Polyethyleneimines (PEIs) are efficient non-viral vectors for gene transfer. Heparan sulfate proteoglycans have been proposed to be the cell-surface receptors for PEI⅐DNA complexes (polyplexes). Here, we investigated if syndecan-1 (SDC1) and syndecan-2 (SDC2) are involved in PEI-mediated transfection. Following addition of polyplexes to HEK293 cells, green fluorescent protein-tagged SDCs rapidly formed clusters with PEI that were dependent of lipid raft integrity. However, although SDC1 overexpression slightly enhanced PEI-mediated gene expression, SDC2 dramatically inhibited it. Confocal microscopy analysis showed that SDC1⅐polyplex endocytosis occurred within minutes after addition of polyplexes, whereas SDC2⅐polyplex endocytosis took hours. Expression of SDC1 cytoplasmic deletion mutants revealed that the SDC1 cytoplasmic tail is required for gene expression, but not for clustering or endocytosis, whereas overexpression of SDC1/SDC2 chimeras showed that the SDC2 ectodomain is responsible for the inhibitory effect on gene transfer. This study provides evidence that SDCs may have opposing effects on PEI-mediated transfection.

No abstract

Several key mutations in the Spike protein receptor binding domain (RBD) have been identified to influence its affinity for the human Angiotensin-Converting Enzyme 2 (ACE2). Here, we perform a comparative study of the ACE2 binding to the wild type (Wuhan) RBD and some of its variants: Alpha B.1.1.7, Beta B.1.351, Delta B.1.617.2, Kappa B.1.617.1, B.1.1.7 + L452R and Omicron B.1.1.529. Using a coiled-coil mediated tethering approach of ACE2 in a novel surface plasmon resonance (SPR)-based assay, we measured interactions at different temperatures. Binding experiments at 10 °C enhanced the kinetic dissimilarities between the RBD variants and allowed a proper fit to a Langmuir 1:1 model with high accuracy and reproducibility, thus unraveling subtle differences within RBD mutants and ACE2 glycovariants. Our study emphasizes the importance of SPR-based assay parameters in the acquisition of biologically relevant data and offers a powerful tool to deepen our understanding of the role of the various RBD mutations in ACE2 interaction binding parameters.

In Romania, with time, settlements located along the main roads have developed and transformed into linear towns, with significant local and connection traffic, important administrative, economic, commercial and touristic activities concentrated in the central area, as well as pedestrian traffic of over 200 pedestrians per hour in the main pedestrian crossings on the route.The object of the present study is made by a series of junctions situated on National Road 1 in Busteni town, on a dangerous road sector. For this study, traffic measurements, simulations and suggestions for improving the existing situation were made.Based on the simulated traffic flows, there were performed capacity analysis with PTV Vissim and Traficware Synchro softwares, and were developed appropriate planning solutions for the intersections, resulting in tables with extracted performance indicators based on micro simulation of the traffic values. Also planning solutions for horizontal design and proposals for traffic lights were made for junctions that can not operate under priority traffic on one direction or which are presenting traffic safety risk.Based on the traffic data, it was taken in consideration the necessity to make planning proposals and to develop design solutions immediately applicable, with minimum intervention.Solutions will refer to the geometric planning of the intersections, but with new plans and timings for traffic lights, including proposals for new equipment; regulating the traffic flow: development/ refurbishment of intersections and pedestrian crossings; optimization of routing programs in order to achieve a higher level of service and more efficient traffic control indicators; segregation of pedestrian movements by vehicles traffic, implementation of physical devices to lock / channel the traffic.