Structural and biological characterization of shortened derivatives of the cathelicidin PMAP-36

Cathelicidins, a family of host defence peptides in vertebrates, play an important role in the innate immune response, exhibiting antimicrobial activity against many bacteria, as well as viruses and fungi. This work describes the design and synthesis of shortened analogues of porcine cathelicidin PMAP-36, which contain structural changes to improve the pharmacokinetic properties. In particular, 20-mers based on PMAP-36 (residues 12-31) and 13-mers (residues 12-24) with modification of amino acid residues at critical positions and introduction of lipid moieties of different lengths were studied to identify the physical parameters, including hydrophobicity, charge, and helical structure, required to optimise their antibacterial activity. Extensive conformational analysis, performed by CD and NMR, revealed that the substitution of Pro25-Pro26 with Ala25-Lys26 increased the alpha-helix content of the 20-mer peptides, resulting in broad-spectrum antibacterial activity against Escherichia coli, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa and Staphylococcus epidermidis strains. Interestingly, shortening to just 13 residues resulted in only a slight decrease in antibacterial activity. Furthermore, two sequences, a 13-mer and a 20-mer, did not show cytotoxicity against HaCat cells up to 64 mu M, indicating that both derivatives are not only effective but also selective antimicrobial peptides. In the short peptide, the introduction of the helicogenic alpha-aminoisobutyric acid forced the helix toward a prevailing 3(10) structure, allowing the antimicrobial activity to be maintained. Preliminary tests of resistance to Ser protease chymotrypsin indicated that this modification resulted in a peptide with an increased in vivo lifespan. Thus, some of the PMAP-36 derivatives studied in this work show a good balance between chain length, antibacterial activity, and selectivity, so they represent a good starting point for the development of even more effective and proteolysis-resistant active peptides

An Interplay between a Face-Centred Composite Experimental Design and Solid-Phase Microextraction for Wine Aroma GC/MS Analysis

For oenological products, most of the intrinsic and extrinsic drivers of perceived quality are associated with specific aromatic profiles. Aromatic diversity has been recognized as a central element in perceived quality as it is able to transmit the complex interactions between grape variety, geographical characteristics, and viticultural and winemaking practices, including the fermentative process. A comprehensive characterization of flavour compounds by headspace solid-phase microextraction (HS-SPME) and gas chromatography coupled to mass spectrometric analysis is often needed in order to ascertain the quality of wine. HS-SPME requires a proper optimization that can be achieved through an adequate experimental design. Here, a HS-SPME/GC-MS based method was developed to investigate the volatile compounds of wine samples obtained by laboratory-scale fermentations. This was performed by inoculating a commercial Saccharomyces cerevisiae strain, which is used both as single starter and as mixed starter, with an indigenous Hanseniaspora osmophila strain. The experimental conditions of HS-SPME (extraction temperature and time) were optimized by applying a face-centred composite experimental design. Up to 95% of the total variance was explained by the proposed model. The optimized method allowed us to confirm the usefulness of combining the inoculation of grapes with selected yeast strains in co-culture situations in order to improve the wine bouquet

Development and Validation of a Reversed-Phase HPLC Method with UV Detection for the Determination of L-Dopa in Vicia faba L. Broad Beans

L-Dopa (LD), a substance used medically in the treatment of Parkinson's disease, is found in several natural products, such as Vicia faba L., also known as broad beans. Due to its low chemical stability, LD analysis in plant matrices requires an appropriate optimization of the chosen analytical method to obtain reliable results. This work proposes an HPLC-UV method, validated according to EURACHEM guidelines as regards linearity, limits of detection and quantification, precision, accuracy, and matrix effect. The LD extraction was studied by evaluating its aqueous stability over 3 months. The best chromatographic conditions were found by systematically testing several C-18 stationary phases and acidic mobile phases. In addition, the assessment of the best storage treatment of Vicia faba L. broad beans able to preserve a high LD content was performed. The best LD determination conditions include sun-drying storage, extraction in HCl 0.1 M, chromatographic separation with a Discovery C-18 column, 250 x 4.6 mm, 5 mu m particle size, and 99% formic acid 0.2% v/v and 1% methanol as the mobile phase. The optimized method proposed here overcomes the problems linked to LD stability and separation, thus contributing to the improvement of its analytical determination

The upstream Variable Number Tandem Repeat polymorphism of the monoamine oxidase type A gene influences trigeminal pain-related evoked responses

Monoamines have an important role in neural plasticity, a key factor in cortical pain processing that promotes changes in neuronal network connectivity. Monoamine oxidase type A (MAOA) is an enzyme that, due to its modulating role in monoaminergic activity, could play a role in cortical pain processing. The X-linked MAOA gene is characterized by an allelic variant of length, the MAOA upstream Variable Number Tandem Repeat (MAOA-uVNTR) region polymorphism. Two allelic variants of this gene are known, the high-activity MAOA (HAM) and low-activity MAOA (LAM). We investigated the role of MAOA-uVNTR in cortical pain processing in a group of healthy individuals measured by the trigeminal electric pain-related evoked potential (tPREP) elicited by repeated painful stimulation. A group of healthy volunteers was genotyped to detect MAOA-uVNTR polymorphism. Electrical tPREPs were recorded by stimulating the right supraorbital nerve with a concentric electrode. The N2 and P2 component amplitude and latency as well as the N2-P2 inter-peak amplitude were measured. The recording was divided into three blocks, each containing 10 consecutive stimuli and the N2-P2 amplitude was compared between blocks. Of the 67 volunteers, 37 were HAM and 30 were LAM. HAM subjects differed from LAM subjects in terms of amplitude of the grand-averaged and first-block N2-P2 responses (HAM>LAM). The N2-P2 amplitude decreased between the first and third block in HAM subjects but not LAM subjects. The MAOA-uVNTR polymorphism seemed to influence the brain response in a repeated tPREP paradigm and suggested a role of the MAOA as a modulator of neural plasticity related to cortical pain processing

Detection of anti-SARS CoV-2 antibodies in human serum by means of Bloch surface waves on 1D photonic crystal biochips

This study presents the development and characterization of a disposable biochip for the detection of antibodies against the SARS-CoV-2 spike protein, a well-known target for vaccine and therapeutic development. This biochip is based on a one-dimensional photonic crystal (1DPC) deposited on a plastic substrate and designed to sustain Bloch surface waves (BSW) in the visible range. The experimental phase was carried out using the biochip in conjunction with a custom-made optical read-out platform capable of real-time refractometric detection and fluorescence-based end-point measurements. Our biochip was functionalized by immobilizing the receptor-binding domain of the spike protein onto the surface using a silanization process. Human serum samples, including a negative control and a positive sample from a recovered COVID-19 patient, were tested on the biochip. The experimental results show that the biochip discriminates between positive and negative samples in a label-free refractometric mode down to a 1:10 dilution of the sera and in quantum dot amplified refractometric and fluorescence mode down to 1:100 dilution. The results demonstrate the potential of the disposable biochip for sensitive and specific detection of COVID-19 antibodies

The actin-binding protein profilin 2 is a novel regulator of iron homeostasis

Cellular iron homeostasis is controlled by the iron regulatory proteins (IRPs) 1 and 2 that bind cis-regulatory iron-responsive elements (IRE) on target messenger RNAs (mRNA). We identified profilin 2 (Pfn2) mRNA, which encodes an actin-binding protein involved in endocytosis and neurotransmitter release, as a novel IRP-interacting transcript, and studied its role in iron metabolism. A combination of electrophoretic mobility shift assay experiments and bioinformatic analyses led to the identification of an atypical and conserved IRE in the 39 untranslated region of Pfn2 mRNA. Pfn2 mRNA levels were significantly reduced in duodenal samples from mice with intestinal IRP ablation, suggesting that IRPs exert a positive effect on Pfn2 mRNA expression in vivo. Overexpression of Pfn2 in HeLa and Hepa1-6 cells reduced their metabolically active iron pool. Importantly, Pfn2-deficient mice showed iron accumulation in discrete areas of the brain (olfactory bulb, hippocampus, and midbrain) and reduction of the hepatic iron store without anemia. Despite low liver iron levels, hepatic hepcidin expression remained high, likely because of compensatory activation of hepcidin by mild inflammation. Splenic ferroportin was increased probably to sustain hematopoiesis. Overall, our results indicate that Pfn2 expression is controlled by the IRPs in vivo and that Pfn2 contributes to maintaining iron homeostasis in cell lines and mice

Legal Cannabis sativa L. Dried Inflorescences: Cannabinoids Content and Cytotoxic Activity against Human HepG2 Cell Line

Cannabis sativa L. has health benefits, principally due to the levels and ratios of two impor- tant cannabinoids, ∆9-tetrahydrocannabinol (THC) and cannabidiol (CBD). THC:CBD ratio affects their pharmacological interaction for the treatment of different diseases as well as its modulation allows for a custom-made product that utilizes the distinguishing effects of CBD, THC, or both, for a peculiar patient or clinical effect. This study aims to investigate the total content of THC, CBD, and their ratio in 34 dried inflorescence legally sold in physical and online stores, by using a validated liquid chromatography-ultraviolet (HPLC-UV) method, after cannabinoids identification performed through MSn studies. Cannabinol (CBN) content was also monitored to evaluate hemp age or con- servation status. CBN content always resulted lower than limit of quantification, thus confirming well-stored fresh hemp. All investigated samples showed a total THC amount below 0.59% w/w, thus responding to legal requirements.. The total CBD amount ranged from 2.62 to 20.27% w/w and it was not related to THC level. THC:CBD ranged among 1:3 and 1:26, thus ascertaining their suitability for different target pharmacological uses. In vitro studies using human hepatoblastoma cell line HepG2 suggested that hemp extracts with THC:CBD ratios of 1:9 exhibited higher toxicity than pure cannabinoids

The ARIEL Instrument Control Unit design for the M4 Mission Selection Review of the ESA's Cosmic Vision Program

The Atmospheric Remote-sensing Infrared Exoplanet Large-survey mission (ARIEL) is one of the three present candidates for the ESA M4 (the fourth medium mission) launch opportunity. The proposed Payload will perform a large unbiased spectroscopic survey from space concerning the nature of exoplanets atmospheres and their interiors to determine the key factors affecting the formation and evolution of planetary systems. ARIEL will observe a large number (>500) of warm and hot transiting gas giants, Neptunes and super-Earths around a wide range of host star types, targeting planets hotter than 600 K to take advantage of their well-mixed atmospheres. It will exploit primary and secondary transits spectroscopy in the 1.2-8 um spectral range and broad-band photometry in the optical and Near IR (NIR). The main instrument of the ARIEL Payload is the IR Spectrometer (AIRS) providing low-resolution spectroscopy in two IR channels: Channel 0 (CH0) for the 1.95-3.90 um band and Channel 1 (CH1) for the 3.90-7.80 um range. It is located at the intermediate focal plane of the telescope and common optical system and it hosts two IR sensors and two cold front-end electronics (CFEE) for detectors readout, a well defined process calibrated for the selected target brightness and driven by the Payload's Instrument Control Unit (ICU).Comment: Experimental Astronomy, Special Issue on ARIEL, (2017

Cosmological Parameters from the 2003 flight of BOOMERANG

We present the cosmological parameters from the CMB intensity and polarization power spectra of the 2003 Antarctic flight of the BOOMERANG telescope. The BOOMERANG data alone constrains the parameters of the $\Lambda$CDM model remarkably well and is consistent with constraints from a multi-experiment combined CMB data set. We add LSS data from the 2dF and SDSS redshift surveys to the combined CMB data set and test several extensions to the standard model including: running of the spectral index, curvature, tensor modes, the effect of massive neutrinos, and an effective equation of state for dark energy. We also include an analysis of constraints to a model which allows a CDM isocurvature admixture.Comment: 18 pages, 10 figures, submitted to Ap