Nanoscale, Voltage-Driven Application of Bioactive Substances onto Cells with Organized Topography

With Scanning Ion Conductance Microscopy (SICM), a non-contact scanning probe technique, it is possible to obtain information about both the surface topography of live cells and to apply molecules onto specific nanoscale structures. The technique is therefore widely used to apply chemical compounds and to study the properties of molecules on the surface of various cell types. The heart muscle cells, the cardiomyocytes, possess a highly elaborate, unique surface topography including T-tubule openings leading into a cell internal system which exclusively har-bors many proteins necessary for the cells physiological function. Here we applied Isoproterenol into these surface openings by changing the applied voltage over the SICM nanopipette. To determine the grade of precision of our application we used finite element simulations to inves-tigate how the concentration profile varies over the cell surface. We first obtained topography scans of the cardiomyocytes using SICM and then determined the electrophoretic mobility of Isoproterenol in a high ion solution to be -7×10-9 m2/Vs. The simulations showed that the delivery to the T-tubule opening is highly confined to the underlying Z-groove and especially to the first T-tubule opening, where the concen-tration is approximately 6.5 times higher compared to on a flat surface under the same delivery settings. Delivery to the crest, instead of the T-tubule opening, resulted in a much lower concentration, emphasizing the importance of topography on agonist delivery. In conclusion SICM, unlike other techniques, can reliably deliver precise quantities of compounds to the T-tubules of cardiomyocyte

Modelling of reservoir sewage currents regime on the basis of software for evaluation of shore strengthening constructions stability

The results of laboratory and field surveys of Vileysko-Minskaya hydrologic system discharge currents were presented. Software for reservoirs currents dynamical pressure counting was developed basing on the results. Modeling in water bodies let invent calculation algorithms for currents speed spread mode counting and banks fixing constructions stability evaluation

Methods and results of natural inspections of Byelorussian water objects as sources of technogenic character emergency situations

Methodology and results of Belarus artificial water bodies’ field surveys were presented. Based on these results potential risk coefficient for hydrodynamic accidents possibility estimation was counted. Belarus water bodies were classified as potential sources of natural emergencies

T-tubule remodelling disturbs localised β2-adrenergic signalling in rat ventricular myocyte during the progression of heart failure

Aims Cardiomyocyte β2-adrenergic receptor (β2AR) cyclic adenosine monophosphate (cAMP) signalling is regulated by the receptors’ subcellular location within transverse tubules (T-tubules), via interaction with structural and regulatory proteins, which form a signalosome. In chronic heart failure (HF), β2ARs redistribute from T-tubules to the cell surface, which disrupts functional signalosomes and leads to diffuse cAMP signalling. However, the functional consequences of structural changes upon β2AR-cAMP signalling during progression from hypertrophy to advanced HF are unknown. Methods and results Rat left ventricular myocytes were isolated at 4-, 8-, and 16-week post-myocardial infarction (MI), β2ARs were stimulated either via whole-cell perfusion or locally through the nanopipette of the scanning ion conductance microscope. cAMP release was measured via a Förster Resonance Energy Transfer-based sensor Epac2-camps. Confocal imaging of di-8-ANNEPS-stained cells and immunoblotting were used to determine structural alterations. At 4-week post-MI, T-tubule regularity, density and junctophilin-2 (JPH2) expression were significantly decreased. The amplitude of local β2AR-mediated cAMP in T-tubules was reduced and cAMP diffused throughout the cytosol instead of being locally confined. This was accompanied by partial caveolin-3 (Cav-3) dissociation from the membrane. At 8-week post-MI, the β2AR-mediated cAMP response was observed at the T-tubules and the sarcolemma (crest). Finally, at 16-week post-MI, the whole cell β2AR-mediated cAMP signal was depressed due to adenylate cyclase dysfunction, while overall Cav-3 levels were significantly increased and a substantial portion of Cav-3 dissociated into the cytosol. Overexpression of JPH2 in failing cells in vitro or AAV9.SERCA2a gene therapy in vivo did not improve β2AR-mediated signal compartmentation or reduce cAMP diffusion. Conclusion Although changes in T-tubule structure and β2AR-mediated cAMP signalling are significant even at 4-week post-MI, progression to the HF phenotype is not linear. At 8-week post-MI the loss of β2AR-mediated cAMP is temporarily reversed. Complete disorganization of β2AR-mediated cAMP signalling due to changes in functional receptor localization and cellular structure occurs at 16-week post-MI

Mechanistic basis for Sgo1-mediated centromere localization and function of the CPC.

Centromere association of the chromosomal passenger complex (CPC; Borealin-Survivin-INCENP-Aurora B) and Sgo1 is crucial for chromosome biorientation, a process essential for error-free chromosome segregation. Phosphorylated histone H3 Thr3 (H3T3ph; directly recognized by Survivin) and histone H2A Thr120 (H2AT120ph; indirectly recognized via Sgo1), together with CPC's intrinsic nucleosome-binding ability, facilitate CPC centromere recruitment. However, the molecular basis for CPC-Sgo1 binding and how their physical interaction influences CPC centromere localization are lacking. Here, using an integrative structure-function approach, we show that the "histone H3-like" Sgo1 N-terminal tail-Survivin BIR domain interaction acts as a hotspot essential for CPC-Sgo1 assembly, while downstream Sgo1 residues and Borealin contribute for high-affinity binding. Disrupting Sgo1-Survivin interaction abolished CPC-Sgo1 assembly and perturbed CPC centromere localization and function. Our findings reveal that Sgo1 and H3T3ph use the same surface on Survivin to bind CPC. Hence, it is likely that these interactions take place in a spatiotemporally restricted manner, providing a rationale for the Sgo1-mediated "kinetochore-proximal" CPC centromere pool