Репозиторий Университета

Human mucin MUC1 plays an important role in cancer development. The increased level of this molecule expression during cancer cell progression induces metastasis and is associated with poor prognosis for patients. There is a large body of experimental data on the role of various functional domains of human mucin MUC1 in metastasis. While, the cytoplasmic domain determined to play a definitive role, the influence of extracellular domain on cancer cell invasiveness still remains unclear. The present paper reveals that the extracellular domain of MUC1 molecule consists of two functional subdomains-the region of tandem repeats (TR) and the region of irregular repeats (IR). We demonstrate the ability of each of these subdomains to alter the invasiveness of cancer cells. The presence of the MUC1 molecules containing TR subdomain (MUC1-TR) on the surface of low-invasive cancer cells leads to the increase in their transendothelial migration potency, while the addition of the IR subdomain to the MUC1-TR molecule (MUC1-IR-TR) restores their natural low invasiveness. J. Cell. Biochem. 118: 4002-4011, 2017. © 2017 Wiley Periodicals, Inc.

Despite rapid progress, many problems and limitations persist and limit applicability of gene editing techniques. Making use of meganucleases, TALENs or CRISPR/Cas9-based tools requires an initial step of pre-screening to determine the efficiency and specificity of the designed tools. This step remains time- and material-consuming. Here, we propose a simple, cheap, reliable, time-saving and highly sensitive method to evaluate a given gene editing tool based on its capacity to induce chromosomal translocations when combined with a reference engineered nuclease. In the proposed technique designated as “ENIT” for Engineered Nucleases-Induced Translocations, a plasmid coding for the DNA-editing tool to be tested is co-transfected into carefully chosen target cells along with that for an engineered nuclease of known specificity and efficiency. If the new enzyme efficiently cuts within the desired region, specific chromosomal translocations will be generated between the two targeted genomic regions and be readily detectable by a one-step PCR or qPCR assay. The PCR product thus obtained can be directly sequenced thereby determining the exact position of the double-strand breaks induced by the gene-editing tools. As a proof of concept, ENIT was successfully tested in different cell types and with different meganucleases, TALENs and CRISPR/Cas9-based editing tools.

(PDF) A one-step PCR-based assay to evaluate efficiency and precision of genomic DNA-editing tools. Available from: https://www.researchgate.net/publication/314714362_A_one-step_PCR-based_assay_to_evaluate_efficiency_and_precision_of_genomic_DNA-editing_tools [accessed Dec 25 2018].

Цель работы - оценка состояния кортикального цитоскелета волокон камбаловидной мышцы крысы в результате 6-часового антиортостатического вывешивания на фоне предшествующего, в течение 3 сут по 100 мкг/сут, введения смеси фосфатидилхолинов (лецитина). Данные о периметре волокон, толщине подмембранного цитоскелета и доле разрывов в нем относительно периметра получали, используя иммуногистохимическую окраску на альфа-актинин-4. Периметр волокон оставался неизменным во всех группах исследования. При этом доля разрывов была выше в группах вывешивания, чем в соответствующих контрольных группах. Толщина окрашенного слоя, соответствующего подмембранному цитоскелету, не менялась в контрольных группах и в группе вывешивания без лецитина, однако в группе вывешивания на фоне лецитина она достоверно увеличивалась по сравнению с соответствующей контрольной группой на 27 % (p < 0,05) соответственно.

Purpose of the work was to assess the cortical cytoskeleton of m. soleus fibers from rats after 6-hr tail-suspension preceded by 3 days of phosphatidylcholine (lecithin) injections at a dose of 100 µг/d. Data about the fiber perimeter, submembrane cytoskeleton thickness and percentage of bonds breaks along the perimeter were obtained using the alpha-actinin-4 anti-body stain. The fiber perimeter remained unchanged in all groups under study. However, the percentage of breaks was high in suspension groups but not in respective groups of control. Thickness of the stained layer commensurate to the submembrane cytoskeleton did not change in the control groups and in the suspension group without lecithin injections but increased reliably in the suspension group that recieved injections by 27 % (p < 0.05) in comparison with its control.

Lanthanide-doped upconversion nanoparticles (UCNPs) have recently attracted great attention in theranostics due to their exceptional optical and physicochemical properties, which enable the design of a novel UCNP-based nanoplatform for luminescent imaging, temperature mapping, sensing, and therapy. In addition, UCNPs are considered to be ideal building blocks for development of multimodal probes for cells and whole body imaging, exploiting simple variation of host matrix, dopant ions, and surface chemistry. Modalities responsible for magnetic resonance imaging (MRI), computed tomography (CT), and positron emission tomography (PET)/single-photon emission computed tomography (SPECT) are embedded in a single UC nanocrystal, providing integrating effect over any modality alone in terms of the efficiency and sensitivity for clinical innovative diagnosis through multimodal bioimaging. In particular, we demonstrate applications of UCNPs as a new nanoplatform for optical and multimodal cancer imaging in vitro and in vivo and extend discussions to delivery of UCNP-based therapeutic agents for photodynamic and photothermal cancer treatments.

Background Prostaspheres-based three dimensional (3D) culture models have provided insight into prostate cancer (PCa) biology, highlighting the importance of cell–cell interactions and the extracellular matrix (EMC) in the tumor microenvironment. Although these 3D classical spheroid platforms provide a significant advance over 2D models mimicking in vivo tumors, the limitations involve no control of assembly and structure with only limited spatial or glandular organization. Here, matrix-free prostaspheres from human metastatic prostate carcinoma PC3 and DU145 cell lines and their respective gemcitabine resistant (GemR) variants were generated by using cyclic Arg-Gly-Asp-D-Phe-Lys peptide modified with 4-carboxybutyl-triphenylphosphonium bromide (cyclo-RGDfK(TPP)). Materials and methods Microscopic imaging, immunocytochemistry (ICC), flow cytometry, sialidase, and WST-1 cell viability assays were used to evaluate the formation of multicellular tumor spheroid (MCTS), cell survival, morphologic changes, and expression levels of α2,6 and α2,3 sialic acid (SA) and E- and N-cadherin in DU145, PC3, and their GemR variants. Results By using the cyclo-RGDfK(TPP) peptide platform in a dose- and time-dependent manner, both DU145 and DU145GemR cells formed small MCTS. In contrast, PC3 and PC3GemR cells formed irregular multicellular aggregates at all concentrations of cyclo-RGDfK(TPP) peptide, even after 6 days of incubation. ICC and flow cytometry results revealed that DU145 cells expressed higher amounts of E-cadherin but lower N-cadherin compared with PC3 cells. By using Maackia amurensis (α2,3-SA-specific MAL-II) and Sambucus nigra (α2,6-SA specific SNA) lectin-based cytochemistry staining and flow cytometry, it was found that DU145 and DU145GemR cells expressed 5 times more α2,6-SA than α2,3-SA on the cell surface. PC3 cells expressed 4 times more α2,3-SA than α2,6-SA, and the PC3GemR cells showed 1.4 times higher α2,6-SA than α2,3-SA. MCTS volume was dose-dependently reduced following pretreatment with α2,6-SA-specific neuraminidase (Vibrio cholerae). Oseltamivir phosphate enhanced cell aggregation and compaction of 3D MCTS formed with PC3 cells. Conclusion The relative levels of specific sialoglycan structures on the cell surface correlate with the ability of PCa cells to form avascular multicellular prostaspheres.

V. Poly(vinyl alcohol) (PVA) hydrogels are widely employed for various biomedical applications, including tissue engineering, due to their biocompatibility, high water solubility, low protein adsorption, and chemical stability. However, non-charged surface of PVA-based hydrogels is not optimal for cell adhesion and spreading. Here, cross-linked macroporous hydrogels based on low molecular weight acrylated PVA (Acr-PVA) was synthesized by modification of the pendant alcohol groups on the PVA with glycidyl methacrylate (GMA). To enhance cell affinity, charged groups were introduced to the hydrogel composition. For this purpose, Acr-PVA was copolymerized with either negatively charged acrylic acid (AA) or positively charged 2-(diethylamino) ethyl methacrylate (DEAEMA) monomers. A surface charge of the obtained hydrogels was found to be in function of the co-monomer type and content. Confocal microscopy observations confirmed that adhesion and spreading of both mouse fibroblasts (L929) and human mesenchymal stem cells (hMSC) on the modified Acr-PVA-AA and Acr-PVA-DEAEMA hydrogels were better than those on the non-modified Acr-PVA hydrogel. The increase of DEAEMA monomer content from 5 to 15 mol% resulted in the enhancement of cell viability which was 1.5-fold higher for Acr-PVA-DEAEMA-15 hydrogel than that of the non-modified Acr-PVA hydrogel sample.

Immunotoxin 4D5scFv-PE40 is a recombinant protein that comprises 4D5scFv antibody as a targeting module and fragment of Pseudomonas exotoxin A as an effector (toxic) one. The immunotoxin has shown pronounced antitumor effect on cancer cells overexpressing HER2 receptor in vitro and on HER2-positive experimental tumors in vivo. We clarified the mechanism of 4D5scFv-PE40 activity that is of particular importance in the case of targeted therapeutic agent aimed at personalizing treatment of disease in relation to molecular genetic characteristics of each patient. After specific binding to HER2 on the cell surface and clathrin-mediated endocytosis the immunotoxin passes through retrograde trafficking route. During this route the immunotoxin molecule is supposed to undergo enzymatic processing that ends in separation of C-terminal and N-terminal fragments of the immunotoxin. Finally, C-terminal functionally active fragment of 4D5scFv-PE40 arrests protein synthesis in cytoplasm followed by cell death via apoptosis.

(PDF) HER2-specific recombinant immunotoxin 4D5scFv-PE40 passes through retrograde trafficking route and forces cells to enter apoptosis. Available from: https://www.researchgate.net/publication/314188519_HER2-specific_recombinant_immunotoxin_4D5scFv-PE40_passes_through_retrograde_trafficking_route_and_forces_cells_to_enter_apoptosis [accessed Dec 25 2018].

Statement of significance: KillerRed is a protein photosensitizer that holds promise as an alternative for the existing hydrophobic photosensitizers that are widely used in clinical photodynamic therapy (PDT). However, applications of KillerRed to deep-seated tumours are limited by the insufficient penetration depth of the excitation light in highly scattering and absorbing biological tissues. Herein, we reported the deployment of upconversion nanoparticles (UCNPs) to enhance the treatment depth of KillerRed by converting the deep-penetrating near-infrared (NIR) light to upconversion photoluminescence and activating the PDT effect of KillerRed under deep tissues. This work demonstrated clear potential of UCNPs as the NIR-to-visible light converter to overcome the light penetration limit that has plagued PDT application for many years.

Formation of long-lived coloured forms of indoline spirocompounds (spirooxazines and spiropyrans) molecules under matrix immobilization in polar polymer of different chemical structure using supercritical fluid impregnation is studied. Some of such systems acquire additional optical characteristics besides colour – they become fluorophors. Mechanisms of observed changes in optical properties and long-term stability of the resulting coloured forms of indoline spirooxazine in polymers are discussed. Electronic absorption spectra of such coloured forms were recorded in supercritical carbon dioxide medium as well as on air. The results can be applied for creation of optical, biomedical and electronic devices and stable fluorescence composite coverings.

Green Chemistry is one of the most important and practically used tools to integrate principles of sustainable development and green economy in the field of chemistry and the chemical industry in various countries. There is a number of metrics in the field of green chemistry. The research presented is an original algorithm of multi-criterion green process assessment for renewable raw materials bioconversion. The algorithm is used when the process of obtaining the same target substance N is possible to be carried out in many ways (or under different conditions). In this case, the researcher task is to choose the best process in compliance with the principles of green chemistry. The multiple-factor complex assessment is to be used to choose the optimum process conditions. The algorithm designed was tested for efficiency in choosing the optimal processes of acid hydrolysis of deproteinized meals. The deproteinized sunflower meal preprocessing efficiency analysis was carried out during the course of its microbiological conversion into a vegetable protein and carbohydrate feed supplement taking into account the principles of green chemistry. The methodology testing was based on the experimental data obtained by chemical hydrolysis of deproteinized residues, and two-stage pretreatment process of deproteinized residues processing comprising the steps of chemical and enzymatic hydrolysis. The combination of chemical and enzymatic hydrolysis in two-stage deproteinized sunflower meal processing was justified. The proposed algorithm allowed not only to determine the effective ranges of parameters of deproteinized sunflower meal processing, but also allowed to justify the possibility of its optimization due to the both processes combined.

Objectives: Central (hypogonadotropic) hypogonadism in women could be a cause of persistent amenorrhea and hypoestrogenemia as observed in postmenopause. This study aimed to compare the clinical, hormonal and biochemical features in women with non-physiological (central hypogonadism) and physiological (postmenopause) hypoestrogenemia.

Methods: A total of 161 young women, median age 24.9 years (interquartile range (IQR) 21.2; 30.5) with central hypogonadism (with isolated hypogonadotropic hypogonadism, n = 76, and with hypopituitarism, n = 85), 53 healthy young women, median age 23.9 years (IQR 23.1; 28.0) and 50 healthy postmenopausal women, median age 56.0 years (IQR 53.1; 58.5), were examined. Psychoemotional, neurovegetative and urogenital symptoms, sex steroid levels, parameters of lipid and mineral metabolism were evaluated.

Results: In young women with central hypogonadism, the frequencies of psychoemotional, neurovegetative and urogenital complaints differed significantly from those in healthy young women and were similar to those in postmenopausal women. Concentrations of estradiol, testosterone, dehydroepiandrosterone sulfate, parameters of lipid and mineral metabolism as well as quality of life in women with central hypogonadism were not typical of healthy young women but were similar to those of postmenopausal women of middle/old age.

Conclusions: Despite the young age of women with central hypogonadism, clinical, hormonal and biochemical abnormalities were similar in many aspects to those in postmenopausal women at middle/old age. These revealed features could be considered as signs of premature aging in young amenorrheic women with low gonadotropin levels.

Metabolomics, a 'budding' discipline, may accurately reflect a specific phenotype which is sensitive to genetic and epigenetic interactions. This rapidly evolving field in science has been proposed as a tool for the evaluation of the effects of epigenetic factors, such as nutrition, environment, drug and lifestyle on phenotype. Urine, being sterile, is easy to obtain and as it contains metabolized or non‑metabolized products, is a favored study material in the field of metabolomics. Urine organic acids (OAs) reflect the activity of main metabolic pathways and have been used to assess health status, nutritional status, vitamin deficiencies and response to xenobiotics. To date, a limited number of studies have been performed which actually define reference OA values in a healthy population and as reference range for epigenetic influences, and not as a reference to congenital metabolic diseases. The aim of the present study was thus the determination of reference values (RVs) for urine OA in a healthy adult population. Targeted metabolomics analysis of 22 OAs in the urine of 122 healthy adults by gas chromatography‑mass spectrometry, was conducted. Percentile distributions of the OA concentrations in urine, as a base for determining the RVs in the respective population sample, were used. No significant differences were detected between female and male individuals. These findings can facilitate the more sensitive determination of OAs in pathological conditions. Therefore, the findings of this study may contribute or add to the information already available on urine metabolite databases, and may thus promote the use of targeted metabolomics for the evaluation of OAs in a clinical setting and for pathophysiological evaluation. However, further studies with well‑defined patients groups exhibiting specific symptoms or diseases are warranted in order to discern between normal and pathological values.

Glycogen synthase kinase 3 (GSK3) has been linked to the mechanisms of stress, mood regulation, and the effects of antidepressants. The functions of the GSK3β isoform have been extensively investigated, but little is known about the α-isoform, although they may functionally related. In a recently established modified swim test with a third delayed swim exposure, brain GSK3β mRNA expression positively correlated with floating behavior on the third test. A two-week-long pretreatment regime with imipramine (7.5mg/kg/day) or thiamine (200mg/kg/day), which is known to have antidepressant properties, reduced the GSK3β over-expression and decreased floating behavior on Day 5. GSK3α mRNA levels were measured in the hippocampus and prefrontal cortex on Days 1, 2 and 5. GSK3α expression was decreased in the prefrontal cortex on Day 2 and increased on Day 5. In this model, GSK3α mRNA changes were prevented by imipramine or thiamine treatment. There was a significant correlation between the expression of the two isoforms in the prefrontal cortex on Day 2 in untreated group. These results provide the first evidence for the potential involvement of GSK3α in depressive-like behaviours and as a target of anti-depressant therapy. Furthermore, the correlations suggest some cross-talk may exist between the two GSK3 isoforms.

A high-calorie diet (HCD) induces two mutually exacerbating effects contributing to diet-induced obesity (DIO): impaired glucose metabolism and increased food consumption. A link between the metabolic and behavioral manifestations is not well understood yet. We hypothesized that chronic inflammation induced by HCD plays a key role in linking together the two components of diet-induced pathology. Based on this hypothesis, we tested if a plasmid (DNA vaccine) encoding p62 (SQSTM1) would alleviate DIO including its metabolic and/or food consumption abnormalities. Previously we reported that injections of the p62 plasmid reduce chronic inflammation during ovariectomy-induced osteoporosis. Here we found that the p62 plasmid reduced levels of pro-inflammatory cytokines IL-1β, IL-12, and INFγ and increased levels of anti-inflammatory cytokines IL-4, IL-10 and TGFβ in HCD-fed animals. Due to this anti-inflammatory response, we further tested whether the plasmid can alleviate HCD-induced obesity and associated metabolic and feeding impairments. Indeed, p62 plasmid significantly reversed effects of HCD on the body mass index (BMI), levels of glucose, insulin and glycosylated hemoglobin (HbA1c). Furthermore, p62 plasmid partially restored levels of the satiety hormone, serotonin, and tryptophan, simultaneously reducing activity of monoamine oxidase (MAO) in the brain affected by the HCD. Finally, the plasmid partially reversed increased food consumption caused by HCD. Therefore, the administering of p62 plasmid alleviates both metabolic and behavioral components of HCD-induced obesity.

Elenagen is a plasmid encoding p62/SQSTM1, the first DNA vaccine possessing two mutually complementing mechanisms of action: it elicits immune response against p62 and mitigates systemic chronic inflammation. Previously, Elenagen demonstrated anti-tumor efficacy and safety in rodent tumor models and spontaneous tumors in dogs. This multicenter I/IIa trial evaluated safety and clinical activity of Elenagen in patients with advanced solid tumors. Fifteen patients were treated with escalating doses of Elenagen (1-5 mg per doses, 5 times weekly) and additional 12 patients received 1 mg dose. Ten patients with breast and ovary cancers that progressed after Elenagen were then treated with conventional chemotherapy. Adverse events (AE) were of Grade 1; no severe AE were observed. Cumulatively twelve patients (44%) with breast, ovary, lung, renal cancer and melanoma achieved stable disease for at least 8 wks, with 4 of them (15%) had tumor control for more than 24 wks, with a maximum of 32 wks. The patients with breast and ovary cancers achieved additional tumor stabilization for 12-28 wks when treated with chemotherapy following Elenagen treatment. Therefore, Elenagen demonstrated good safety profile and antitumor Oncotarget 2 www.impactjournals.com/oncotarget

(PDF) Safety and efficacy of p62 DNA vaccine ELENAGEN in a first-in-human trial in patients with advanced solid tumors. Available from: https://www.researchgate.net/publication/316148229_Safety_and_efficacy_of_p62_DNA_vaccine_ELENAGEN_in_a_first-in-human_trial_in_patients_with_advanced_solid_tumors [accessed Dec 25 2018].

The formation of hydrophilic pores in lipid bilayer during phase transition is described using Smolukhowski's equation with an additional term of the hydrophobic pore source. This term is added to account for defects in lipid packing during phase transition. We assume that the temporal sequence of the pores is a stochastic process, a non -stationary second- order Erlang flow. Flow characteristics depend on the equation solution and determine the formation times of the hydrophilic pore. The calculated distribution of the durations of intervals between hydrophilic pore formations is in a good agree ment with experimental data published before. In terms of this model we describe the influence of poly (ethylene glycol) on the pore formation frequency.

Background: Colonic microbiome is thought to be involved in auto-immune multiple sclerosis (MS). Interactions between diet and the colonic microbiome in MS are unknown. Methods: We compared the composition of the colonic microbiota quantitatively in 25 MS patients and 14 healthy controls.Fluorescence in situ hybridization (FISH) with 162 ribosomal RNA derived bacterial FISH probes was used. Ten of the MS patients received a ketogenic diet for 6 months. Changes in concentrations of 35 numerically substantial bacterial groups were monitored at baseline and at 2, 12, and 23/24 weeks. Results: No MS typical microbiome pattern was apparent.The total concentrations and diversity of substantial bacterial groups were reduced in MS patients (P < 0.001). Bacterial groups detected with EREC (mainly Roseburia), Bac303 (Bacteroides), and Fprau (Faecalibacterium prausnitzii) probes were diminished the most. The individual changes were multidirectional and inconsistent. The effects of a ketogenic diet were biphasic. In the short term, bacterial concentrations and diversity were further reduced. They started to recover at week 12 and exceeded significantly the baseline values after 23–24 weeks on the ketogenic diet. Conclusions: Colonic biofermentative function is markedly impaired in MS patients.The ketogenic diet normalized concentrations of the colonic microbiome after 6 months.

Our investigation of 1004 faecal specimens from European bats for picornaviruses by broadly reactive nested reverse transcription-PCR found picornaviral RNA in 28 samples (2.8 %). Phylogenetic analysis of the partial 3D genomic region suggested that one bat virus belonged to the species Enterovirus G (EV-G, formerly Porcine enterovirus B). Bat infection was supported by relatively high EV-G concentrations of 1.1×106 RNA copies per gram of faeces. All other bat viruses belonged either to the bat-associated genus Mischivirus, or to an unclassified Picornaviridae group distantly related to the genus Sapelovirus. Members of this unclassified sapelovirus-related group had RNA secondary structures in their 3′-nontranslated regions that were typical of enteroviruses and that resembled structures that occur in bat-associated coronaviruses, suggesting ancient recombination events. Based on sequence distances, several picornaviruses from European and Chinese bats were likely conspecific, suggesting connectivity of virus populations. Due to their high mutation rates and their diversity, picornaviruses may be useful tools for studies of bat and virus ecology.

Цель данного исследования — изучение функциональных последствий эстетической ринопластики. Авторы на достаточно большом объеме клинического материала (253 пациента) сравнили результаты операций в зависимости от хирургического доступа, объема и техники вмешательства и выявили, что постринопластические нарушения носового дыхания достоверно чаще встречаются после «эндоназального» хирургического доступа, чем после «открытой» ринопластики (75% против 68%), функциональный результат вмешательства существенно лучше при одноэтапном проведении риносептопластики и внутриносового вмешательства (конхопластика, синусотомия) (СОП 629±98 см3/с против 397±65 см3/с по объективной оценке, p<0,05) и применение во время первичной ринопластики структуросохраняющих приемов (расширяющие трансплантаты, расширяющие лоскуты, укрепляющие трансплантаты) позволяет уменьшить выраженность и частоту функциональных нарушений в отдаленные сроки наблюдения (СОП 550±85 см3/с против 303±50 см3/с по объективной оценке, p<0,05).