A Resonant System for In Vitro Studies Emulating Wireless Power Transfer Exposure at 13.56 MHz

This paper presents the design of a resonant system for in vitro studies to emulate the exposure of a monolayer of cells to a wireless power transfer system operating at 13.56 MHz. The design procedure targets a system, which maximizes the specific absorption rate (SAR) uniformity on the plane where the layer is cultured, as well as SAR efficiency (defined as SAR over the input power), within the size constraints of a standard incubator. Three resonant wireless power transfer systems with different commonly used loop/coil geometries (cylindrical with circular and square cross-sections and annular) were compared with assess the configuration maximizing the considered design criteria. The system performance in terms of reflection and transmission coefficients, as well as generated E- and H-fields, was characterized numerically and experimentally inside the incubator. Moreover, SAR was computed at the monolayer level. The system equipped with cylindrical coils with square cross-sections led to a high electromagnetic field uniformity in in vitro biological samples. In particular, the uniformities in E and SAR at the layer level were within 7.9% and 5.5%, respectively. This was achieved with the variation in H below the usually considered ±5% limit. © 2020 Bioelectromagnetics Society     

Seed dispersal networks in tropical forest fragments: Area effects,remnant species,and interaction diversity

Seed dispersal interactions involve key ecological processes in tropical forests that help to maintain ecosystem functioning. Yet this functionality may be threatened by increasing habitat loss, defaunation, and fragmentation. However, generalist species, and their interactions, can benefit from the habitat change caused by human disturbance while more specialized interactions mostly disappear. Therefore, changes in the structure of the local, within fragment, networks can be expected. Here we investigated how the structure of seed dispersal networks changes along a gradient of increasing habitat fragmentation. We analyzed 16 bird seed dispersal assemblages from forest fragments of a biodiversity-rich ecosystem. We found significant species–, interaction–, and network–area relationships, yet the later was determined by the number of species remaining in each community. The number of frugivorous bird and plant species, their interactions, and the number of links per species decreases as area is lost in the fragmented landscape. In contrast, network nestedness has a negative relationship with fragment area, suggesting an increasing generalization of the network structure in the gradient of fragmentation. Network specialization was not significantly affected by area, indicating that some network properties may be invariant to disturbance. Still, the local extinction of partner species, paralleled by a loss of interactions and specialist–specialist bird–plant seed dispersal associations, suggests the functional homogenization of the system as area is lost. Our study provides empirical evidence for network–area relationships driven by the presence/absence of remnant species and the interactions they perform.     

Normal hematopoiesis and hematologic malignancies: Role of canonical Wnt signaling pathway and stromal microenvironment

Wnts are a family of evolutionary-conserved secreted signaling molecules critically involved in a variety of developmental processes and in cell fate determination. A growing body of evidence suggests that Wnt signaling plays a crucial role in the influence of bone marrow stromal microenvironment on the balance between hematopoietic stem cell self-renewal and differentiation. Emerging clinical and experimental evidence also indicates Wnt signaling involvement in the disruption of the latter balance in hematologic malignancies, where the stromal microenvironment favors the homing of cancer cells to the bone marrow, as well as leukemia stem cell development and chemoresistance. In the present review, we summarize and discuss the role of the canonical Wnt signaling pathway in normal hematopoiesis and hematologic malignancies, with regard to recent findings on the stromal microenvironment involvement in these process and diseases.     

Biochemical modifications of gliadins induced by microbial transglutaminase on wheat flour

Background

Celiac disease (CD) is an immune-mediated disorder caused by the ingestion of wheat gluten. A lifelong, gluten-free diet is required to normalize the intestinal mucosa. We previously found that transamidation by microbial transglutaminase (mTGase) suppressed the gliadin-specific immune response in intestinal T-cell lines from CD patients and in models of gluten sensitivity.

Methods

SDS-PAGE, Western blot, ELISA, tissue transglutaminase (tTGase) assay and nano-HPLC–ESI-MS/MS experiments were used to analyze prolamins isolated from treated wheat flour.

Results

Gliadin and glutenin yields decreased to 7.6 ± 0.5% and 7.5 ± 0.3%, respectively, after a two-step transamidation reaction that produced a water-soluble protein fraction (spf). SDS-PAGE, Western blot and ELISA analyses confirmed the loss of immune cross-reactivity with anti-native gliadin antibodies in residual transamidated gliadins (K-gliadins) and spf as well as the occurrence of neo-epitopes. Nano-HPLC–ESI-MS/MS experiments identified some native and transamidated forms of celiacogenic peptides including p31–49 and confirmed that mTGase had similar stereo-specificity of tTGase. Those peptides resulted to be 100% and 57% modified in spf and K-gliadins, respectively. In particular, following transamidation p31–49 lost its ability to increase tTGase activity in Caco-2 cells. Finally, bread manufactured with transamidated flour had only minor changes in baking characteristics.

Conclusions

The two-step transamidation reaction modified the analyzed gliadin peptides, which are known to trigger CD, without influencing main technological properties.

General significance

Our data shed further light on a detoxification strategy alternative to the gluten free diet and may have important implications for the management of CD patients.     

GABAA Receptors of Cerebellar Granule Cells in Culture: Interaction with Benzodiazepines

GABA_A receptor mediated inhibition plays an important role in modulating the input/output dynamics of cerebellum. A characteristic of cerebellar GABA_A receptors is the presence in cerebellar granule cells of subunits such as α6 and δ which give insensitivity to classical benzodiazepines. In fact, cerebellar GABA_A receptors have generally been considered a poor model for testing drugs which potentially are active at the benzodiazepine site. In this overview we show how rat cerebellar granule cells in culture may be a useful model for studying new benzodiazepine site agonists. This is based on the pharmacological separation of diazepam-sensitive α1 β2/3 γ2 receptors from those which are diazepam-insensitive and contain the α6 subunit. This is achieved by utilizing furosemide/Zn²⁺ which block α6 containing and incomplete receptors.     

Retinal Cell Death Induced by TRPV1 Activation Involves NMDA Signaling and Upregulation of Nitric Oxide Synthases

The activation of the transient receptor potential vanilloid type 1 channel (TRPV1) has been correlated with oxidative and nitrosative stress and cell death in the nervous system. Our previous results indicate that TRPV1 activation in the adult retina can lead to constitutive and inducible nitric oxide synthase-dependent protein nitration and apoptosis. In this report, we have investigated the potential effects of TRPV1 channel activation on nitric oxide synthase (NOS) expression and function, and the putative participation of ionotropic glutamate receptors in retinal TRPV1-induced protein nitration, lipid peroxidation, and DNA fragmentation. Intravitreal injections of the classical TRPV1 agonist capsaicin up-regulated the protein expression of the inducible and endothelial NOS isoforms. Using 4,5-diaminofluorescein diacetate for nitric oxide (NO) imaging, we found that capsaicin also increased the production of NO in retinal blood vessels. Processes and perikarya of TRPV1-expressing neurons in the inner nuclear layer of the retina were found in the vicinity of nNOS-positive neurons, but those two proteins did not colocalize. Retinal explants exposed to capsaicin presented high protein nitration, lipid peroxidation, and cell death, which were observed in the inner nuclear and plexiform layers and in ganglion cells. This effect was partially blocked by AP-5, a NMDA glutamate receptor antagonist, but not by CNQX, an AMPA/kainate receptor antagonist. These data support a potential role for TRPV1 channels in physiopathological retinal processes mediated by NO, which at least in part involve glutamate release.     

Effective Binding Force Calculation in Dimeric Proteins

We apply the Blue Moon constrained Molecular Dynamics technique to study a particular case of molecular recognition, one of the main issues of modern molecular biology. We investigate the effects of mutation of interface residues on the binding strength of the dimeric protein superoxide dismutase from Photobacterium leiognathi. With our technique we produce a specific path describing the separation of the dimers and we calculate the effective mean force involved in the process. We apply the method to two mutants and compare the results with those obtained in an earlier calculation on the native enzyme. The method is sensitive to the mutations and allows us to establish a semi-quantitative hierarchy for the association strengths of the three enzymes.     

Blue Moon Approach to Rare Events

The "Blue Moon" ensemble is a computationally efficient molecular dynamics method to estimate the rate constants of rare activated events when the process can be described by a reaction coordinate ξ(r), a well-defined function in configuration space. By means of holonomic constraints a number of values of ξ(r) can be prescribed along the relevant path to identify the "bottleneck" region first and to sample an ensemble of starting conditions to generate activated trajectories. These MD trajectories sample phase space according to a biased configurational distribution. With a suitable re-weighting of averages from such ensemble of trajectories one can characterize completely rare events.     

DNA Topological Context Affects Access to Eukaryotic DNA Topoisomerase I

Abstract

We have analyzed the reactivity of a 217 base pair segment of the intrinsically curved Crithidia fasciculata kinetoplast DNA towards eukaryotic DNA topoisomerase I. The substrates were open [linear fragment and nicked circle] and closed minidomains [closed relaxed circle and circles with linking differences of ?1 and ?2], We interpreted the results with the aid of a model that was used to predict the structures of the topoisomers. The modelling shows that the ΔLk(?l) form is unusually compact because of the curvature in the DNA. To determine the role of sequence-directed curvature in both the experimental and modeling studies, controls were examined in which the curved Crithidia sequence was replaced by an uncurved sequence obtained from the plasmid pBR322.

Reactivity of the Crithidia DNA [as analyzed both by the cleavage and the topoisomerization reactions] markedly varied among the DNA forms: (i) the hierarchy of overall reactivity observed is: linear fragment > nicked circular, closed circular [ΔLk(O)], interwound [ΔLk(?2)] > bent interwound [ΔLk(?l)]; (ii) the intensity of several cleavage positions differs among DNA forms.

The results show that eukaryotic DNA topoisomerase I is very sensitive to the conformation of the substrates and that its reactivity is modulated by the variation of the compactness of the DNA molecule. The C. fasciculata sequence contains a highly curved segment that determines the conformation of the closed circle in a complex way.