Label-Free Study of the Global Cell Behavior during Exposure to Environmental Radiofrequency Fields in the Presence or Absence of Pro-Apoptotic or Pro-Autophagic Treatments

It remains controversial whether exposure to environmental radiofrequency signals (RF) impacts cell status or response to cellular stress such as apoptosis or autophagy. We used two label-free techniques, cellular impedancemetry and Digital Holographic Microscopy (DHM), to assess the overall cellular response during RF exposure alone, or during co-exposure to RF and chemical treatments known to induce either apoptosis or autophagy. Two human cell lines (SH-SY5Y and HCT116) and two cultures of primary rat cortex cells (astrocytes and co-culture of neurons and glial cells) were exposed to RF using an 1800 MHz carrier wave modulated with various environmental signals (GSM: Global System for Mobile Communications, 2G signal), UMTS (Universal Mobile Telecommunications System, 3G signal), LTE (Long-Term Evolution, 4G signal, and Wi-Fi) or unmodulated RF (continuous wave, CW). The specific absorption rates (S.A.R.) used were 1.5 and 6 W/kg during DHM experiments and ranged from 5 to 24 W/kg during the recording of cellular impedance. Cells were continuously exposed for three to five consecutive days while the temporal phenotypic signature of cells behavior was recorded at constant temperature. Statistical analysis of the results does not indicate that RF-EMF exposure impacted the global behavior of healthy, apoptotic, or autophagic cells, even at S.A.R. levels higher than the guidelines, provided that the temperature was kept constant.     

Metabolic and functional consequences of successive no-flow and sustained low-flow ischaemia; a 31P MRS study in rat hearts

Sawmill workers in British Columbia (B.C.), Canada, have been exposed to chlorophenate fungicides which are known to be contaminated with polychlorinated dibenzo-p-dioxins (PCDDs) and polychlorinated dibenzofurans (PCDFs). Due to concern about the potential of these workers to have significant body burdens of PCDD/Fs, and the absence of measurements in these workers, a single-compartment pharmacokinetic model was developed to estimate the concentration of PCDD/Fs in the fat tissue of the sawmill workers. Data from a large cohort of B.C. sawmill workers and literature-based data on chlorophenate exposures and PCDD/F concentrations in chlorophenates were used in Monte Carlo simulations to predict a PCDD/F body burden distribution. The median concentrations of HxCDF and HpCDF predicted using the model for the B.C. sawmill worker population exceeded the range measured in unexposed populations. PeCDF and OCDF concentrations exceeded the range measured in unexposed populations at the 70th percentile of the model-predicted distribution, and PeCDD at the 90th percentile. The primary limitation of the model was the scarcity of input data about actual dermal and inhalation exposures to chlorophenates.     

Kinetic and stoichiometric characterisation of streptavidin-binding aptamers

Aptamers are oligonucleotide ligands that are selected for high-affinity binding to molecular targets. Only limited knowledge relating to relations between structural and kinetic properties that define aptamer-target interactions is available. To this end, streptavidin-binding aptamers were isolated and characterised by distinct analytical techniques. Binding kinetics of five broadly similar aptamers were determined by surface plasmon resonance (SPR); affinities ranged from 35-375 nM with large differences in association and dissociation rates. Native mass spectrometry showed that streptavidin can accommodate up to two aptamer units. In a 3D model of one aptamer, conserved regions are exposed, strongly suggesting that they directly interact with the biotin-binding pockets of streptavidin. Mutational studies confirmed both conserved regions to be crucial for binding. An important result is the observation that the most abundant aptamer in our selections is not the tightest binder, emphasising the importance of having insight into the kinetics of complex formation. To find the tightest binder it might be better to perform fewer selection rounds and to focus on post-selection characterisation, through the use of complementary approaches as described in this study.     

Preclinical Assessment of the Combination of PSMA-Targeting Radionuclide Therapy with PARP Inhibitors for Prostate Cancer Treatment

Prostate specific membrane antigen targeted radionuclide therapy (PSMA-TRT) is a promising novel treatment for prostate cancer (PCa) patients. However, PSMA-TRT cannot be used for curative intent yet, thus additional research on how to improve the therapeutic efficacy is warranted. A potential way of achieving this, is combining TRT with poly ADP-ribosylation inhibitors (PARPi), which has shown promising results for TRT of neuroendocrine tumor cells. Currently, several clinical trials have been initiated for this combination for PCa, however so far, no evidence of synergism is available for PCa. Therefore, we evaluated the combination of PSMA-TRT with three classes of PARPi in preclinical PCa models. In vitro viability and survival assays were performed using PSMA-expressing PCa cell lines PC3-PIP and LNCaP to assess the effect of increasing concentrations of PARPi veliparib, olaparib or talazoparib in combination with PSMA-TRT compared to single PARPi treatment. Next, DNA damage analyses were performed by quantifying the number of DNA breaks by immunofluorescent stainings. Lastly, the potential of the combination treatments was studied in vivo in mice bearing PC3-PIP xenografts. Our results show that combining PSMA-TRT with PARPi did not synergistically affect the in vitro clonogenic survival or cell viability. DNA-damage analysis revealed only a significant increase in DNA breaks when combining PSMA-TRT with veliparib and not in the other combination treatments. Moreover, PSMA-TRT with PARPi treatment did not improve tumor control compared to PSMA-TRT monotherapy. Overall, the data presented do not support the assumption that combining PSMA-TRT with PARPi leads to a synergistic antitumor effect in PCa. These results underline that extensive preclinical research using various PCa models is imperative to validate the applicability of the combination strategy for PCa, as it is for other cancer types.     

Microcircuitry and function of the inferior olive

The inferior olive, which provides the climbing fibers to Purkinje cells in the cerebellar cortex, has been implicated in various functions, such as learning and timing of movements, and comparing intended with achieved movements. For example, climbing-fiber activity could transmit error signals during eye-blink conditioning or adaptation of the vestibulo-ocular reflex, or it could carry motor command signals beating on the rhythm of the oscillating and synchronous firing of ensembles of olivary neurons, or both. In this review, we approach the controversial issue of olivocerebellar function from the perspective of the unique organization of the microcircuitry of the olivary neuropil. The characteristic glomeruli are formed by a core of long dendritic or axonal spines, each of which is innervated by both an inhibitory terminal derived from the hindbrain and an excitatory terminal derived from either an ascending or descending input. The dendritic spines, which originate from dendrites with varicosities carrying dendritic lamellar bodies, are coupled by gap junctions. By drawing a comparison with a computational model by Segev and Rall,which might be applicable to the typical olivary spine with its unique morphological features and combined excitatory and inhibitory input, we propose that the microcircuitry of the inferior olive is capable of functioning both in motor learning and motor timing, but does not directly compare intended with achieved movements.     

Human factors evaluations of Free Flight Issues solved and issues remaining

The Dutch National Aerospace Laboratory (NLR) has conducted extensive human-in-the-loop simulation experiments in NLR's Research Flight Simulator (RFS), focussed on human factors evaluation of Free Flight. Eight years of research, in co-operation with partners in the United States and Europe, has shown that Free Flight has the potential to increase airspace capacity by at least a factor of 3. Expected traffic loads and conflict rates for the year 2020 appear to be no major problem for professional airline crews participating in flight simulation experiments. Flight efficiency is significantly improved by user-preferred routings, including cruise climbs, while pilot workload is only slightly increased compared to today's reference. Detailed results from three projects and six human-in-the-loop experiments in NLR's Research Flight Simulator are reported. The main focus of these results is on human factors issues and particularly workload, measured both subjectively and objectively. An extensive discussion is included on many human factors issues resolved during the experiments, but also open issues are identified. An intent-based Conflict Detection and Resolution (CD&R) system provides "benefits" in terms of reduced pilot workload, but also "costs" in terms of complexity, need for priority rules, potential compatibility problems between different brands of Flight Management Systems and large bandwidth. Moreover, the intent-based system is not effective at solving multi-aircraft conflicts. A state-based CD&R system also provides "benefits" and "costs". Benefits compared to the full intent-based system are simplicity, low bandwidth requirements, easy to retrofit (no requirements to change avionics infrastructure) and the ability to solve multi-aircraft conflicts in parallel. The "costs" involve a somewhat higher pilot workload in similar circumstances, the smaller look-ahead time which results in less efficient resolution manoeuvres and the sometimes false/nuisance alerts due to missing intent information. The optimal CD&R system (in terms of costs versus benefits) has been suggested to be state-based CD&R with the addition of intended or target flight level. This combination of state-based CD&R with a limited amount of intent provides "the best of both worlds". Studying this CD&R system is still an open issue.     

Adenovirus type 40 virions contain two distinct fibers

Human subgroup F adenoviruses, types 40 and 41, have two genes that could specify different fiber proteins, a long fiber with 21 or 22 pseudorepeat motifs in the shaft, and a short fiber with 12 motifs. We show that for adenovirus type 40, both proteins are expressed from separate late regions of the genome. They are incorporated into the virion with only one fiber per penton base, unlike avian adenoviruses which have two fibers extending from each penton. Comparison of known adenovirus fiber sequences suggests that the two fibers in adenovirus 40 and 41 arose through an external recombination event rather than having evolved from each other directly. Two alternative fibers on the virion may imply the existence of two distinct receptors.