The effect of differences in the third domain of the glycoprotein E of tick-borne encephalitis virus of the Far Eastern, Siberian and European subtypes on the binding of recombinant D3 proteins with a chimeric antibody

Currently, a therapeutic drug based on recombinant antibodies for the prevention and treatment of tick-borne encephalitis virus (TBEV) is developed in ICBFM SB RAS, and the chimeric antibody ch14D5 is considered as one of the key components of this drug. It was previously shown that this antibody is directed to the domain D3 of the glycoprotein E of TBEV. It was previously shown that this antibody is able to protect mice from the European subtype of TBEV, strain “Absettarov”, and the presence of virus-neutralizing activity against the Far Eastern subtype of TBEV, strain 205 was also shown for this antibody. However, it remains unclear whether this antibody exhibits selectivity for different subtypes of TBEV. The aim of this study was to investigate the effect of amino acid sequence differences of recombinant D3 domains derived from the glycoprotein E of TBEV of the Far Eastern, Siberian and European subtypes on the binding of the protective antibody ch14D5 to these proteins. Using Western blot analysis and surface plasmon resonance, it was shown that ch14D5 antibody has the highest affinity (KD= 1.7±0.5 nM) for the D3 domain of the TBEV of the “Sofjin-Ru” strain belonging to the Far Eastern subtype of the virus. At the same time, the affinity of ch14D5 antibody for similar D3 proteins derived from “Zausaev”, “1528-99” and “Absettarov” strains of the Siberian and European subtypes of TBEV was noticeably lower (KD= 25±4, 300±50, 250±50 nM, respectively). In addition, information about the spatial arrangement of amino acid residues that are different for the studied recombinant proteins indicates that the epitope recognized by the ch14D5 antibody is in close proximity to the lateral ridge of D3 domain of E glycoprotein

Microtubule Dynamics Regulate Cyclic Stretch-Induced Cell Alignment in Human Airway Smooth Muscle Cells

Microtubules are structural components of the cytoskeleton that determine cell shape, polarity, and motility in cooperation with the actin filaments. In order to determine the role of microtubules in cell alignment, human airway smooth muscle cells were exposed to cyclic uniaxial stretch. Human airway smooth muscle cells, cultured on type I collagen-coated elastic silicone membranes, were stretched uniaxially (20% in strain, 30 cycles/min) for 2 h. The population of airway smooth muscle cells which were originally oriented randomly aligned near perpendicular to the stretch axis in a time-dependent manner. However, when the cells treated with microtubule disruptors, nocodazole and colchicine, were subjected to the same cyclic uniaxial stretch, the cells failed to align. Lack of alignment was also observed for airway smooth muscle cells treated with a microtubule stabilizer, paclitaxel. To understand the intracellular mechanisms involved, we developed a computational model in which microtubule polymerization and attachment to focal adhesions were regulated by the preexisting tensile stress, pre-stress, on actin stress fibers. We demonstrate that microtubules play a central role in cell re-orientation when cells experience cyclic uniaxial stretching. Our findings further suggest that cell alignment and cytoskeletal reorganization in response to cyclic stretch results from the ability of the microtubule-stress fiber assembly to maintain a homeostatic strain on the stress fiber at focal adhesions. The mechanism of stretch-induced alignment we uncovered is likely involved in various airway functions as well as in the pathophysiology of airway remodeling in asthma

Контролируемые исследования по эффективности поликомпонентной вакцины при иммунотерапии у больных с хроническими обструктивными заболеваниями органов дыхания

During the hard controlled study, the effeciency of polycomponent vaccina designed by SRI of I.I.Mechnikov was investigated. The satisfactory and excelent effect was found in 71.4% of patients, that expressed in the listening of remission, the decrease of the exacerbation frequency and the severity of the disease course, the decrease of the quantity of medication, the improvement of respiration. Vaccina therapy induces the increase of the level of antibodies to all the antigen components of the vaccina in the patients. The number of patients with low titres of the antibodies decreases and the number of patients with high ones increases. Vaccina therapy with VP-4 used in according to the offered medication scheme does not induce the increase of the total IgE level in serum. There is the sharp decrease of this parameter in patients with the initial high level of IgE.В строго контролируемом опыте изучена эффективность поликомпонентной вакцины НИИ ВС им. И.И. Мечникова. Установлен хороший и отличный терапевтический эффект у 71,4% больных, что выражается в удлинении сроков ремиссии у больных, снижении частоты обострений и тяжести течения заболевания, сокращении количества принимаемых лекарств, улучшении показателей функции внешнего дыхания. Вакцинотерапия вызывает у больных подъем уровня антител ко всем антигенным компонентам вакцины. Уменьшается число больных с низкими титрами и увеличивается количество больных с высокими титрами антител. Вакцинотерапия ВП-4 по предложенной схеме введения не вызывает подъем уровня общего иммуноглобулина Е в сыворотке крови больных. У больных с исходно высоким уровнем иммуноглобулина Е отмечается выраженное снижение этого показателя

Lasp-1 Regulates Podosome Function

Eukaryotic cells form a variety of adhesive structures to connect with their environment and to regulate cell motility. In contrast to classical focal adhesions, podosomes, highly dynamic structures of different cell types, are actively engaged in matrix remodelling and degradation. Podosomes are composed of an actin-rich core region surrounded by a ring-like structure containing signalling molecules, motor proteins as well as cytoskeleton-associated proteins

Dynamic Modeling of Cell Migration and Spreading Behaviors on Fibronectin Coated Planar Substrates and Micropatterned Geometries

An integrative cell migration model incorporating focal adhesion (FA) dynamics, cytoskeleton and nucleus remodeling, actin motor activity, and lamellipodia protrusion is developed for predicting cell spreading and migration behaviors. This work is motivated by two experimental works: (1) cell migration on 2-D substrates under various fibronectin concentrations and (2) cell spreading on 2-D micropatterned geometries. These works suggest (1) cell migration speed takes a maximum at a particular ligand density (~1140 molecules/µm2) and (2) that strong traction forces at the corners of the patterns may exist due to combined effects exerted by actin stress fibers (SFs). The integrative model of this paper successfully reproduced these experimental results and indicates the mechanism of cell migration and spreading. In this paper, the mechanical structure of the cell is modeled as having two elastic membranes: an outer cell membrane and an inner nuclear membrane. The two elastic membranes are connected by SFs, which are extended from focal adhesions on the cortical surface to the nuclear membrane. In addition, the model also includes ventral SFs bridging two focal adhesions on the cell surface. The cell deforms and gains traction as transmembrane integrins distributed over the outer cell membrane bond to ligands on the ECM surface, activate SFs, and form focal adhesions. The relationship between the cell migration speed and fibronectin concentration agrees with existing experimental data for Chinese hamster ovary (CHO) cell migrations on fibronectin coated surfaces. In addition, the integrated model is validated by showing persistent high stress concentrations at sharp geometrically patterned edges. This model will be used as a predictive model to assist in design and data processing of upcoming microfluidic cell migration assays

Comparative Dynamics of Retrograde Actin Flow and Focal Adhesions: Formation of Nascent Adhesions Triggers Transition from Fast to Slow Flow

Dynamic actin network at the leading edge of the cell is linked to the extracellular matrix through focal adhesions (FAs), and at the same time it undergoes retrograde flow with different dynamics in two distinct zones: the lamellipodium (peripheral zone of fast flow), and the lamellum (zone of slow flow located between the lamellipodium and the cell body). Cell migration involves expansion of both the lamellipodium and the lamellum, as well as formation of new FAs, but it is largely unknown how the position of the boundary between the two flow zones is defined, and how FAs and actin flow mutually influence each other. We investigated dynamic relationship between focal adhesions and the boundary between the two flow zones in spreading cells. Nascent FAs first appeared in the lamellipodium. Within seconds after the formation of new FAs, the rate of actin flow decreased locally, and the lamellipodium/lamellum boundary advanced towards the new FAs. Blocking fast actin flow with cytochalasin D resulted in rapid dissolution of nascent FAs. In the absence of FAs (spreading on poly-L-lysine-coated surfaces) retrograde flow was uniform and the velocity transition was not observed. We conclude that formation of FAs depends on actin dynamics, and in its turn, affects the dynamics of actin flow by triggering transition from fast to slow flow. Extension of the cell edge thus proceeds through a cycle of lamellipodium protrusion, formation of new FAs, advance of the lamellum, and protrusion of the lamellipodium from the new base

Tubulin Tyrosination Is Required for the Proper Organization and Pathfinding of the Growth Cone

International audienceBACKGROUND: During development, neuronal growth cones integrate diffusible and contact guidance cues that are conveyed to both actin and microtubule (MT) cytoskeletons and ensure axon outgrowth and pathfinding. Although several post-translational modifications of tubulin have been identified and despite their strong conservation among species, their physiological roles during development, especially in the nervous sytem, are still poorly understood. METHODOLOGY/FINDINGS: Here, we have dissected the role of a post-translational modification of the last amino acid of the alpha-tubulin on axonal growth by analyzing the phenotype of precerebellar neurons in Tubulin tyrosin ligase knock-out mice (TTL(-/-)) through in vivo, ex vivo and in vitro analyses. TTL(-/-) neurons are devoid of tyrosinated tubulin. Their pathway shows defects in vivo, ex vivo, in hindbrains open-book preparations or in vitro, in a collagen matrix. Their axons still orient toward tropic cues, but they emit supernumerary branches and their growth cones are enlarged and exhibit an emission of mis-oriented filopodia. Further analysis of the TTL(-/-) growth cone intracellular organization also reveals that the respective localization of actin and MT filaments is disturbed, with a decrease in the distal accumulation of Myosin IIB, as well as a concomitant Rac1 over-activation in the hindbrain. Pharmacological inhibition of Rac1 over-activation in TTL(-/-) neurons can rescue Myosin IIB localization. CONCLUSIONS/SIGNIFICANCE: In the growth cone, we propose that tubulin tyrosination takes part in the relative arrangement of actin and MT cytoskeletons, in the regulation of small GTPases activity, and consequently, in the proper morphogenesis, organization and pathfinding of the growth cone during development

Nudel and FAK as Antagonizing Strength Modulators of Nascent Adhesions through Paxillin

Competition for binding to the cellular protein paxillin by the proteins Nudel and focal adhesion kinase is important for the proper regulation of cell adhesion and migration