-19 mortality (AUC = 0.91 [0.83, 1.00], p < 0.0001; OR = 72.0 [7.5, 689], p = 0.0002). In addition, [Urea] × 1000/(BChEb × [ALB]) was a good predictor of mortality (AUC = 0.95 [0.89, 1.00], p < 0.0001; OR = 31.5 [3.4, 293], p = 0.0024). The index [Urea] × [AGWF] × 1000/(BChEb × [ALB]) was the best predictor of mortality associated with COVID-19 infection, followed by [Urea] × 1000/(BChEb × [ALB]). After validation in a subsequent cohort, these two indices could be recommended for diagnostic laboratories.

Glioma is one of the most intractable types of cancer, due to delayed diagnosis at advanced stages. The clinical symptoms of glioma are unclear and due to a variety of glioma subtypes, available low-invasive testing is not effective enough to be introduced into routine medical laboratory practice. Therefore, recent advances in the clinical diagnosis of glioma have focused on liquid biopsy approaches that utilize a wide range of techniques such as next-generation sequencing (NGS), droplet-digital polymerase chain reaction (ddPCR), and quantitative PCR (qPCR). Among all techniques, NGS is the most advantageous diagnostic method. Despite the rapid cheapening of NGS experiments, the cost of such diagnostics remains high. Moreover, high-throughput diagnostics are not appropriate for molecular profiling of gliomas since patients with gliomas exhibit only a few diagnostic markers. In this review, we highlighted all available assays for glioma diagnosing for main pathogenic glioma DNA sequence alterations. In the present study, we reviewed the possibility of integrating routine molecular methods into the diagnosis of gliomas. We state that the development of an affordable assay covering all glioma genetic aberrations could enable early detection and improve patient outcomes. Moreover, the development of such molecular diagnostic kits could potentially be a good alternative to expensive NGS-based approaches., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Penkova, Kuziakova, Gulaia, Tiasto, Goncharov, Lanskikh, Zhmenia, Baklanov, Farniev and Kumeiko.)

The most favorable targets for retrospectively determining human exposure to organophosphorus pesticides, insecticides, retardants, and other industrial organophosphates (OPs) are adducts of OPs with blood plasma butyrylcholinesterase (BChE) and human serum albumin (HSA). One of the methods for determining OP exposure is the reactivation of modified BChE using a concentrated solution of KF in an acidic medium. It is known that under the action of fluoride ion, OPs or their fluoroanhydrides can be released not only from BChE adducts but also from the adducts with albumin; however, the contribution of albumin to the total pool of released OPs after plasma treatment with KF has not yet been studied. The efficiency of OP release can be affected by many factors associated with the experimental technique, but first, the structure of the adduct must be taken into account. We report a comparative analysis of the structure and conformation of organophosphorus adducts on HSA and BChE using molecular modeling methods and the mechanism of OP release after fluoride ion exposure. The conformational analysis of the organophosphorus adducts on HSA and BChE was performed, and the interaction of fluoride ions with modified proteins was studied by molecular dynamics simulation. The geometric and energy characteristics of the studied adducts and their complexes with fluoride ion were calculated using molecular mechanics and semiempirical approaches. The structural features of modified HSA and BChE that can affect the efficiency of OP release after fluoride ion exposure were revealed. Using the proposed approach, the expediency of using KF for establishing exposure to different OPs, depending on their structure, can be assessed.

The aim of this work was to explore the involvement of 5-HT 1B and 5-HT 2B receptors (5-HT 1B R and 5-HT 2B R) in the regulation of free cytoplasmic calcium concentration ([Ca 2+ ] i ) in human umbilical vein endothelial cells (HUVEC). We have shown by quantitative PCR analysis, that 5-HT 1B R and 5-HT 2B R mRNAs levels are almost equal in HUVEC. Immunofluorescent staining demonstrated, that 5-HT 1B R and 5-HT 2B R are expressed both in plasma membrane and inside the cells. Intracellular 5-HT 1B R are localized mainly in the nuclear region, whereas 5-HT 2B R receptors are almost evenly distributed in HUVEC. 5-HT, 5-HT 1B R agonist CGS12066B, or 5-HT 2B R agonist BW723C86 added to HUVEC caused a slight increase in [Ca 2+ ] i , which was much lower than that of histamine, ATP, or SFLLRN, an agonist of protease-activated receptors (PAR1). However, activation of 5-HT 1B R with CGS12066B followed by activation of 5-HT 2B R with BW723C86 manifested a synergism of response, since several-fold higher rise in [Ca 2+ ] i occurred. CGS12066B caused more than a 5-fold increase in [Ca 2+ ] i rise in HUVEC in response to 5-HT. This 5-HT induced [Ca 2+ ] i rise was abolished by 5-HT 2B R antagonist RS127445, indicating that extracellular 5-HT acts through 5-HT 2B R. Synergistic [Ca 2+ ] i rise in response to activation of 5-HT1BR and 5-HT 2B R persisted in a calcium-free medium. It was suppressed by the phospholipase C inhibitor U73122 and was not inhibited by the ryanodine and NAADP receptors antagonists dantrolene and NED-19. [Ca 2+ ] i measurements in single cells demonstrated that activation of 5-HT 2B R alone by BW723C86 caused single asynchronous [Ca 2+ ] i oscillations in 19.8 ± 4.2% ( n = 3) of HUVEC that occur with a long delay (66.1 ± 4.3 s, n = 71). On the contrary, histamine causes a simultaneous and almost immediate increase in [Ca 2+ ] i in all the cells. Pre-activation of 5-HT 1B R by CGS12066B led to a 3-4 fold increase in the number of HUVEC responding to BW723C86, to synchronization of their responses with a delay shortening, and to the bursts of [Ca 2+ ] i oscillations in addition to single oscillations. In conclusion, to get a full rise of [Ca 2+ ] i in HUVEC in response to 5-HT, simultaneous activation of 5-HT 1B R and 5-HT 2B R is required. 5-HT causes an increase in [Ca 2+ ] i via 5-HT 2B R while 5-HT 1B R could be activated by the membrane-permeable agonist CGS12066B. We hypothesized that CGS12066B acts via intracellular 5-HT 1B R inaccessible to extracellular 5-HT. Intracellular 5-HT 1B R might be activated by 5-HT which could be accumulated in EC under certain pathological conditions.

The etiology and pathogenesis of Alzheimer's disease are multifactorial, so one of the treatment strategies is the development of the drugs that affect several targets associated with the pathogenesis of the disease. Within this roadmap, we investigated the interaction of several substituted 1,3-dihydro-2-oxo-1 H -benzimidazol-2-ones with their potential molecular targets: cholinesterases (ChE) and three types of the G s -protein-coupled serotonin receptors (5-HTR) 5-HT 6 , 5-HT 4 and 5-HT 7 (5-HT 4 R, 5-HT 6 R and 5-HT 7 R, respectively). A microplate modification of the Ellman method was used for the biochemical analysis of the inhibitory ability of the drugs towards ChE. Molecular modeling methods, such as molecular docking and molecular dynamics (MD) simulation in water and the lipid bilayer, were used to study the interaction of the compounds with ChE and 5-HTR. In vitro experiments showed that the tested compounds had moderate anticholinesterase activity. With the help of molecular modeling methods, the mechanism of interaction of the tested compounds with ChE was investigated, the binding sites were described and the structural features of the drugs that determine the strength of their anticholinesterase activity were revealed. Primary in silico evaluation showed that benzimidazole-carboxamides effectively bind to 5-HT 4 R and 5-HT 7 R. The pool of the obtained data allows us to choose N-[2-(diethylamino)ethyl]-2-oxo-3-(tert-butyl)-2,3-dihydro-1 H -benzimidazole-1-carboxamide hydrochloride (compound 13 ) as the most promising for further experimental development.

A primary in vitro analysis of the anticholinesterase properties of substituted 1,3-dihydro-2-oxo-1H-benzimidazol-2-ones was performed along with in silico calculation of their oral toxicity. These compounds are analogs of BIMU-8, a well-known agonist of serotonin 5-HT 4 receptors, and are supposed to combine the functions of cholinesterase inhibitors and serotonin receptor agonists. Biochemical analysis showed the ability of the obtained chemicals to inhibit acetyl- and butyrylcholinesterase. A compound with minimal toxicity, high inhibitory ability against butyrylcholinesterase, and low inhibitory ability against acetylcholinesterase has been identified, which is of greatest interest for further experimental development., (© 2023. Springer Science+Business Media, LLC, part of Springer Nature.)

The esterase status of blood plasma can claim to be one of the universal markers of various diseases; therefore, it deserves attention when searching for markers of the severity of COVID-19 and other infectious and non-infectious pathologies. When analyzing the esterase status of blood plasma, the esterase activity of serum albumin, which is the major protein in the blood of mammals, should not be ignored. The purpose of this study is to expand understanding of the esterase status of blood plasma and to evaluate the relationship of the esterase status, which includes information on the amount and enzymatic activity of human serum albumin (HSA), with other biochemical parameters of human blood, using the example of surviving and deceased patients with confirmed COVID-19. In experiments in vitro and in silico, the activity of human plasma and pure HSA towards various substrates was studied, and the effect of various inhibitors on this activity was tested. Then, a comparative analysis of the esterase status and a number of basic biochemical parameters of the blood plasma of healthy subjects and patients with confirmed COVID-19 was performed. Statistically significant differences have been found in esterase status and biochemical indices (including albumin levels) between healthy subjects and patients with COVID-19, as well as between surviving and deceased patients. Additional evidence has been obtained for the importance of albumin as a diagnostic marker. Of particular interest is a new index, [Urea] × [MDA] × 1000/(BChEb × [ALB]), which in the group of deceased patients was 10 times higher than in the group of survivors and 26 times higher than the value in the group of apparently healthy elderly subjects.

Human artificial chromosomes (HACs) have provided a useful tool to study kinetochore structure and function, gene delivery, and gene expression. The HAC propagates and segregates properly in the cells. Recently, we have developed an experimental high-throughput imaging (HTI) HAC-based assay that allows the identification of genes whose depletion leads to chromosome instability (CIN). The HAC carries a GFP transgene that facilitates quantitative measurement of CIN. The loss of HAC/GFP may be measured by flow cytometry or fluorescence scanning microscope. Therefore, CIN rate can be measured by counting the proportion of fluorescent cells. Here, the HAC/GFP-based assay has been adapted to screen anticancer compounds for possible induction or elevation of CIN. We analyzed 24 cytotoxic plant extracts. Punica granatum leaf extract (PLE) indeed sharply increases CIN rate in HT1080 fibrosarcoma cells. PLE treatment leads to cell cycle arrest, reduction of mitotic index, and the increased numbers of micronuclei (MNi) and nucleoplasmic bridges (NPBs). PLE-mediated increased CIN correlates with the induction of double-stranded breaks (DSBs). We infer that the PLE extract contains a component(s) that elevate CIN, making it a candidate for further study as a potential cancer treatment. The data also provide a proof of principle for the utility of the HAC/GFP-based system in screening for natural products and other compounds that elevate CIN in cancer cells., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Goncharov, Kovalskaia, Romanishin, Shved, Belousov, Tiasto, Gulaia, Neergheen, Rummun, Liskovykh, Larionov, Kouprina and Kumeiko.)

The carrageenans isolated from red algae demonstrated a variety of activities from antiviral and immunomodulatory to antitumor. The diverse structure and sulfation profile of carrageenans provide a great landscape for drug development. In this study, we isolated, purified and structurally characterized κo- and λo- oligosaccharides from the marine algae Chondrus armatus . We further examined the tumor suppressive activity of both carrageenans in gastrointestinal cancer models. Thus, using MTT assay, we could demonstrate a pronounced antiproliferative effect of the carrageenans in KYSE-30 and FLO-1 as well as HCT-116 and RKO cell lines with IC 50 184~405 μg/mL, while both compounds were less active in non-cancer epithelial cells RPE-1. This effect was stipulated by the inhibition of cell cycle progression in the cancer cells. Specifically, flow cytometry revealed an S phase delay in FLO-1 and HCT-116 cells under κo-carrageenan treatment, while KYSE-30 demonstrated a pronounced G 2 /M cell cycle delay. In line with this, western blotting revealed a reduction of cell cycle markers CDK2 and E2F2. Interestingly, κo-carrageenan inhibited cell cycle progression of RKO cells in G 1 phase. Finally, isolated κo- and λo- carrageenans induced apoptosis on adenocarcinomas, specifically with high apoptosis induction in RKO cells. Overall, our data underline the potential of κo- and λo- carrageenans for colon and esophageal carcinoma drug development.

An increasing number of the population all around the world suffer from age-associated neurodegenerative diseases including Parkinson's disease (PD). This disorder presents different signs of genetic, epigenetic and environmental origin, and molecular, cellular and intracellular dysfunction. At the molecular level, α-synuclein (αSyn) was identified as the principal molecule constituting the Lewy bodies (LB). The gut microbiota participates in the pathogenesis of PD and may contribute to the loss of dopaminergic neurons through mitochondrial dysfunction. The most important pathogenetic link is an imbalance of Ca 2+ ions, which is associated with redox imbalance in the cells and increased generation of reactive oxygen species (ROS). In this review, genetic, epigenetic and environmental factors that cause these disorders and their cause-and-effect relationships are considered. As a constituent of environmental factors, the example of organophosphates (OPs) is also reviewed. The role of endothelial damage in the pathogenesis of PD is discussed, and a 'triple hit hypothesis' is proposed as a modification of Braak's dual hit one. In the absence of effective therapies for neurodegenerative diseases, more and more evidence is emerging about the positive impact of nutritional structure and healthy lifestyle on the state of blood vessels and the risk of developing these diseases.

Organophosphates (OPs) are toxic chemicals produced by an esterification process and some other routes. They are the main components of herbicides, pesticides, and insecticides and are also widely used in the production of plastics and solvents. Acute or chronic exposure to OPs can manifest in various levels of toxicity to humans, animals, plants, and insects. OPs containing insecticides were widely used in many countries during the 20th century, and some of them continue to be used today. In particular, 36 OPs have been registered in the USA, and all of them have the potential to cause acute and sub-acute toxicity. Renal damage and impairment of kidney function after exposure to OPs, accompanied by the development of clinical manifestations of poisoning back in the early 1990s of the last century, was considered a rare manifestation of their toxicity. However, since the beginning of the 21st century, nephrotoxicity of OPs as a manifestation of delayed toxicity is the subject of greater attention of researchers. In this article, we present a modern view on the molecular pathophysiological mechanisms of acute nephrotoxicity of organophosphate compounds.

The development of drugs for the therapy of COVID-19 is one of the main problems of modern physiology, biochemistry and pharmacology. Taking into account the available information on the participation of mast cells and the role of histamine in the pathogenesis of COVID-19, as well as information on the positive role of famotidine in the prevention and treatment of coronavirus infection, an experiment was carried out using famotidine in a mouse model. We used a type A/PR/8/34 (H1N1) virus adapted to mice. The antiviral drug oseltamivir (Tamiflu), which belongs to the group of neuraminidase inhibitors, was used as a reference drug. The use of famotidine in combination with oseltamivir can increase survival, improve the dynamics of animal weight, reduce the level of NKT cells and increase the level of naive T-helpers. Further studies of famotidine in vivo should be aimed at optimizing the regimen of drug use at a higher viral load, as well as with a longer use of famotidine., Competing Interests: CONFLICTS OF INTERESTThe authors declare no obvious and potential conflicts of interest related to the publication of this article., (© Pleiades Publishing, Ltd. 2022.)

The delayed effects of acute intoxication by organophosphates (OPs) are poorly understood, and the various experimental animal models often do not take into account species characteristics. The principal biochemical feature of rodents is the presence of carboxylesterase in blood plasma, which is a target for OPs and can greatly distort their specific effects. The present study was designed to investigate the nephrotoxic effects of paraoxon (O,O-diethyl O-(4-nitrophenyl) phosphate, POX) using three models of acute poisoning in outbred Wistar rats. In the first model ( M1 , POX2x group), POX was administered twice at doses 110 µg/kg and 130 µg/kg subcutaneously, with an interval of 1 h. In the second model ( M2 , CBPOX group), 1 h prior to POX poisoning at a dose of 130 µg/kg subcutaneously, carboxylesterase activity was pre-inhibited by administration of specific inhibitor cresylbenzodioxaphosphorin oxide (CBDP, 3.3 mg/kg intraperitoneally). In the third model ( M3 ), POX was administered subcutaneously just once at doses of LD16 (241 µg/kg), LD50 (250 µg/kg), and LD84 (259 µg/kg). Animal observation and sampling were performed 1, 3, and 7 days after the exposure. Endogenous creatinine clearance (ECC) decreased in 24 h in the POX2x group ( p = 0.011). Glucosuria was observed in rats 24 h after exposure to POX in both M1 and M2 models. After 3 days, an increase in urinary excretion of chondroitin sulfate (CS, p = 0.024) and calbindin ( p = 0.006) was observed in rats of the CBPOX group. Morphometric analysis revealed a number of differences most significant for rats in the CBPOX group. Furthermore, there was an increase in the area of the renal corpuscles ( p = 0.0006), an increase in the diameter of the lumen of the proximal convoluted tubules (PCT, p = 0.0006), and narrowing of the diameter of the distal tubules ( p = 0.001). After 7 days, the diameter of the PCT lumen was still increased in the nephrons of the CBPOX group ( p = 0.0009). In the M3 model, histopathological and ultrastructural changes in the kidneys were revealed after the exposure to POX at doses of LD50 and LD84. Over a period from 24 h to 3 days, a significant ( p = 0.018) expansion of Bowman's capsule was observed in the kidneys of rats of both the LD50 and LD84 groups. In the epithelium of the proximal tubules, stretching of the basal labyrinth, pycnotic nuclei, and desquamation of microvilli on the apical surface were revealed. In the epithelium of the distal tubules, partial swelling and destruction of mitochondria and pycnotic nuclei was observed, and nuclei were displaced towards the apical surface of cells. After 7 days of the exposure to POX, an increase in the thickness of the glomerular basement membrane (GBM) was observed in the LD50 and LD84 groups ( p = 0.019 and 0.026, respectively). Moreover, signs of damage to tubular epithelial cells persisted with blockage of the tubule lumen by cellular detritus and local destruction of the surface of apical cells. Comparison of results from the three models demonstrates that the nephrotoxic effects of POX, evaluated at 1 and 3 days, appear regardless of prior inhibition of carboxylesterase activity.

C1q domain-containing (C1qDC) proteins are a group of biopolymers involved in immune response as pattern recognition receptors (PRRs) in a lectin-like manner. A new protein MkC1qDC from the hemolymph plasma of Modiolus kurilensis bivalve mollusk widespread in the Northwest Pacific was purified. The isolation procedure included ammonium sulfate precipitation followed by affinity chromatography on pectin-Sepharose. The full-length MkC1qDC sequence was assembled using de novo mass-spectrometry peptide sequencing complemented with N-terminal Edman's degradation, and included 176 amino acid residues with molecular mass of 19 kDa displaying high homology to bivalve C1qDC proteins. MkC1qDC demonstrated antibacterial properties against Gram-negative and Gram-positive strains. MkC1qDC binds to a number of saccharides in Ca 2+ -dependent manner which characterized by structural meta-similarity in acidic group enrichment of galactose and mannose derivatives incorporated in diversified molecular species of glycans. Alginate, κ-carrageenan, fucoidan, and pectin were found to be highly effective inhibitors of MkC1qDC activity. Yeast mannan, lipopolysaccharide (LPS), peptidoglycan (PGN) and mucin showed an inhibitory effect at concentrations three orders of magnitude greater than for the most effective saccharides. MkC1qDC localized to the mussel hemal system and interstitial compartment. Intriguingly, MkC1qDC was found to suppress proliferation of human adenocarcinoma HeLa cells in a dose-dependent manner, indicating to the biomedical potential of MkC1qDC protein.

Serum albumin possesses esterase and pseudo-esterase activities towards a number of endogenous and exogenous substrates, but the mechanism of interaction of various esters and other compounds with albumin is still unclear. In the present study, proton nuclear magnetic resonance ( 1 H NMR) has been applied to the study of true esterase activity of albumin, using the example of bovine serum albumin (BSA) and p -nitrophenyl acetate (NPA). The site of BSA esterase activity was then determined using molecular modelling methods. According to the data obtained, the accumulation of acetate in the presence of BSA in the reaction mixture is much more intense as compared with the spontaneous hydrolysis of NPA, which indicates true esterase activity of albumin towards NPA. Similar results were obtained for p -nitophenyl propionate (NPP) as substrate. The rate of acetate and propionate release confirms the assumption that there is a site of true esterase activity in the albumin molecule, which is different from the site of the pseudo-esterase activity Sudlow II. The results of molecular modelling of BSA and NPA interaction make it possible to postulate that Sudlow site I is the site of true esterase activity of albumin.

Being one of the main proteins in the human body and many animal species, albumin plays a decisive role in the transport of various ions-electrically neutral and charged molecules-and in maintaining the colloidal osmotic pressure of the blood. Albumin is able to bind to almost all known drugs, as well as many nutraceuticals and toxic substances, largely determining their pharmaco- and toxicokinetics. Albumin of humans and respective representatives in cattle and rodents have their own structural features that determine species differences in functional properties. However, albumin is not only passive, but also an active participant of pharmacokinetic and toxicokinetic processes, possessing a number of enzymatic activities. Numerous experiments have shown esterase or pseudoesterase activity of albumin towards a number of endogeneous and exogeneous esters. Due to the free thiol group of Cys34, albumin can serve as a trap for reactive oxygen and nitrogen species, thus participating in redox processes. Glycated albumin makes a significant contribution to the pathogenesis of diabetes and other diseases. The interaction of albumin with blood cells, blood vessels and tissue cells outside the vascular bed is of great importance. Interactions with endothelial glycocalyx and vascular endothelial cells largely determine the integrative role of albumin. This review considers the esterase, antioxidant, transporting and signaling properties of albumin, as well as its structural and functional modifications and their significance in the pathogenesis of certain diseases.

We studied the effect of different doses of ammonium chloride (ACl) and ammonium carbonate (ACr) on immunological parameters of the peripheral blood in rats during high-intensity exercise. Changes in the absolute and relative numbers of granulocytes, lymphocytes, natural killers, naive and mature effector cells one day after the end of the forced swimming cycle were found by using a hematological analyzer and a flow cytometer. Immunological indicators were analyzed relative to swimming duration on the last day of ultimate load. The revealed changes indicate the onset of the effector phase of the development of the inflammatory processes in the positive control group (physiological saline) and in rats receiving a higher dose of ACr (20 mg/kg), while administration of ACl prevented the development of inflammatory processes and shifts in the physiological balance of lymphocyte subpopulations. Immunological profiling indicates that ACl in a dose of 20 mg/kg most effectively improved physical performance in our forced swimming model.

Preconditioning is often used in medicine to protect organs from ischemic damage and in athletes to enhance the performances. We tested whether low-dose ammonium preconditioning (AMP) could have a beneficial effect on physical exercises (PE). We used Cardiopulmonary Exercise Testing (CPET) on a treadmill to investigate the effects of low-dose AMP on the physical exercise capacity of professional track and field athletes and tested twenty-five athletes. Because of the individual differences between athletes, we performed a preliminary treadmill test (Pre-test) and, according to the results, the athletes were randomly allocated into the AMP and control (placebo, PL) group based on the similarity of the total distance covered on a treadmill. In the AMP group, the covered distance increased (11.3 ± 3.6%, p < 0.02) compared to Pre-test. Similarly, AMP significantly increased O 2 uptake volume-VO 2 (4.6 ± 2.3%, p < 0.03) and pulmonary CO 2 output-VCO 2 (8.7 ± 2.8%, p < 0.01). Further, the basic blood parameters (pH, pO 2 , and lactate) shift was lower despite the greater physical exercise progress in the AMP group compared to Pre-test, whereas in the placebo group there were no differences between Pre-test and Load-test. Importantly, the AMP significantly increased red blood cell count (6.8 ± 2.0%, p < 0.01) and hemoglobin concentration (5.3 ± 1.9%, p < 0.01), which might explain the beneficial effects in physical exercise progress. For the first time, we showed that low-dose AMP had clear beneficial effects on submaximal PE., Competing Interests: The authors declare no conflict of interest.

Here, we propose a new approach for quantitative estimation of von Willebrand factor (vWF) exposed on the surface of endothelial cells (ECs) using the ARC1779 aptamer that interacts with the vWF A1 domain. To visualize complex formation between vWF and the aptamer, the latter was conjugated with the Cy5 fluorescent label. Cultured human umbilical vein endothelial cells (HUVEC) were stained with the ARC1779-Cy5 conjugate and imaged with a fluorescence microscope. The images were analyzed with the CellProfiler software. vWF released from the Weibel-Palade bodies was observed as bright dot-like structures of round and irregular shape, the number of which increased several times after HUVEC exposure to histamine or thrombin. Staining with ARC1779-Cy5 also revealed long filamentous vWF structures on the surface of activated HUVEC. vWF secretion by ECs is activated by the second messengers cAMP and Ca2+. There is evidence that hydrogen peroxide also acts as a second messenger in ECs. In addition, exogenous H2O2 formed in leukocytes can enter ECs. The aim of our study was to determine the effect of H2O2 on the vWF exposure at the surface of HUVEC using the proposed method. It was shown that hydrogen peroxide at concentration 100 µM, which is lower than the cytotoxicity threshold of H2O2 for cultured HUVEC, increased several times the number of dot-like structures and total amount of vWF exposed on plasma membrane of HUVEC, which suggest that H2O2 acts as a mediator that activates exocytosis of Weibel-Palade bodies and vWF secretion in the vascular endothelium during inflammation and upon elevated generation of endogenous reactive oxygen species in ECs.

Being one of the main proteins in the human body and many animal species, albumin plays a crucial role in the transport of various ions, electrically neutral molecules and in maintaining the colloidal osmotic pressure of the blood. Albumin is able to bind almost all known drugs, many nutraceuticals and toxic substances, determining their pharmaco- and toxicokinetics. However, albumin is not only the passive but also the active participant of the pharmacokinetic and toxicokinetic processes possessing a number of enzymatic activities. Due to the thiol group of Cys34, albumin can serve as a trap for reactive oxygen and nitrogen species, thus participating in redox processes. The interaction of the protein with blood cells, blood vessels, and also with tissue cells outside the vascular bed is of great importance. The interaction of albumin with endothelial glycocalyx and vascular endothelial cells largely determines its integrative role. This review provides information of a historical nature, information on evolutionary changes, inflammatory and antioxidant properties of albumin, on its structural and functional modifications and their significance in the pathogenesis of some diseases., Competing Interests: CONFLICT OF INTERESTSThe authors declare no obvious and potential conflicts of interest related to the publication of this article., (© Pleiades Publishing, Ltd. 2021.)

As a carrier of many biologically active compounds, blood is exposed to oxidants to a greater extent than the intracellular environment. Serum albumin plays a key role in antioxidant defence under both normal and oxidative stress conditions. This review evaluates data published in the literature and from our own research on the mechanisms of the enzymatic and non-enzymatic activities of albumin that determine its participation in redox modulation of plasma and intercellular fluid. For the first time, the results of numerous clinical, biochemical, spectroscopic and computational experiments devoted to the study of allosteric modulation of the functional properties of the protein associated with its participation in antioxidant defence are analysed. It has been concluded that it is fundamentally possible to regulate the antioxidant properties of albumin with various ligands, and the binding and/or enzymatic features of the protein by changing its redox status. The perspectives for using the antioxidant properties of albumin in practice are discussed., Competing Interests: The authors declare no conflict of interest. The funders had no role in the design of the study; in the collection, analyses or interpretation of data; in the writing of the manuscript, or in the decision to publish the results.

We compared the effects of two doses of ammonium chloride and ammonium carbonate (10 and 20 mg/kg) on the duration of swimming and blood lactate level. Ammonium chloride in a dose of 20 mg/kg was more efficient than in a dose of 10 mg/kg. The efficiency of ammonium carbonate in a dose of 10 mg/kg was similar to that of ammonium chloride in a dose of 20 mg/kg. Increasing the dose of ammonium carbonate to 20 mg/kg led to a decrease in the duration of swimming. On the last day of the experiment, lactate level in 5 min after exhausting load was maximum in control rats, while in rats treated with 10 mg/kg ammonium carbonate and 20 mg/kg ammonium chloride it was lower by 27 and 33%, respectively. In the control group, the amplitude of the decrease in lactate concentration in 1 h after load was 2-fold greater than in the group receiving ammonium chloride in a dose of 20 mg/kg and 1.6-fold greater that in groups treated with ammonium carbonate in a dose of 10 mg/kg and ammonium chloride in a dose of 20 mg/kg.

Ammonium, an end-product of catabolism, in low doses can promote adaptation of metabolic pathways in erythrocytes under conditions of extreme physical exercise. We compared the effects of two ammonium salts, ammonium chloride and ammonium carbonate, in two doses on biochemical parameters of rat erythrocytes 1 day after extreme physical exercise in a 4-week cycle of forced swimming. Of 16 analyzed parameters, the maximum number of significant shifts from the control was revealed in the groups of rats receiving ammonium chloride in doses of 20 and 10 mg/kg, and the minimal number of differences was found in groups treated with ammonium carbonate in the same doses. The comparison of the levels of reduced glutathione and 2.3-bisphosphoglicerate and activities of 5'-nucleotidase and Ca 2+ - and Na/K-ATPases attested to more rigorous control of the mechanism of oxygen delivery to tissues by erythrocytes after administration of ammonium chloride in a dose of 20 mg/kg.

Endothelial cells (ECs) line the blood vessels and lymphatic vessels, as well as heart chambers, forming the border between the tissues, on the one hand, and blood or lymph, on the other. Such a strategic position of the endothelium determines its most important functional role in the regulation of vascular tone, hemostasis, and inflammatory processes. The damaged endothelium can be both a cause and a consequence of many diseases. The state of the endothelium is indicated by the phenotype of these cells, represented mainly by (trans)membrane markers (surface antigens). This review defines endothelial markers, provides a list of them, and considers the mechanisms of their expression and the role of the endothelium in certain pathological conditions., Competing Interests: The authors declare that they have no conflict of interest., (© Pleiades Publishing, Ltd. 2020.)

One of the hallmarks of cancer is c hromosome in stability (CIN), which leads to aneuploidy, translocations, and other chromosome aberrations. However, in the vast majority of human tumors the molecular basis of CIN remains unknown, partly because not all genes controlling chromosome transmission have yet been identified. To address this question, we developed an experimental high-throughput imaging (HTI) siRNA assay that allows the identification of novel CIN genes. Our method uses a human artificial chromosome (HAC) expressing the GFP transgene. When this assay was applied to screen an siRNA library of protein kinases, we identified PINK1 , TRIO , IRAK1 , PNCK , and TAOK1 as potential novel genes whose knockdown induces various mitotic abnormalities and results in chromosome loss. The HAC-based assay can be applied for screening different siRNA libraries (cell cycle regulation, DNA damage response, epigenetics, and transcription factors) to identify additional genes involved in CIN. Identification of the complete spectrum of CIN genes will reveal new insights into mechanisms of chromosome segregation and may expedite the development of novel therapeutic strategies to target the CIN phenotype in cancer cells., (© 2019 Liskovykh et al.; Published by Cold Spring Harbor Laboratory Press.)

Second messenger nicotinic acid adenine dinucleotide phosphate (NAADP) triggers Ca 2+ release via two-pore channels (TPCs) localized in endolysosomal vesicles. The aim of the present work is to evaluate the role of TPCs in the action of norepinephrine (NE), angiotensin II (AngII), vasopressin (AVP), and 5-hydroxytriptamine (5-HT) on free cytoplasmic calcium concentration ([Ca 2+ ] i ) in smooth muscle cells (SMCs) isolated from rat aorta and on aorta contraction. To address this issue, the NAADP structural analogue and inhibitor of TPCs, NED 19, was applied. We have demonstrated a high degree of colocalization of the fluorescent signals of cis -NED 19 and endolysosmal probe LysoTracker in SMCs. Both cis - or trans -NED 19 inhibited the rise of [Ca 2+ ] i in SMCs induced by 100 μM NE by 50-60%. IC 50 for cis - and trans -NED 19 were 2.7 and 8.9 μM, respectively. The inhibition by NED 19 stereoisomers of the effects of AngII, AVP, and 5-HT was much weaker. Both forms of NED 19 caused relaxation of aortic rings preconstricted by NE, with relative potency of cis -NED 19 several times higher than that of trans -NED 19. Inhibition by cis -NED 19 of NE-induced contraction was maintained after intensive washing and slowly reversed within an hour of incubation. Cis - and trans -NED 19 did not cause decrease in the force of aorta contraction in response to Ang II and AVP, and only slightly relaxed aorta preconstricted by 5-HT and by KCl. Suppression of TPC1 in SMCs with siRNA caused a 40% decrease in [Ca 2+ ] i in response to NE, whereas siRNA against TPC2 did not change NE calcium signaling. These data suggest that TPC1 is involved in the NE-stimulated [Ca 2+ ] i rise in SMCs. Inhibition of TPC1 activity by NED 19 could be the reason for partial inhibition of aortic rings contraction in response to NE.

Poisoning by organophosphates (OPs) takes one of the leading places in the total number of exotoxicoses. Detoxication of OPs at the first stage of the poison entering the body could be achieved with the help of DNA- or RNA-aptamers, which are able to bind poisons in the bloodstream. The aim of the research was to develop an approach to rational in silico design of aptamers for OPs based on the example of paraoxon. From the published sequence of an aptamer binding organophosphorus pesticides, its three-dimensional model has been constructed. The most probable binding site for paraoxon was determined by molecular docking and molecular dynamics (MD) methods. Then the nucleotides of the binding site were mutated consequently and the values of free binding energy have been calculated using MD trajectories and MM-PBSA approach. On the basis of the energy values, two sequences that bind paraoxon most efficiently have been selected. The value of free binding energy of paraoxon with peripheral anionic site of acetylcholinesterase (AChE) has been calculated as well. It has been revealed that the aptamers found bind paraoxon more effectively than AChE. The peculiarities of paraoxon interaction with the aptamers nucleotides have been analyzed. The possibility of improving in silico approach for aptamer selection is discussed., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

In this study, we investigated the effects of NAD(P)H oxidase (NOX) inhibitor VAS2870 (3-benzyl-7-(2-benzoxazolyl)thio-1,2,3-triazolo[4,5-d]pyrimidine) on the histamine-induced elevation of free cytoplasmic calcium concentration ([Ca 2+ ] i ) and the secretion of von Willebrand factor (vWF) in human umbilical vein endothelial cells (HUVECs) and on relaxation of rat aorta in response to histamine. At 10 μM concentration, VAS2870 suppressed the [Ca 2+ ] i rise induced by histamine. Inhibition was not competitive, with IC50 3.64 and 3.22 μM at 1 and 100 μM concentrations of histamine, respectively. There was no inhibition of [Ca 2+ ] i elevation by VAS2870 in HUVECs in response to the agonist of type 1 protease-activated receptor SFLLRN. VAS2870 attenuated histamine-induced secretion of vWF and did not inhibit basal secretion. VAS2870 did not change the degree of histamine-induced relaxation of rat aortic rings constricted by norepinephrine. We suggest that NOX inhibitors might be used as a tool for preventing thrombosis induced by histamine release from mast cells without affecting vasorelaxation.

Hydrogen peroxide, formed in the endothelium, acts as a factor contributing to the relaxation of blood vessels. The reason for this vasodilatory effect could be modulation by H 2 O 2 of calcium metabolism, since mobilization of calcium ions in endothelial cells is a trigger of endothelium-dependent relaxation. The aim of this work was to investigate the influence of H 2 O 2 on the effects of Ca 2+ -mobilizing agonists in human umbilical vein endothelial cells (HUVEC). We have found that H 2 O 2 in concentration range 10-100  μ M increases the rise of [Ca 2+ ] i induced by 5-hydroxytryptamine (5-HT) and carbachol and does not affect the calcium signals of ATP, agonist of type 1 protease-activated receptor SFLLRN, histamine and bradykinin. Using specific agonists of 5-HT1B and 5-HT2B receptors CGS12066B and BW723C86, we have demonstrated that H 2 O 2 potentiates the effects mediated by these types of 5-HT receptors. Potentiation of the effect of BW723C86 can be produced by the induction of endogenous oxidative stress in HUVEC. We have shown that the activation of 5-HT2B receptor by BW723C86 causes production of reactive oxygen species (ROS). Inhibitor of NADPH oxidases VAS2870 suppressed formation of ROS and partially inhibited [Ca 2+ ] i rise induced by BW723C86. Thus, it can be assumed that vasorelaxation induced by endogenous H 2 O 2 in endothelial cells partially occurs due to the potentiation of the agonist-induced calcium signaling.

We studied the role of calcium-regulating structures of slow- (m. soleus, SOL) and fast-twitch (m. extensor digitorum longus, EDL) skeletal muscles of rats during adaptation to exhausting physical activity and the possibility of modulating this adaptation with decaffeinated green tea extract. It was established that EDL adaptation is mainly aimed at Са 2+ elimination from the sarcoplasm by Са-ATPase and its retention in the reticulum by calsequestrin. Administration of green tea extract increased endurance due to involvement of slow-twitch muscles whose adaptation is associated with enhanced expression of all the studied genes responsible for the regulation of Ca 2+ balance.

The albumin molecule, in contrast to many other plasma proteins, is not covered with a carbohydrate moiety and can bind and transport various molecules of endogenous and exogenous origin. The enzymatic activity of albumin, the existence of which many scientists perceive skeptically, is much less studied. In toxicology, understanding the mechanistic interactions of organophosphates with albumin is a special problem, and its solution could help in the development of new types of antidotes. In the present work, the history of the issue is briefly examined, then our in silico data on the interaction of human serum albumin with soman, as well as comparative in silico data of human and bovine serum albumin activities in relation to paraoxon, are presented. Information is given on the substrate specificity of albumin and we consider the possibility of its affiliation to certain classes in the nomenclature of enzymes., Competing Interests: The authors confirm that there are no conflicts of interest.

Glycosaminoglycans (GAGs) in the urine of Wistar rats, and on the surface of the epithelium and lamina propria of the bladder, were quantitatively assessed before and after acute intoxication with paraoxon or cyclophosphamide. Paraoxon was administered subcutaneously (s.c.) twice with an interval of 1h to a final dose of 275mg/kg; cyclophosphamide was administered intraperitoneally (i.p.) with a single dose of 100mg/kg or to a final dose of 240mg/kg (three times per 80mg/kg every 12h). GAGs on the surface of the epithelium and lamina propria of the urinary bladder of rats were quantitatively determined by Alcian blue dye staining. GAGs in the urine were determined spectrophotometrically with 1-9-dimethyl methylene blue. By 48-96h after intoxication with either paraoxon or cyclophosphamide, statistically significant increases in the concentration of GAGs were obtained both for the tissues of the bladder and the urine of rats. Cyclophosphamide, in contrast to paraoxon, caused the development of hemorrhagic cystitis in the animals. The synthesis of GAGs is considered to be compensatory response to the toxic xenobiotics., (Copyright © 2017 Elsevier GmbH. All rights reserved.)

Hydrogen peroxide at concentrations below cytotoxic ones causes an increase in the cytoplasmic calcium concentration in human umbilical vein endothelial cells as a result of calcium release from intracellular stores. Two-pore calcium channel blocker trans-NED19 partially suppresses the increase in the level of calcium ions in the cells in response to the addition of hydrogen peroxide. The staining of endothelial cells with the fluorescent stereoisomer cis-NED19 and LysoTracker confirmed the localization of two-pore calcium channels in lysosomes and endolysosomal vesicles.

In blood vessels, serotonin 5-HT2B receptors mainly mediate relaxation, although their activation by the selective agonist BW723C86 is known to exert contraction of aorta in deoxycorticosterone acetate (DOCA)-salt and N(omega)-nitro-l-arginine (l-NAME) hypertensive rats [Russel et al., 2002; Banes et al., 2003] and in mice with type 2 diabetes [Nelson et al., 2012]. The unmasking effect on vasoconstriction can be caused by a shift in the balance of tyrosine phosphorylation in smooth muscle cells (SMC) due to oxidative stress induced inhibition of protein tyrosine phosphatases (PTP). We have demonstrated that BW723C86 which does not cause contraction of rat aorta and mesenteric artery rings, evoked a vasoconstrictor effect in the presence of PTP inhibitors sodium orthovanadate (Na 3 VO 4 ) or BVT948. BW723C86 induced a weak rise of [Ca 2+ ] i in the SMC isolated from rat aorta; however, after pre-incubation with Na 3 VO 4 the response to BW723C86 increased more than 5-fold. This effect was diminished by protein tyrosine kinase (PTK) inhibitor genistein, inhibitor of Src-family kinases PP2, inhibitor of NADPH-oxidase VAS2870 and completely suppressed by N-acetylcysteine and 5-HT2B receptor antagonist RS127445. Using fluorescent probe DCFH-DA we have shown that Na 3 VO 4 induces oxidative stress in SMC. In the presence of Na 3 VO 4 BW723C86 considerably increased formation of reactive oxygen species while alone had no appreciable effect on DCFH oxidation. We suggest that oxidative stress causes inhibition of PTP and unmasking of 5-HT2B receptors functional activity., (Copyright © 2016 Elsevier Inc. All rights reserved.)

The metabolic profile of plasma of white non-linear rats was investigated under normal conditions and after chronic inhalational exposure to low doses of aliphatic hydrocarbons with the number of carbon atoms from 6 to 10. Metabolic profile was determined with combination of gas chromatography-mass spectrometry and high performance, high resolution liquid chromatography-mass spectrometry with subsequent use of chemometrical methods for data treatment and presentation. It was shown that continuous 90-day exposure to a mixture of C₆-C₁₀ saturated hydrocarbons at concentration of 160 ± 20.5 mg/m³ results in various impairments of metabolic processes in liver and kidneys. Exposure to hydrocarbons at doses of 31.4 ± 5.6 mg/m³ and 5.2 ± 1.8 mg/m³ evoked significantly smaller changes. Novel metabolic markers of the toxic effect of low concentrations of C₆-C₁₀ aliphatic hydrocarbons were revealed. The ratio of concentrations of pyrophosphoric and oxalic acids in rat blood plasma was found to be the most sensitive marker called <>. A hypothesis is put forward about the redox balance violation as the leading pathogenetic mechanism of neuropathies and concomitant pathologies associated with hydrocarbon chronic intoxication.

Endothelium is a community of endothelial cells (ECs), which line the blood and lymphatic vessels, thus forming an interface between the tissues and the blood or lympha. This strategic position of endothelium infers its indispensable functional role in controlling vasoregulation, haemostasis, and inflammation. The state of endothelium is simultaneously the cause and effect of many diseases, and this is coupled with modifications of endothelial phenotype represented by markers and with biochemical profile of blood represented by biomarkers. In this paper, we briefly review data on the functional role of endothelium, give definitions of endothelial markers and biomarkers, touch on the methodological approaches for revealing biomarkers, present an implicit role of endothelium in some toxicological mechanistic studies, and survey the role of reactive oxygen species (ROS) in modulation of endothelial status.

The blocker of two-pore channels trans-NED 19 and hydrogen peroxide were found to inhibit histamine-induced relaxation of rat-aorta. The degree of inhibition depended on histamine concentration. The relaxation in response to I μM histamine of rat aorta preconstricted with 30 mM KCI, serotonin, or endothelin- 1, was completely abolished by 30 μM trans-NED 19. On the other hand, trans-NED 19 decreased the relaxation of the aorta in the presence of 10 μM histamine only by 2.1-fold to 2.4-fold, and there was almost no inhibition by trans-NED 19 of the relaxationinduced by 100 ptM histamine.) Relaxation of precontracted with serotonin aorta in response to 10 and 100 μM histamine was reduced by hydrogen peroxide (200 M) by 10- and 2.5-fold, respectively. Suppression of aorta relaxation by trans-NED 19 and H202 correlated with their inhibitory effect on the histamine-induced increase in the cytoplasmic free calcium concentration in human umbilical vein endothelial cells. With the use of a fluorescent probe LysoTracker, the cis-NED19 binding sites were demonstrated to be localized in endolysosomes of the endothelial cells. These data indicate that two-pore calcium channels participate in the histamine-induced endothelium-dependent relaxation of rat aorta. Furthermore, their functional role is exhibited much more clearly at low histamine concentrations. We suggest that hydrogen peroxide evokes depletion of intracellular calcium depots thereby suppressing the response to histamine.

A general and concise approach to thermally and hydrolytically stable alkyl 2,3-dihydroazete-2,3-di-/2,2,3-tricarboxylates from alkyl 2-bromoazirine-2-carboxylates or 4-bromo-5-alkoxyisoxazoles is reported. The synthesis involves the formation of 2-azabuta-1,3-diene by the reaction of rhodium carbenoid with isoxazole or azirine followed by cyclization/hydrodebromination cascade. The latter reaction is the first example of the selective hydrodehalogenation of a valence isomer under equilibrium conditions. In vitro cytotoxicity tests on THP-1 cell line revealed that the 2,3-dihydroazetes greatly differ in their ability to induce apoptosis and/or necrosis. To adequately describe and quantitatively assess these properties, the difference between the two areas under the curves of concentration dependency of apoptosis/necrosis induction within the concentration range was used. Trimethyl 4-phenyl-2,3-dihydroazete-2,2,3-tricarboxylate was found to display the maximal apoptotic potential coupled with high cytotoxic and minimal necrotic potential.

Accumulating data indicates that chromosome instability (CIN) common to cancer cells can be used as a target for cancer therapy. At present the rate of chromosome mis-segregation is quantified by laborious techniques such as coupling clonal cell analysis with karyotyping or fluorescence in situ hybridization (FISH). Recently, a novel assay was developed based on the loss of a non-essential human artificial chromosome (HAC) carrying a constitutively expressed EGFP transgene ("loss of signal" assay). Using this system, anticancer drugs can be easily ranked on by their effect on HAC loss. However, it is problematic to covert this "loss of signal" assay into a high-throughput screen to identify drugs and mutations that increase CIN levels. To address this point, we re-designed the HAC-based assay. In this new system, the HAC carries a constitutively expressed shRNA against the EGFP transgene integrated into human genome. Thus, cells that inherit the HAC display no green fluorescence, while cells lacking the HAC do. We verified the accuracy of this "gain of signal" assay by measuring the level of CIN induced by known antimitotic drugs and added to the list of previously ranked CIN inducing compounds, two newly characterized inhibitors of the centromere-associated protein CENP-E, PF-2771 and GSK923295 that exhibit the highest effect on chromosome instability measured to date. The "gain of signal" assay was also sensitive enough to detect increase of CIN after siRNA depletion of known genes controlling mitotic progression through distinct mechanisms. Hence this assay can be utilized in future experiments to uncover novel human CIN genes, which will provide novel insight into the pathogenesis of cancer. Also described is the possible conversion of this new assay into a high-throughput screen using a fluorescence microplate reader to characterize chemical libraries and identify new conditions that modulate CIN level.

Angiotensin II (Ang II) is an important mammalian neurohormone involved in reninangiotensin system. Ang II is produced both constitutively and locally by RAS systems, including white fat adipocytes. The influence of Ang II on adipocytes is complex, affecting different systems of signal transduction from early Са(2+) responses to cell proliferation and differentiation, triglyceride accumulation, expression of adipokine-encoding genes and adipokine secretion. It is known that white fat adipocytes express all RAS components and Ang II receptors (АТ1 and АТ2). The current work was carried out with the primary white adipocytes culture, and Са(2+) signaling pathways activated by Ang II were investigated using fluorescent microscopy. Са(2+)-oscillations and transient responses of differentiated adipocytes to Ang II were registered in cells with both small and multiple lipid inclusions. Using inhibitory analysis and selective antagonists, we now show that Ang II initiates periodic Са(2+)-oscillations and transient responses by activating АТ1 and АТ2 receptors and involving branched signaling cascades: 1) Ang II → Gq → PLC → IP3 → IP3Rs → Ca(2+) 2) Gβγ → PI3Kγ → PKB 3) PKB → eNOS → NO → PKG 4) CD38 → cADPR → RyRs → Ca(2+) In these cascades, AT1 receptors play the leading role. The results of the present work open a perspective of using Ang II for correction of signal resistance of adipocytes often observed during obesity and type 2 diabetes., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Albumin molecule, unlike molecules of many other plasma proteins, is not covered with carbohydrate shell. It plays a crucial role in maintaining of colloid osmotic pressure of the blood, and is able to bind and transport various endogenous and exogenous molecules. The enzymatic activity of albumin, the existence and the role of which most researchers are still skeptical to accept, is of the main interest to us. In this review, a history of the issue is traced, with particular attention to the esterase activity of albumin. The kinetic and thermodynamic characteristics of the interaction of albumin with some substrates are adduced, and possibility of albumin being attributed to certain groups of Enzyme Nomenclature is considered.