Advanced vehicle-to-everything (V2X) safety applications must operate with ultra-low latency and be highly reliable. Therefore, they require sophisticated supporting technologies. This is especially ...true for cooperative applications, such as Collective Perception (CP), where a large amount of data constantly flows among vehicles and between vehicles and a network intelligence server. Both low and high-level support is needed for such an operation, meaning that various access technologies and other architectural elements also need to incorporate features enabling the effective use of V2X applications with strict requirements. The new 5G core architecture promises even more supporting technologies, like Multi-access Edge Computing (MEC). To test the performance of these technologies, an integrated framework for V2X simulations with 5G network elements is proposed in the form of combining Simu5G, a standalone 5G implementation, with the go-to V2X-simulator, Artery. As a first step toward a fully functional MEC-assisted CP Service, an extension to Simu5G’s edge implementation is introduced. The edge application is responsible for dispatching the Collective Perception Messages generated by the vehicles via the 5G connectivity so that a MEC server provided by the network can process incoming data. Simulation results prove the operability of the proposed integrated system and edge computing’s potential in assisting V2X scenarios.
A prior peripheral nerve injury in vivo, promotes a rapid elongated mode of sensory neurons neurite regrowth in vitro. This in vitro model of conditioned axotomy allows analysis of the cellular and ...molecular mechanisms leading to an improved neurite re-growth. Our differential interference contrast microscopy and immunocytochemistry results show that conditioned axotomy, induced by sciatic nerve injury, did not increase somatic size of adult lumbar sensory neurons from mice dorsal root ganglia sensory neurons but promoted the appearance of larger neurites and growth cones. Using atomic force microscopy on live neurons, we investigated whether membrane mechanical properties of growth cones of axotomized neurons were modified following sciatic nerve injury. Our data revealed that neurons having a regenerative growth were characterized by softer growth cones, compared to control neurons. The increase of the growth cone membrane elasticity suggests a modification in the ratio and the inner framework of the main structural proteins.
Autophagy is mediated by membrane-bound organelles and it is an intrinsic catabolic and recycling process of the cell, which is very important for the health of organisms. The biogenesis of ...autophagic membranes is still incompletely understood. In vitro studies suggest that Atg2 protein transports lipids presumably from the ER to the expanding autophagic structures. Autophagy research has focused heavily on proteins and very little is known about the lipid composition of autophagic membranes. Here we describe a method for immunopurification of autophagic structures from Drosophila melanogaster (an excellent model to study autophagy in a complete organism) for subsequent lipidomic analysis. Western blots of several organelle markers indicate the high purity of the isolated autophagic vesicles, visualized by various microscopy techniques. Mass spectrometry results show that phosphatidylethanolamine (PE) is the dominant lipid class in wild type (control) membranes. We demonstrate that in Atg2 mutants (Atg2−), phosphatidylinositol (PI), negatively charged phosphatidylserine (PS), and phosphatidic acid (PA) with longer fatty acyl chains accumulate on stalled, negatively charged phagophores. Tandem mass spectrometry analysis of lipid species composing the lipid classes reveal the enrichment of unsaturated PE and phosphatidylcholine (PC) in controls versus PI, PS and PA species in Atg2−. Significant differences in the lipid profiles of control and Atg2− flies suggest that the lipid composition of autophagic membranes dynamically changes during their maturation. These lipidomic results also point to the in vivo lipid transport function of the Atg2 protein, pointing to its specific role in the transport of short fatty acyl chain PE species.
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•Immunoisolation of early autophagic organelles from Drosophila for lipidomics•Autophagosomal membranes are dominated by PE species.•Phagophores in Atg2− are enriched in PI and negatively charged PS and PA species.•Autophagic membranes are highly unsaturated and dynamic in their lipid composition.•PE species are likely in vivo targets of Atg2 lipid transporter.
Reverse transcription-quantitative real-time PCR (RT-qPCR) is a ubiquitously used method in biological research, however, finding appropriate reference genes for normalization is challenging. We ...aimed to identify genes characterized with low expression variability among human cancer and normal cell lines. For this purpose, we investigated the expression of 12 candidate reference genes in 13 widely used human cancer cell lines (HeLa, MCF-7, A-549, K-562, HL-60(TB), HT-29, MDA-MB-231, HCT 116, U-937, SH-SY5Y, U-251MG, MOLT-4 and RPMI-8226) and, in addition, 7 normal cell lines (HEK293, MRC-5, HUVEC/TERT2, HMEC, HFF-1, HUES 9, XCL-1). In our set of genes, we included SNW1 and CNOT4 as novel candidate reference genes based on the RNA HPA cell line gene data from The Human Protein Atlas. HNRNPL and PCBP1 were also included along with the "classical" reference genes ACTB, GAPDH, IPO8, PPIA, PUM1, RPL30, TBP and UBC. Results were evaluated using GeNorm, NormFiner, BestKeeper and the Comparative ΔCt methods. In conclusion, we propose IPO8, PUM1, HNRNPL, SNW1 and CNOT4 as stable reference genes for comparing gene expression between different cell lines. CNOT4 was also the most stable gene upon serum starvation.
In human cells two dUTPase isoforms have been described: one nuclear (DUT-N) and one mitochondrial (DUT-M), with cognate localization signals. In contrast, here we identified two additional isoforms; ...DUT-3 without any localization signal and DUT-4 with the same nuclear localization signal as DUT-N. Based on an RT-qPCR method for simultaneous isoform-specific quantification we analysed the relative expression patterns in 20 human cell lines of highly different origins. We found that the DUT-N isoform is expressed by far at the highest level, followed by the DUT-M and the DUT-3 isoform. A strong correlation between expression levels of DUT-M and DUT-3 suggests that these two isoforms may share the same promoter. We analysed the effect of serum starvation on the expression of dUTPase isoforms compared to non-treated cells and found that the mRNA levels of DUT-N decreased in A-549 and MDA-MB-231 cells, but not in HeLa cells. Surprisingly, upon serum starvation DUT-M and DUT-3 showed a significant increase in the expression, while the expression level of the DUT-4 isoform did not show any changes. Taken together our results indicate that the cellular dUTPase supply may also be provided in the cytoplasm and starvation stress induced expression changes are cell line dependent.
The enzyme dUTPase has an essential role in maintaining genomic integrity. In mouse, nuclear and mitochondrial isoforms of the enzyme have been described. Here we present the isoform-specific mRNA ...expression levels in different murine organs during development using RT-qPCR. In this study, we analyzed organs of 14.5-day embryos and of postnatal 2-, 4-, 10-week- and 13-month-old mice. We demonstrate organ-, sex- and developmental stage-specific differences in the mRNA expression levels of both isoforms. We found high mRNA expression level of the nuclear isoform in the embryo brain, and the expression level remained relatively high in the adult brain as well. This was surprising, since dUTPase is known to play an important role in proliferating cells, and mass production of neural cells is completed by adulthood. Thus, we investigated the pattern of the dUTPase protein expression specifically in the adult brain with immunostaining and found that dUTPase is present in the germinative zones, the subventricular and the subgranular zones, where neurogenesis occurs and in the rostral migratory stream where neuroblasts migrate to the olfactory bulb. These novel findings suggest that dUTPase may have a role in cell differentiation and indicate that accurate dTTP biosynthesis can be vital, especially in neurogenesis.
The oscillation and collective behavior of convective flows is studied by a computational fluid dynamics approach. More specifically, the rising dynamics of heated fluid columns is simulated in ...gravitational field using a simplified 2D geometry. The numerical method uses the FEniCS package for solving the coupled Navier–Stokes and heat-diffusion equations. For the flow of a single heated fluid column, the effect of the inflow yield and the nozzle diameter is studied. In agreement with the experiments, for a constant nozzle diameter the oscillation frequency increases approximately linearly as a function of the the flow rate, while for a constant flow rate the frequency decreases as a power law with the increased nozzle diameter. For the collective behavior of two nearby flows, we find a counter-phase synchronization and a decreasing trend of the common oscillation frequency with the distance between the jets. These results are in agreement with the experiments, and our computational study also suggests that the phenomenon is present on largely different length-scales.
The emergence of newer SARS-CoV-2 variants of concern (VOCs) profoundly changed the ICU demography; this shift in the virus's genotype and its correlation to lethality in the ICUs is still not fully ...investigated. We aimed to survey ICU patients' clinical and laboratory parameters in correlation with SARS-CoV-2 variant genotypes to lethality. 503 COVID-19 ICU patients were included in our study beginning in January 2021 through November 2022 in Hungary. Furthermore, we implemented random forest (RF) as a potential predictor regarding SARS-CoV-2 lethality among 649 ICU patients in two ICU centers. Survival analysis and comparison of hypertension (HT), diabetes mellitus (DM), and vaccination effects were conducted. Logistic regression identified DM as a significant mortality risk factor (OR: 1.55, 95% CI 1.06-2.29, p = 0.025), while HT showed marginal significance. Additionally, vaccination demonstrated protection against mortality (p = 0.028). RF detected lethality with 81.42% accuracy (95% CI 73.01-88.11%, AUC: 91.6%), key predictors being PaO
/FiO
ratio, lymphocyte count, and chest Computed Tomography Severity Score (CTSS). Although a smaller number of patients require ICU treatment among Omicron cases, the likelihood of survival has not proportionately increased for those who are admitted to the ICU. In conclusion, our RF model supports more effective clinical decision-making among ICU COVID-19 patients.
Oscillation and collective behavior of diffusion flames is a fascinating phenomena. Considering candle bundles with different sizes in variable oxygen concentration, the flickering dynamics of the ...flames are experimentally and theoretically investigated. Trends for the flickering frequency as a function of the candle number in the bundle and oxygen concentration is revealed for various topologies of the candles packing. The collective behavior of the flames as a function of their separation distance is studied by measuring an appropriate synchronization order parameter and through the common oscillation frequency. In agreement with previous results we find a discontinuous phase transition between an in-phase synchronized state at small separation distance and a counter-phase synchronized state at larger separation distances. A previously used dynamical model is modified in order to accommodate our experimental findings.
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