Evaluation of Reference Gene Stability for Investigations of Intracellular Signalling in Human Cancer and Non-Malignant Mesenchymal Stromal Cells.

Background: Real-time reverse transcription quantitative polymerase chain reaction (RT-qPCR) is a powerful tool for analysing target gene expression in biological samples. To achieve reliable results by RT-qPCR, the most stable reference genes must be selected for proper data normalisation, particularly when comparing cells of different types. We aimed to choose the least variable candidate reference genes among eight housekeeping genes tested within a set of human cancer cell lines (HeLa, MCF-7, SK-UT-1B, A549, A431, SK-BR-3), as well as four lines of normal, non-malignant mesenchymal stromal cells (MSCs) of different origins.

EGF, TGF- and Amphiregulin Differently Regulate Endometrium-Derived Mesenchymal Stromal/Stem Cells.

The prototypical receptor tyrosine kinase epidermal growth factor receptor (EGFR) is regulated by a set of its ligands, which determines the specificity of signaling and intracellular fate of the receptor. The EGFR signaling system is well characterized in immortalized cell lines such as HeLa derived from tumor tissues, but much less is known about EGFR function in untransformed multipotent stromal/stem cells (MSCs). We compared the effect of epidermal growth factor (EGF), transforming growth factor-α (TGF-α) and amphiregulin (AREG) on physiological responses in endometrial MSCs (enMSC) and HeLa cells.

Effect of Ply Orientation on the Mechanical Performance of Carbon Fibre Honeycomb Cores.

Carbon fibres used as a honeycomb core material (subject to a proper in-depth analysis of their reinforcement patterns) allows solving the thermo-dimensional stability problem of the units for space systems. Based on the results of numerical simulations with the support of finite element analysis, the paper provides an evaluation of the accuracy of analytical dependencies for the determination of the moduli of elasticity of a carbon fibre honeycomb core in tension/compression and shear. It is shown that a carbon fibre honeycomb reinforcement pattern has a significant impact on the mechanical performance of the carbon fibre honeycomb core.

Outcomes of Deferoxamine Action on HO-Induced Growth Inhibition and Senescence Progression of Human Endometrial Stem Cells.

Mesenchymal stem cells (MSCs) are broadly applied in regenerative therapy to replace cells that are lost or impaired during disease. The low survival rate of MSCs after transplantation is one of the major limitations heavily influencing the success of the therapy. Unfavorable microenvironments with inflammation and oxidative stress in the damaged regions contribute to MSCs loss.

[Creating the conditions for skeletal traction in mandibular condyle fracture in children and adolescents].

In this article the possibility of using the Clark appliance (Twin-block) for creating conditions for skeletal traction in case of fraction of mandibular condyle in children and teenagers is presented. The proposed method of using the double-jaw orthodontic appliance Twin-block in case of mandibular fractures with condylar displacement promotes release of traumatized condyle and mandible replacement to correct position, promotes optimal conditions for myofunctional balance, fragment consolidation and remodeling of affected temporomandibular joint.

Functional cycle of EEA1-positive early endosome: Direct evidence for pre-existing compartment of degradative pathway.

Early endosomes, regarded as the main sorting station on endocytic pathway, are characterized by high frequency of homotypic fusions mediated by tethering protein EEA1. Despite intensive investigations, biogenesis of endosomes, boundaries between early and late endosomes, and process of cargo transition though them remain obscure. Here, using EGF/EGFR endocytosis as a model and confocal microscopy of fixed and live cells, we provide evidence favoring EEA1-vesicles being pre-existed vesicular compartment, that maintains its resident proteins' level and is sensitive to biosynthetic, but not endocytic pathway disturbance.

Improving the selection efficiency of the counter-selection marker pheS* for the genetic engineering of Bacillus amyloliquefaciens.

Bacillus subtilis pheS was genetically modified to obtain a counter-selection marker with high selection efficiency in Bacillus amyloliquefaciens. The application of the new replication-thermosensitive integrative vector pNZTM1, containing this marker, pheS, with a two-step replacement recombination procedure provides an effective tool for the genetic engineering of industrially important Bacillus species.

Senescence-messaging secretome factors trigger premature senescence in human endometrium-derived stem cells.

Accumulating evidence suggests that the senescence-messaging secretome (SMS) factors released by senescent cells play a key role in cellular senescence and physiological aging. Phenomenon of the senescence induction in human endometrium-derived mesenchymal stem cells (MESCs) in response to SMS factors has not yet been described. In present study, we examine a hypothesis whether the conditioned medium from senescent cells (CM-old) may promote premature senescence of young MESCs.

Quantitative analysis of the heterogeneous population of endocytic vesicles.

The quantitative characterization of endocytic vesicles in images acquired with microscope is critically important for deciphering of endocytosis mechanisms. Image segmentation is the most important step of quantitative image analysis. In spite of availability of many segmentation methods, the accurate segmentation is challenging when the images are heterogeneous with respect to object shapes and signal intensities what is typical for images of endocytic vesicles.

Mobility of tethering factor EEA1 on endosomes is decreased upon stimulation of EGF receptor endocytosis in HeLa cells.

Tethering factor EEA1, mediating homotypic fusion of early endosomes, was shown to be localized in membrane-bound state both in serum-deprived and stimulated for EGF receptor endocytosis cells. However, it is not known whether dynamics behavior of EEA1 is affected by EGF stimulation. We investigated EEA1 cytosol-to-membrane exchange rate in interphase HeLa cells by FRAP analysis.

EEA1 CARRYING VESICLES ARE NOT AUTOPHAGOSOMES IN SERUM-DEPRIVED HeLa CELLS.

According to current model, stimulation of EGF-receptor endocytosis results in recruitment onto early endosomes cytosolic tether protein EEA1 necessary for their further fusions. However, EEA1-positive vesicles are found in the cells not treated with growth factor, that were incubated in serum-free conditions. It is known also that prolonged serum deprivation induces autophagosomes formation, the process possibly involving endocytic compartments.

Quantum dots implementation as a label for analysis of early stages of EGF receptor endocytosis: a comparative study on cultured cells.

EGF complexed to fluorescent photostable quantum dots by biotin-streptavidin system (bEGF-savQD) is attractive for both the basic research and therapeutic application such as targeted drug delivery in EGF-receptor (EGFR) expressing cancers. However, compared to native EGF, the large size of QD and its quasi-multivalency can have unpredictable effects on EGFR endocytosis changing the internalization portal and/or endosomal processing tightly bound to EGF signaling. We have found that bEGF-savQDs enter HeLa cells via the temperature-dependent clathrin-mediated EGF-receptor-specific pathway characteristic for native EGF.

[Analysis of vesicle subpopulations carrying early endosomal autoantigen EEA1].

Confocal immunofluorescent analysis of interphase HeLa cells has demonstrated that involved in regulation of homotypic fusions early endosomal autoantigene EEA1 is associated with vesicles represented by two populations differing in apparent size, localization and the level of bound EEA1. Before analysis the cells have been preincubated in serum-deprived medium for 12 h to minimize ligand-dependent endocytosis of serum growth factors. The first subpopulation is mainly represented by large vesicles strongly decorated with EEA1.

[Changes in the heart of neonatal rats after cryptosporidial gastroenteritis of different degrees of severity].

Disturbances at the childhood age increase risk of the appearance of cardiovascular diseases decades later. The nature of this interconnection called ontogenetic programming is not completely understood. Valuable sources of knowledge about mechanisms of ontogenetic programming are data of interspecies study of biology of the body life cycles and of heart physiological capabilities.

[Adherence to guidelines on management of acute coronary syndrome in Russian hospitals and outcomes of hospitalization (data from the RECORD-2 Registry)].

Background: Complete following existing guidelines for management of acute coronary syndrome (ACS) is known to be associated with better outcomes. Partly this is explained by lesser adherence to recommendations in high risk patients. Aim of our study was to assess relationship between degree of following current guidelines and in hospital outcomes independently from initial assessment of risk.

[Threshold rat neonatal cardiomyocyte response to gradual cryptosporidial infection severity increase].

Infectious gastroenteritis is one of the common causes of tachyarrythmia, malabsorbtion and growth retardation in children. Our recent studies have indicated that neonatal.cryptosporidial gastroenteritis is associated with long-term cardiomyocyte abnormalities.

[Analysis of EGF receptor endocytosis dynamics based on semiquantative processing of confocal immunofluorescent images of fixed cells].

Endocytosis of signaling receptors EGF receptor in particular, starting at the plasma membrane and finishing in perinuclear lysosomes implies endosomes' multiple interactions with homotypic endosomes and vesicles of other origin (lysosomes, trans-Golgi network), which results in endosomal size changes. In addition, the characteristics of endocytic pathway is endosomes' translocation from cell periphery to juxtanuclear region. Thus, endocytosis as a highly dynamic process develops in time and space.

[Assessment of the prognostic value of the CRUSADE score in patients with acute coronary syndromes hospitalized in a noninvasive hospital].

Background. The CRUSADE score was created for assessment of bleeding risk in patients with acute coronary syndrome (ACS). However its accuracy was not confirmed in populations of patients treated in Russian hospitals, in noninvasively managed patients, in patients without myocardial infarction (MI).

[Cystatin C and bleedings during hospitalization of noninvasively treated patients with acute coronary syndromes].

Purpose: To assess relationship between blood level of a marker of renal function cystatin C (CC) and bleeding events in patients with acute coronary syndromes (ACS).

Material And Methods: CC level was measured in 160 patients from the ACS registry conducted in a noninvasive hospital in Moscow. Each month we included in this registry 25 consecutively hospitalized patients and recorded deaths and bleeding events (major and moderate by TIMI and/or GUSTO criteria) during hospitalization.

[Factors associated with inhospital severe bleeding in noninvasively treated patients with non ST elevation acute coronary syndrome].

Unlabelled: Risk factors of bleeding in acute coronary syndrome (ACS) were derived from data of clinical trials and registers with high proportion of invasively treated patients. Aim of this study was to elucidate factors associated with severe bleeding in patients with non ST-elevation (NSTE) ACS treated in a noninvasive hospital.

Material And Methods: From November 2009 to April 2011 24 NSTEACS patients had severe GUSTO or TIMI bleeding.

[Cardiomyocyte myosin heavy chain composition change after cryptosporidial gastroenteritis].

Cardiovascular diseases are the most common case of human death in developed countries. Thus, the discovering of their new risk factors is of primary importance. Based on epidemiology studies, vertebrate life-history traits comparison and cross-species cardiomyocyte transcriptome analysis, we suggest that one of these factors could be infectious gastroenteritis.

[Generation of dopamine neurons from human embryonic stem cells in vitro].

The aim of the study was to generate dopaminergic (DA) neurons from human embryonic stem cells (ESC) in vitro. It was shown that human ESCs are able to differentiated into DA neurons without co-culture with stromal cells. Terminal differentiation into DA neurons was reached by successive application of noggin and bFGF growth factors on collagen and matrigel substrates during 3-4 weeks.

[Acetylation of microtubules during endocytosis of epidermal growth factor receptor (c-ErbB1) in interphase HeLa cells].

Acetylation of microtubules (MT) during endocytosis of epidermal growth factor receptor, c-ErbB1, was studied by confocal immunofluorescence microscopy. It was found that stimulation of endocytosis of c-ErbB1 complexed with the epidermal growth factor (EGF), resulted in continuous raising of MT acetylation that reached its maximum at 60-90 min and then went down to the control level. Simultaneously, the receptor-containing endosomes grew in size and were redistributed into juxtranuclear region.

[Remodeling of microtubule system during EGF receptor endocytosis].

The idea of microtubules (MTs) as of passive railway tracks, along which transport vesicles travel by use of motor proteins, is widely accepted. In the present work the organization of MT system during EGF-receptor endocytosis was investigated by indirect double immunofluorescence in HeLa and A431 cell lines. Stimulation of cells with EGF resulted in formation of EGF receptor-containing peripheral vesicular endosomes.