High-throughput profiling of ion channel activity in primary human lymphocytes

We present a high-throughput method to quantify the functional activity of potassium (K (+)) ion channels in primary human lymphocytes. This method is rapid, automated, specific (here for the voltage-gated Kv1.3 ion channel), and capable of measuring, in parallel, the electrical currents of over 200 individual lymphocytes isolated from freshly drawn blood. The statistics afforded by high-throughput measurements allowed direct comparison of Kv1.3 activity in different subsets of lymphocytes, including CD4 (+) and CD8 (+) T cells, gammadelta T cells, and B cells. High-throughput measurements made it possible to quantify the heterogeneous, functional response of Kv1.3 ion channel activity upon stimulation of CD4 (+) and CD8 (+) T cells with mitogen. These experiments enabled elucidation of time-courses of functional Kv1.3 activity upon stimulation as well as studies of the effects of the concentration of mitogenic antibodies on Kv1.3 levels. The results presented here suggest that Kv1.3 ion channel activity can be used as a functional activation marker in T cells and that it correlates to cell size and levels of a surface antigen, CD25. Moreover, this work presents an enabling methodology that can be applied widely, allowing high-throughput screening of specific voltage-gated ion channels in a variety of primary cells.     

Hydrolysis of Alkyl Ester on Lipase/Silicalite-1 Catalyst

Pure silica zeolites have been prepared in powder and pellets form with different surface chemical properties in order to investigate how the binding forces in the lipase enzyme adsorption influence the final conformation of the immobilized enzyme and its catalytic activity. The catalytic activity of the adsorbed lipase have been tested in the alkyl esters hydrolysis reaction. The higher alkyl-esters conversion is observed for the lipase immobilized on the supports prepared in OH⁻ media compared to the fluoride media supports. The obtained results can be explained by the two different non-covalent interactions between the external surface of zeolites and the enzyme.     

High‐pressure high‐temperature extraction of phenolic compounds from grape skins

This study focused on the use of a non‐conventional extraction technology by employing high‐pressure high‐temperature stirred reactor to extract polyphenols from grape skins. Extraction time (15–330 min) and temperature (30–150 °C) were selected as independent variables, and their effects were studied. A preliminary kinetic study on polyphenols extraction revealed that the second‐order model fitted satisfactorily the experimental results (R² ≥ 0.9798). Total polyphenol yield and total flavonoid (TF) yield, as well as the antiradical power (ARP) of the extracts, were analysed. The use of high‐pressure high‐temperature technology resulted in obtaining extracts rich in polyphenols with high ARP. The highest total polyphenol (60.7 mg_GAE ) and TF (15.1 mg_CE ) yields were obtained at 150 °C for 270 min and 150 °C for 15 min, respectively. HPLC was employed to analyse phenolic compounds. Considerable quantities of single phenolic compounds were extracted. The highest yields of gallic acid, 5‐hydroxymethylfurfural, protocatechuic acid, catechin, vanillic acid, syringic acid, cumaric acid, trans‐resveratrol and quercetin (163.2, 20.0, 69.9, 420.0, 20.6, 603.0, 20.1, 42.4 and 117.1 mg per 100 g_DS, respectively) were found. ARP values were found between 8.45 and 52.17 μg_DPPH .     

Fibers from fruit by-products enhance probiotic viability and fatty acid profile and increase CLA content in yoghurts

This study evaluated the effect of the supplementation of total dietary fiber from apple, banana or passion fruit processing by-products on the post-acidification, total titratable acidity, bacteria counts and fatty acid profiles in skim milk yoghurts co-fermented by four different probiotics strains: Lactobacillus acidophilus L10 and Bifidobacterium animalis subsp. lactis BL04, HN019 and B94. Apple and banana fibers increased the probiotic viability during shelf-life. All the fibers were able to increase the short chain and polyunsaturated fatty acid contents of yoghurts compared to their respective controls. A synergistic effect between the type of fiber and the probiotic strain on the conjugated linoleic acid content was observed, and the amount of α-linolenic acid was increased by banana fiber. The results of this study demonstrate, for the first time, that fruit fibers can improve the fatty acid profile of probiotic yoghurts and point out the suitability of using fibers from fruit processing the by-products to develop new high value-added fermented dairy products.     

Recognition of organic solvents molecules by simultaneous detection using SAW oscillator sensors and optical fiber devices coated by Langmuir-Blodgett cadmium arachidate films

Surface acoustic waves (SAW) 433 and 315 MHz, two-port resonator-based oscillators coated with a Langmuir-Blodgett (LB) thin layer of chemosensitive cadmium arachidate (CdA) provide highly sensitive chemical acoustic sensors for detection and monitoring of organic vapors, at room temperature. LB CdA film-coated silica optical fibers (SOF) have been successfully fabricated and studied for organic solvents molecules sensing applications. The sensing performance of both types of acoustic and optical transducers has been compared for detecting six molecular species. Simultaneous measurements of frequency changes (delta f) and optoelectronic signal changes (deltaV) of the LB CdA film assembled onto SAW sensors and SOF devices have been realized for organic vapors recognition purposes. Six molecular species such as ethanol, methanol, isopropanol, ethylacetate, acetone, and toluene have been identified and recognized by a specific index (deltaf/deltaV), which can be considered a characteristic property of the chemosensitive material. The discrimination of the six molecular species examined also has been obtained by chemical patterns using a couple of specific index (deltaf433/deltaV; deltaf315/deltaV) measured by combining SAW 433 or 315 MHz oscillators and SOF sensing devices. Transient responses, calibration curves, intertransducer relationships, and chemical patterns are presented and discussed.     

Development of microbial and chemical MST tools to identify the origin of the faecal pollution in bathing and shellfish harvesting waters in France

The microbiological quality of coastal or river waters can be affected by faecal pollution from human or animal sources. An efficient MST (Microbial Source Tracking) toolbox consisting of several host-specific markers would therefore be valuable for identifying the origin of the faecal pollution in the environment and thus for effective resource management and remediation. In this multidisciplinary study, after having tested some MST markers on faecal samples, we compared a selection of 17 parameters corresponding to chemical (steroid ratios, caffeine, and synthetic compounds), bacterial (host-specific Bacteroidales, Lactobacillus amylovorus and Bifidobacterium adolescentis) and viral (genotypes I-IV of F-specific bacteriophages, FRNAPH) markers on environmental water samples (n = 33; wastewater, runoff and river waters) with variable Escherichia coli concentrations. Eleven microbial and chemical parameters were finally chosen for our MST toolbox, based on their specificity for particular pollution sources represented by our samples and their detection in river waters impacted by human or animal pollution; these were: the human-specific chemical compounds caffeine, TCEP (tri(2-chloroethyl)phosphate) and benzophenone; the ratios of sitostanol/coprostanol and coprostanol/(coprostanol+24-ethylcopstanol); real-time PCR (Polymerase Chain Reaction) human-specific (HF183 and B. adolescentis), pig-specific (Pig-2-Bac and L. amylovorus) and ruminant-specific (Rum-2-Bac) markers; and human FRNAPH genogroup II.     

Plasma Exosome Profile in ST-Elevation Myocardial Infarction Patients with and without Out-of-Hospital Cardiac Arrest

The identification of new biomarkers allowing an early and more accurate characterization of patients with ST-segment elevation myocardial infarction (STEMI) is still needed, and exosomes represent an attractive diagnostic tool in this context. However, the characterization of their protein cargo in relation to cardiovascular clinical manifestation is still lacking. To this end, 35 STEMI patients (17 experiencing resuscitated out-of-hospital cardiac arrest (OHCA-STEMI) and 18 uncomplicated) and 32 patients with chronic coronary syndrome (CCS) were enrolled. Plasma exosomes were characterized by the nanoparticle tracking analysis and Western blotting. Exosomes from STEMI patients displayed a higher concentration and size and a greater expression of platelet (GPIIb) and vascular endothelial (VE-cadherin) markers, but a similar amount of cardiac troponin compared to CCS. In addition, a difference in exosome expression of acute-phase proteins (ceruloplasmin, transthyretin and fibronectin) between STEMI and CCS patients was found. GPIIb and brain-associated marker PLP1 accurately discriminated between OHCA and uncomplicated STEMI. In conclusion, the exosome profile of STEMI patients has peculiar features that differentiate it from that of CCS patients, reflecting the pathophysiological mechanisms involved in STEMI. Additionally, the exosome expression of brain- and platelet-specific markers might allow the identification of patients experiencing ischemic brain injury in STEMI.     

Exploiting the Metabolic Consequences of PTEN Loss and Akt/Hexokinase 2 Hyperactivation in Prostate Cancer: A New Role for δ-Tocotrienol

The Warburg effect is commonly recognized as a hallmark of nearly all tumors. In prostate cancer (PCa), it has been shown to be driven by PTEN loss- and Akt hyperactivation-associated upregulation of hexokinase 2 (HK2). δ-Tocotrienol (δ-TT) is an extensively studied antitumor compound; however, its role in affecting PCa glycolysis is still unclear. Herein, we demonstrated that δ-TT inhibits glucose uptake and lactate production in PTEN-deficient LNCaP and PC3 PCa cells, by specifically decreasing HK2 expression. Notably, this was accompanied by the inhibition of the Akt pathway. Moreover, the nutraceutical could synergize with the well-known hypoglycemic agent metformin in inducing PCa cell death, highlighting the crucial role of the above metabolic phenotype in δ-TT-mediated cytotoxicity. Collectively, these results unravel novel inhibitory effects of δ-TT on glycolytic reprogramming in PCa, thus providing new perspectives into the mechanisms of its antitumor activity and into its use in combination therapy.