Nebulette knockout mice have normal cardiac function, but show Z-line widening and up-regulation of cardiac stress markers

Aims: Nebulette is a 109 kDa modular protein localized in the sarcomeric Z-line of the heart. In vitro studies have suggested a role of nebulette in stabilizing the thin filament, and missense mutations in the nebulette gene were recently shown to be causative for dilated cardiomyopathy and endocardial fibroelastosis in human and mice. However, the role of nebulette in vivo has remained elusive. To provide insights into the function of nebulette in vivo, we generated and studied nebulette-deficient (nebl-/-) mice. Methods and results: Nebl-/- mice were generated by replacement of exon 1 by Cre under the control of the endogenous nebulette promoter, allowing for lineage analysis using the ROSA26 Cre reporter strain. This revealed specific expression of nebulette in the heart, consistent with in situ hybridization results. Nebl-/- mice exhibited normal cardiac function both under basal conditions and in response to transaortic constriction as assessed by echocardiography and haemodynamic analyses. Furthermore, histological, IF, and western blot analysis showed no cardiac abnormalities in nebl-/- mice up to 8 months of age. In contrast, transmission electron microscopy showed Z-line widening starting from 5 months of age, suggesting that nebulette is important for the integrity of the Z-line. Furthermore, up-regulation of cardiac stress responsive genes suggests the presence of chronic cardiac stress in nebl-/- mice. Conclusion: Nebulette is dispensable for normal cardiac function, although Z-line widening and up-regulation of cardiac stress markers were found in nebl-/- heart. These results suggest that the nebulette disease causing mutations have dominant gain-of-function effects

Bioinspired negatively charged calcium phosphate nanocarriers for cardiac delivery of MicroRNAs

Aim: To develop biocompatible and bioresorbable negatively charged calcium phosphate nanoparticles (CaP-NPs) as an innovative therapeutic system for the delivery of bioactive molecules to the heart. Materials & methods: CaP-NPs were synthesized via a straightforward one-pot biomineralization-inspired protocol employing citrate as a stabilizing agent and regulator of crystal growth. CaP-NPs were administered to cardiac cells in vitro and effects of treatments were assessed. CaP-NPs were administered in vivo and delivery of microRNAs was evaluated. Results: CaP-NPs efficiently internalized into cardiomyocytes without promoting toxicity or interfering with any functional properties. CaP-NPs successfully encapsulated synthetic microRNAs, which were efficiently delivered into cardiac cells in vitro and in vivo. Conclusion: CaP-NPs are a safe and efficient drug-delivery system for potential therapeutic treatments of polarized cells such as cardiomyocytes

Effect of PEF pre-treatment and extraction temperature on the recovery of carotenoids from tomato wastes

In this work, the influence of pulsed electric fields (PEF) pre-treatment and extraction temperature on the recovery of carotenoid compounds from tomato peels, achieved after peeling of steam blanched whole tomato fruits, was investigated. PEF pre-treatments were carried out at different field strengths (E = 0.5–5 kV/cm) and total specific energy input (WT = 0.5-20 kJ/kg). The cell disintegration index (ZP) was used to identify the optimal PEF processing conditions for the pre-treatment of tomato peels before the extraction with acetone at different temperatures (20-50°C). Extracts from untreated and PEF treated samples were analysed in terms of content and composition of carotenoids and antioxidant power. According to the ZP values, an energy input of 5 kJ/kg was enough for reaching the maximal permeabilization of tomato peel tissues at any investigated field strength. At this optimal energy input, only the extracts obtained from PEF pre-treated peels at 5 kV/cm showed a significantly higher total carotenoids content (47.3%) and antioxidant power (68%), as compared to the untreated samples. Regardless of the application of a PEF pre-treatment, the increase of the extraction temperature did not lead to further intensification in the recovery yield of total carotenoids in the extracts. Moreover, HPLC analyses revealed that lycopene was the main carotenoid extracted and no degradation/isomerization phenomena occurred. Results obtained in this work demonstrated that the PEF pretreatment of tomato processing wastes coupled with room or moderate extraction temperature could add new value to the tomato processing chain, improving economic performances and decreasing waste problems

Pulsed electric fields-assisted extraction of valuable compounds from Arthrospira platensis : effect of pulse polarity and mild heating

The present study aimed to investigate the effect of the main pulsed electric field (PEF) process parameters on the cell damages of A. platensis microalgae and the extractability of valuable compounds [water-soluble proteins (WSP), C-phycocyanin (C-PC), and carbohydrates (CH)]. Aqueous microalgae suspensions (2%, w/w) were PEF-treated at variable field strength (E = 10, 20, 30 kV/cm), total specific energy (WT = 20, 60, 100 kJ/kgsusp), and inlet temperature (25, 35, 45°C), with either monopolar or bipolar square wave pulses (5 μs of width, delay time between pulses of opposite polarities = 1, 5, 10, 20 μs), prior to extraction with water at room temperature (25°C) for up to 3 h. High-pressure homogenization (HPH) treatment (P = 150 MPa, 3 passes) was used to achieve complete cell disruption to quantify the total extractable content of target intracellular compounds. Scanning electron microscopy (SEM) and optical microscopy analyses clearly showed that PEF merely electroporated the membranes of algae cell, without damaging the cell structure and forming cell debris. The application of PEF treatment (monopolar pulses, 20 kV/cm and 100 kJ/kgsusp) at room temperature significantly enhanced the extraction yield of WSP [17.4% dry weight (DW)], CH (10.1% DW), and C-PC (2.1% DW), in comparison with the untreated samples. Bipolar pulses appeared less effective than monopolar pulses and led to extraction yields dependent on the delay time. Additionally, regardless of pulse polarity, a clear synergistic effect of the combined PEF (20 kV/cm and 100 kJ/kgsusp)-temperature (35°C) treatment was detected, which enabled the extraction of up to 37.4% (w/w) of total WSP, 73.8% of total CH, and 73.7% of total C-PC. Remarkably, the PEF treatment enabled to obtain C-phycocyanin extract with higher purity than that obtained using HPH treatment. The results obtained in this work suggest that the application of PEF combined with mild heating could represent a suitable approach for the efficient recovery of water-soluble compounds microalgal biomass

Changes of structural and techno-functional properties of high hydrostatic pressure (HHP) treated whey protein isolate over refrigerated storage

This work assessed the effect of storage time on the structure and functionality of HHP-treated whey protein isolate (WPI). Different pressures (100–600 MPa) and treatment times (15–30 min) were applied to aqueous WPI dispersions (5% w/v). The induced degree of unfolding was evaluated to select optimal HHP treatment conditions of WPI before storage at 4 °C. Conformational and techno-functional properties of untreated and optimally HHP-treated WPI samples were determined by UV–Vis and IR spectroscopy, foaming capacity, and interfacial tension measurements, respectively. Further tests of HHP-assisted hydrolysis of WPI were performed by α-chymotrypsin, bromelain, or their mixture (1:1 w/w), with the degree of hydrolysis (DH%) and electrophoretic patterns analyzed. The maximum unfolding degree was detected after a treatment of 400 MPa and 15 min and, at these processing conditions, no aggregation occurred. However, the structural changes achieved upon HHP were gradually lost during storage through a first-order refolding process (kREF. = 0.031 h−1), with restoring of native functionality. Hydrolysis performances of selected enzymes towards WPI were significantly promoted by high-pressure. Interestingly, a clear synergistic effect of α-chymotrypsin and bromelain combination on the WPI hydrolysis yield was detected, which resulted in the highest protein rupture (DH = 17%)

Ohmic heating for polyphenol extraction from grape berries: An innovative prefermentary process

High energy consumption is often required to increase the extraction of phenolic compounds from grapes during alcoholic fermentation. Processes such as thermovinification require significant temperature changes over a long period of time to ensure the diffusion of phenolic compounds from the grape skin layer to the must. In this study, the capability of the ohmic heating (OH) process (E = 55 V/cm, t = 60-90 s, T = 72 °C) to improve the extractability of valuable intracellular compounds from grape skins of Aglianico and Barbera grape matrices before the alcoholic fermentation stepAs similarly reported by Donsì et al. (2010), any tissue damage to grape skins occurring after the application of either conventional or ohmic heating was not found to influence the rate of fermentation. was investigated and compared with both untreated and conventional thermally (CH) treated (T = 72 °C, t = 90 s) samples. Total phenolics and antioxidant capacity were monitored during fermentation (10 days). In comparison to the conventional thermal treatment, the results showed that the phenolic compound content of musts was twice as high immediately after OH treatment. This process could drastically improve the classic prefermentary maceration (thermovinifications, cold macerations, etc.) time. In finished wines produced from the treated musts, the total polyphenolic content of OH wines was up to 17 % higher than that of CH wines, and 30 % higher than that of untreated wines. No differences in concentrations of total tannins and anthocyanins were observed between conventional and ohmic heated musts. However, an increase of 30 to 200 % for some aromatic esters was observed in wines from ohmic heated musts. Overall, the outcomes of this work proved that, in addition to the thermal effect, the moderate electric field (MEF) applied during ohmic heating has the potential to induce an instantaneous release of polyphenolic compounds due to the electroporation phenomenon of cell membranes, thus saving energy and reducing processing time

Improving the extraction yield of juice and bioactive compounds from sweet cherries and their by-products by pulsed electric fields

The influence of pulsed electric fields (PEF) pre-treatment of sweet cherry (Prunus Avium) fruits of "Duroni Nero" variety both on the extraction yield and antioxidant properties of juice obtained by pressing, and on the subsequent extraction with solvent of bioactive compounds from cherry by-products (press-cake) was investigated. PEF pre-treatments were performed at constant specific energy input (Wt = 10 kJ/kg) and different electric field strength (E= 0.5-3 kV/cm) before applying a pressure of 1.64 bar for 5 min. PEF-assisted pressing (E = 1 kV/cm) led to a significant increase of juice yield (+40%), anthocyanins (+80%), and antioxidant power (+27%) with respect to untreated samples. However, PEF treatment intensity higher than 1 kV/cm did not significantly improved the quantitative and qualitative characteristics of juice. Compared to the untreated samples, the extracts from electroporated press-cakes achieved after PEF-assisted pressing of cherry fruits at 0.5 kV/cm showed the highest increases in anthocyanins (+38%) and antioxidant activity (+21%). No evidence of selective release or degradation of individual anthocyanins in juice and press cake extracts due to PEF application was observed. Overall, the results of this investigation demonstrated the potential of PEF as a mild technology to improve the efficiency of industrial processing of cherry fruits

L’app Immuni quale requisito per lo svolgimento di attività a rischio (di diffusione del contagio)? Una proposta per incentivare l’uso delle ICT nella lotta alla pandemia

Alla luce delle numerose misure restrittive adottate in Italia, come in molti altri paesi, per contenere l’epidemia di SARS-CoV-2, gli autori esaminano i termini entro i quali l’uso dell’App Immuni potrebbe essere qualificato come condizione legittimante per lo svolgimento di attività a rischio di contagio al fine di dare maggiore effettività alle misure di contenimento dell’epidemia e, quindi, meglio calibrare le limitazioni alle libertà personali. In questa prospettiva, l’attenzione degli autori si concentra sull’analisi del funzionamento dell’app Immuni, specie in un’ottica di protezione dei dati personali ed alla luce dell’analisi delle norme europee in materia, analizzate anche attraverso il prisma del principio di proporzionalità.The Immuni app as a requirement for carrying out activities at risk (of widespreading the contagion)? A proposal to encourage the use of ICT in the fight against the pandemic. In light of the numerous restrictive measures adopted in Italy, as in many other countries, to contain the SARS-CoV-2 epidemic, the authors examine the terms with which the use of the “App Immuni” could be qualified as a legitimate condition for carrying out activities at risk of infection. Digital contact tracing could give greater effectiveness to the measures taken to contain the epidemic and, therefore, lead to a better calibration of the limitations to personal freedoms. In this perspective, the authors’ attention is focused on analyzing the functioning of the app Immuni, especially in view of protecting personal data and in light of the analysis of the relevant European standards, also analyzed through the prism of the principle of proportionality

A screen-printed electrode modified with gold nanoparticles/cellulose nanocrystals for electrochemical detection of 4,4′-methylene diphenyl diamine

Developing simple, cost-effective, easy-to-use, and reliable analytical devices if of utmost importance for the food industry for rapid in-line checks of their products that must comply with the provisions set by the current legislation. The purpose of this study was to develop a new electrochemical sensor for the food packaging sector. More specifically, we propose a screen-printed electrode (SPE) modified with cellulose nanocrystals (CNCs) and gold nanoparticles (AuNPs) for the quantification of 4,4′-methylene diphenyl diamine (MDA), which is one of the most important PAAs that can transfer from food packaging materials into food stuffs. The electrochemical performance of the proposed sensor (AuNPs/CNCs/SPE) in the presence of 4,4′-MDA was evaluated using cyclic voltammetry (CV). The modified AuNPs/CNCs/SPE showed the highest sensitivity for 4,4′-MDA detection, with a peak current of 9.81 μA compared with 7.08 μA for the bare SPE. The highest sensitivity for 4,4′-MDA oxidation was observed at pH = 7, whereas the detection limit was found at 57 nM and the current response of 4,4′-MDA rose linearly as its concentration increased from 0.12 μM to 100 μM. Experiments using real packaging materials revealed that employing nanoparticles dramatically improved both the sensitivity and the selectivity of the sensor, which can be thus considered as a new analytical tool for quick, simple, and accurate measurement of 4,4′-MDA during converting operations

Rheological and tribological characterization of different commercial hazelnut-based spreads

Five commercial hazelnut/cocoa spreads with different compositions were tested by rheology/tribology. The impact of each formulation on the structural/lubricant performances was investigated. Rotational/oscillatory rheology was chosen to assess material behavior during flow. Viscosity variation as a function of temperature and chamber geometry was evaluated. Oscillatory mode tests were carried out to obtain information on product viscoelasticity. Tribological analysis was performed at different temperatures aiming at simulating the chewing/swallowing process. All samples were categorized as pseudo-plastic and viscoelastic materials, with the elastic component prevailing over the viscous one. Major differences were detected in terms of consistency index, depending on the total lipid content. Temperature increase enhanced spread fluidity with a decreasing viscosity according to the Arrhenius model (R2 > 0.942) and greater values of activation energy reflecting higher sensitivity to microstructural changes. An inverse relationship between Casson viscosity (Formula presented.) and sugar/fat ratio highlighted additional correlations between structural parameters and spread formulation. Tribological measurements at 25°C highlighted that, at the initial eating stage, the friction factor (0.112–0.262 at sliding velocity of 8 × 10−6 m/s) was strongly affected by either the amounts of solid fat or hazelnut percentage. Tribological data corroborated the theory for which tribology and rheology cover different domains