Stages of neuronal network formation

Graph theoretical approaches have become a powerful tool for investigating the architecture and dynamics of complex networks. The topology of network graphs revealed small-world properties for very different real systems among these neuronal networks. In this study, we observed the early development of mouse retinal ganglion cell (RGC) networks in vitro using timelapse video microscopy. By means of a time-resolved graph theoretical analysis of the connectivity, shortest path length and the edge length, we were able to discover the different stages during the network formation. Starting from single cells, at the first stage neurons connected to each other ending up in a network with maximum complexity. In the further course, we observed a simplification of the network which manifested in a change of relevant network parameters such as the minimization of the path length. Moreover, we found that RGC networks self-organized as small-world networks at both stages; however, the optimization occurred only in the second stage

Ligand Mediated Sequestering of Integrins in Raft-Mimicking Lipid Mixtures: The Role of Bilayer Asymmetry and Cholesterol Content

poster abstractLipid microdomains play an important functional role in plasma membranes. However, the small size and transient nature of lipid/membrane heterogeneities in the plasma membrane make characterization of microdomains and microdomain-related membrane processes quite challenging. To address this issue, we recently introduced a powerful model membrane system that allows the investigation of membrane protein sequestering and oligomerization in raft-mimicking lipid mixtures using combined confocal fluorescence spectroscopy, photon counting histogram (PCH), and epifluorescence microscopy. Our experiments on bilayer-spanning domains showed that αvβ3 and α5β1 integrins predominantly exist as monomers and sequester preferentially to the liquid-disordered (ld) phase in the absence of ligands. Notably, addition of vitronectin (αvβ3) and fibronectin (α5β1) caused substantial translocations of integrins into the liquid-ordered (lo) phase without altering receptor oligomerization state. Here we expand our previous studies and report on the sequestering and oligomerization state of αvβ3 and α5β1 in asymmetric bilayer compositions containing coexisting lo and ld phases located exclusively in the top leaflet of the bilayer (bottom leaflet shows only ld phase). Remarkably, in such a membrane environment, both integrins show a higher affinity for the top leaflet-restricted lo domains in the absence of their respective ligands. A slight change in the integrin sequestration was observed after addition of their respective ligands. We also present experimental findings, which show that cholesterol content has a substantial influence on integrin sequestering and oligomerization in raft-mimicking lipid mixtures. The described experimental results highlight the potential importance of membrane asymmetry and lipid composition in the sequestering of membrane proteins in biological membranes

Conjugates of Superoxide Dismutase 1 with Amphiphilic Poly(2-oxazoline) Block Copolymers for Enhanced Brain Delivery: Synthesis, Characterization and Evaluation in Vitro and in Vivo

Superoxide dismutase 1 (SOD1) efficiently catalyzes dismutation of superoxide but its poor delivery to the target sites in the body, such as brain, hinders its use as a therapeutic agent for superoxide-associated disorders. Here to enhance the delivery of SOD1 across the blood brain barrier (BBB) and in neurons the enzyme was conjugated with poly(2-oxazoline) (POx) block copolymers, P(MeOx-b-BuOx) or P(EtOx-b-BuOx), comprised of 1) hydrophilic 2-methyl-2-oxazoline (MeOx) or 2-ethyl-2-oxazoline (EtOx) and 2) hydrophobic 2-butyl-2-oxazoline (BuOx) repeating units. The conjugates contained from 2 to 3 POx chains joining the protein amino groups via cleavable -(ss)- or non-cleavable –(cc)- linkers at the BuOx block terminus. They retained 30% to 50% of initial SOD1 activity, were conformationally and thermally stable and assembled in 8 or 20 nm aggregates in aqueous solution. They had little if any toxicity to CATH.a neurons and displayed enhanced uptake in these neurons as compared to native or PEGylated SOD1. Of the two conjugates, SOD1-(cc)-P(MeOx-b-BuOx) and SOD1-(cc)-P(EtOx-b-BuOx) compared, the latter was entering cells 4 to 7 times faster and at 6 h colocalized predominantly with endoplasmic reticulum (41 ± 3%) and mitochondria (21 ± 2%). Colocalization with endocytosis markers and pathway inhibition assays suggested that it was internalized through lipid raft/caveolae, also employed by the P(EtOx-b-BuOx) copolymer. The SOD activity in cell lysates and ability to attenuate angiotensin II (Ang II)-induced superoxide in live cells were increased for this conjugate compared to SOD1 and PEG-SOD1. Studies in mice showed that SOD1-POx had ca. 1.75 times longer half-life in blood than native SOD1 (28.4 vs 15.9 min) and after i.v. administration penetrated the BBB significantly faster than albumin to accumulate in brain parenchyma. The conjugate maintained high stability both in serum and in brain (77% vs. 84% at 1 h post injection). Its amount taken up by the brain reached a maximum value of 0.08%ID/g (percent of the injected dose taken up per gram of brain) 4 h post injection. The entry of SOD1-(cc)-P(EtOx-b-BuOx) to the brain was mediated by a non-saturable mechanism. Altogether, SOD1-POx conjugates are promising candidates as macromolecular antioxidant therapies for superoxide-associated diseases such as Ang II induced neuro-cardiovascular diseases

No Conclusive Evidence for Transits of Proxima b in MOST photometry

The analysis of Proxima Centauri's radial velocities recently led Anglada-Escud\'e et al. (2016) to claim the presence of a low mass planet orbiting the Sun's nearest star once every 11.2 days. Although the a-priori probability that Proxima b transits its parent star is just 1.5%, the potential impact of such a discovery would be considerable. Independent of recent radial velocity efforts, we observed Proxima Centauri for 12.5 days in 2014 and 31 days in 2015 with the MOST space telescope. We report here that we cannot make a compelling case that Proxima b transits in our precise photometric time series. Imposing an informative prior on the period and phase, we do detect a candidate signal with the expected depth. However, perturbing the phase prior across 100 evenly spaced intervals reveals one strong false-positive and one weaker instance. We estimate a false-positive rate of at least a few percent and a much higher false-negative rate of 20-40%, likely caused by the very high flare rate of Proxima Centauri. Comparing our candidate signal to HATSouth ground-based photometry reveals that the signal is somewhat, but not conclusively, disfavored (1-2 sigmas) leading us to argue that the signal is most likely spurious. We expect that infrared photometric follow-up could more conclusively test the existence of this candidate signal, owing to the suppression of flare activity and the impressive infrared brightness of the parent star.Comment: Accepted to ApJ. Posterior samples, MOST photometry and HATSouth photometry are all available at https://github.com/CoolWorlds/Proxim

A high capacity polymeric micelle of paclitaxel: Implication of high dose drug therapy to safety and in vivo anti-cancer activity

The poor solubility of paclitaxel (PTX), the commercially most successful anticancer drug, has long been hampering the development of suitable formulations. Here, we present translational evaluation of a nanoformulation of PTX, which is characterized by a facile preparation, extraordinary high drug loading of 50 % wt. and PTX solubility of up to 45 g/L, excellent shelf stability and controllable, sub-100 nm size. We observe favorable in vitro and in vivo safety profiles and a higher maximum tolerated dose compared to clinically approved formulations. Pharmacokinetic analysis reveals that the higher dose administered leads to a higher exposure of the tumor to PTX. As a result, we observed improved therapeutic outcome in orthotopic tumor models including particularly faithful and aggressive “T11” mouse claudin-low breast cancer orthotopic, syngeneic transplants. The promising preclinical data on the presented PTX nanoformulation showcase the need to investigate new excipients and is a robust basis to translate into clinical trials

Response to letter to the editor by Jan Kühnisch

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Parasites dominate hyperdiverse soil protist communities in Neotropical rainforests

High animal and plant richness in tropical rainforest communities has long intrigued naturalists. It is unknown if similar hyperdiversity patterns are reflected at the microbial scale with unicellular eukaryotes (protists). Here we show, using environmental metabarcoding of soil samples and a phylogeny-aware cleaning step, that protist communities in Neotropical rainforests are hyperdiverse and dominated by the parasitic Apicomplexa, which infect arthropods and other animals. These host-specific parasites potentially contribute to the high animal diversity in the forests by reducing population growth in a density-dependent manner. By contrast, too few operational taxonomic units (OTUs) of Oomycota were found to broadly drive high tropical tree diversity in a host-specific manner under the Janzen-Connell model. Extremely high OTU diversity and high heterogeneity between samples within the same forests suggest that protists, not arthropods, are the most diverse eukaryotes in tropical rainforests. Our data show that protists play a large role in tropical terrestrial ecosystems long viewed as being dominated by macroorganisms

Common and low Frequency variants in MERTK are independently associated with multiple sclerosis susceptibility with discordant association dependent upon HLA-DRB1*15:01 status

Multiple Sclerosis (MS) is a chronic inflammatory demyelinating disease of the central nervous system. The risk of developing MS is strongly influenced by genetic predisposition, and over 100 loci have been established as associated with susceptibility. However, the biologically relevant variants underlying disease risk have not been defined for the vast majority of these loci, limiting the power of these genetic studies to define new avenues of research for the development of MS therapeutics. It is therefore crucial that candidate MS susceptibility loci are carefully investigated to identify the biological mechanism linking genetic polymorphism at a given gene to the increased chance of developing MS. MERTK has been established as an MS susceptibility gene and is part of a family of receptor tyrosine kinases known to be involved in the pathogenesis of demyelinating disease. In this study we have refined the association of MERTK with MS risk to independent signals from both common and low frequency variants. One of the associated variants was also found to be linked with increased expression of MERTK in monocytes and higher expression of MERTK was associated with either increased or decreased risk of developing MS, dependent upon HLA-DRB1*15:01 status. This discordant association potentially extended beyond MS susceptibility to alterations in disease course in established MS. This study provides clear evidence that distinct polymorphisms within MERTK are associated with MS susceptibility, one of which has the potential to alter MERTK transcription, which in turn can alter both susceptibility and disease course in MS patients

Fluoride bioavailability in saliva and plaque

<p>Abstract</p> <p>Background</p> <p>Different fluoride formulations may have different effects on caries prevention. It was the aim of this clinical study to assess the fluoride content, provided by NaF compared to amine fluoride, in saliva and plaque.</p> <p>Methods</p> <p>Eight trained volunteers brushed their teeth in the morning for 3 minutes with either NaF or amine fluoride, and saliva and 3-day-plaque-regrowth was collected at 5 time intervals during 6 hours after tooth brushing. The amount of collected saliva and plaque was measured, and the fluoride content was analysed using a fluoride sensitive electrode. All subjects repeated all study cycles 5 times, and 3 cycles per subject underwent statistical analysis using the Wilcoxon-Mann-Whitney test.</p> <p>Results</p> <p>Immediately after brushing the fluoride concentration in saliva increased rapidly and dropped to the baseline level after 360 minutes. No difference was found between NaF and amine fluoride. All plaque fluoride levels were elevated after 30 minutes until 120 minutes after tooth brushing, and decreasing after 360 minutes to baseline. According to the highly individual profile of fluoride in saliva and plaque, both levels of bioavailability correlated for the first 30 minutes, and the fluoride content of saliva and plaque was back to baseline after 6 hours.</p> <p>Conclusions</p> <p>Fluoride levels in saliva and plaque are interindividually highly variable. However, no significant difference in bioavailability between NaF and amine fluoride, in saliva, or in plaque was found.</p