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Following publication of the original article [1], the authors flagged that the ‘Availability of data and materials’ declaration is incomplete. Namely, it does not report the official accession number for the zebrafish PepT1a nucleotide sequence. With concern to this, please see the (correct) ‘Availability of data and materials’ declaration here: “Zebrafish pept1a (slc15a1a) nucleotide sequence has been submitted to GenBank (https://www.ncbi.nlm. nih.gov/nuccore/) and is available with the following accession number: GenBank Acc. No. MN723161” The authors apologize for any inconvenience caused.

POT transporters drive the concentrative uptake of their substrates by coupling to the transmembrane H+ electrochemical gradient. POTs recognize highly diverse di/tripeptides. Substrate extent expands with mammalian PepT1/PepT2, which also transport β-lactam antibiotics and peptide-based prodrugs. While substrate recognition changes, protonation sites seem conserved among members. In POTs, transport is achieved through the movement of the gating helices around the central binding site. The extracellular (EC) gate, formed by TM1,2 and TM7,8, serves to control access to the binding site from the EC side of the membrane. The intracellular (IC) gate, formed by TM4,5 and TM10,11, controls the release of peptide and protons on the inside of the cell. Two salt bridges coordinate these helices and control protein conformation. Whereas the IC gate contains a conserved Lys-Glu pair, the EC gate salt bridge is less conserved. In most of the bacterial POTs, the EC gate salt bridge is an Arg-Glu pair, while in mammalian PepT1, a conserved His on TM2 combined with an Arg-Asp salt bridge on TM1 and TM7 is seen. TM2 His is also found in “mammalian-like” bacterial members. In zebrafish PepT1b, the only vertebrate PepT1 known to work at alkaline pH, Lys replaces Arg on TM1. Likewise, PepT1 equally diverging from the “mammalian-like” transporters were retrieved (GenBank) from teleost fish (Cypriniformes, Cyprinodontiformes, Gymnotiformes, Gadiformes), birds (Apodiformes, Trochiliformes, Passeriformes, Piciformes), and even mammals (Chiroptera, Macroscelidea, Insectivora, Primates). Our findings extend the number of PepT1 prone to structure-function analyses and open to understand how their molecular diversity meets the physiology of the species and/or the environment where the species lives.

GI inflammation involves pathological processes affecting the gut epithelial barrier that are poorly reversible. A direct target of such processes is the enterocyte monolayer, which absorptive function is challenged by inflammation-induced cytoskeleton dynamics. Here, we evaluate the proinflammatory effects of PMA (phorbol-12-myristate-13-acetate) on Caco-2 intestinal cells at two different stages of spontaneous differentiation i.e. undifferentiated (7 dps, days post seeding) and differentiated enterocyte-like cells (>21 dps) in the presence/absence of the natural dipeptide carnosine (CAR) as potential anti-inflammatory molecule. By analysing actin cytoskeleton in 7 dps Caco-2 monolayers versus 21 dps monolayers, we revealed opposite effects of PMA which respectively induced disruption vs. intensification of actin rings and fibers; in both cases, simultaneous administration of PMA and CAR showed counteracting effects. Remarkably, the mRNA expression analysis of the ACTB (actin b) gene mirrored the morphological evidences and the counteraction of CAR. Moreover, the mRNA expression analysis of the AIF-1 gene (Allograft Inflammatory Factor 1) revealed opposite trends in 7 dps- versus 21 dps-grown Caco-2 with respect to PMA effects, and the same holds true for the SLC15A4/PHT1 gene involved both in intestinal peptide absorption and inflammatory responses; nevertheless, AIF-1 mRNA were not affected by CAR, whilst SLC15A4/PHT1 mRNA showed CAR-dependent up-regulation regardless of the differentiation stage. Overall, our results describe cytoskeletal and gene expression modulations which hint a model to distinguish physiological and inflammatory responses based on the differentiation stage of the Caco-2 monolayer and on the effects of CAR as differentially protective substrate.

The SoLute Carrier 15 (Slc15) family includes H+-dependent transporters that play a key role in the cellular uptake/reuptake of di/tripeptides and peptidomimetics. In mammals, in the epithelial cells of intestine and renal tubules two di/tripeptide transport systems have been characterized: the low-affinity/high-capacity system Slc15a1 (PepT1) and the high-affinity/low-capacity system Slc15a2 (PepT2). While PepT1 transporters have been studied in many teleost species, data on PepT2 is still lacking, except for zebrafish (Danio rerio). Here, we performed basic functional and expression analyses of a newly cloned Atlantic salmon (Salmo salar) PepT2. In Xenopus laevis oocytes, transient currents analysis showed that both total amount of charges moved (Q) and decay time (τ) vs membrane voltage shifted to more positive potential values when extracellular pH decreased, highlighting the role of H+ in the first step of transport cycle. Transport current vs voltage relations, as from Gly-L-Gln dose-response experiments, allowed kinetic parameters to be determined as a function of potential (from -140 to +20 mV) and external pH (5.5, 6.5, 7.6). Salmon PepT2 showed the higher apparent Gly-L-Gln affinity (K0.5) at pH 5.5 and 6.5 at the physiological membrane potential and an increase of maximal relative current (Imax) for more negative potentials and more acidic conditions. Notably, mRNA tissue expression analysis revealed that it is highly expressed in midgut and hindgut. Similar to zebrafish, this salmon PepT2 is a high-affinity/low-capacity transporter (K0.5 for Gly-Gln 4,4 μM, Imax -10 nA at -40 mV at pH 6.5), but its specific expression in the mid-to-distal portions of the gut opens to distinct and not yet known roles for a PepT2-type protein in fish physiology.

Di/tripeptides are key nutrients in animal diets, and fundamental for growth. They are transported across the membrane of enterocytes via the Slc15a1/PepT1 peptide transporter, that uses an inwardly-directed proton electrochemical gradient to drive the uptake. Due to a genome duplication event, PepT1a and PepT1b paralogues are found in teleost fish. Two PepT1a transporters, respectively cloned from the Atlantic salmon (Salmo salar) and zebrafish (Danio rerio), namely AsPepT1a and zfPepT1a, have been characterized. For both orthologs, function was verified by heterologous expression in Xenopus laevis oocytes, highlighting electrogenic, Na+-independent and pH-dependent transport similarly to the well-known PepT1b. The transient currents and the transport currents recorded from AsPept1a and zfPepT1a indicate significant functional differences with respect to PepT1b. PepT1a can be described as a low‐affinity/high‐capacity system, but its substrates preference profile is peculiar in the species, particularly for charged dipeptides. Moreover, the pre-steady state (PSS) currents, that reflect the first steps of transport cycle display in the charge/voltage relationship differences with respect to Pept1b in the voltage dependence. Considering that the PSS are consequence of the rearrangement of the protein in the membrane electric field, PepT1a transporters interact with the substrate differently from PepT1b. In addition, PepT1a and PepT1b have similar expression profile but different expression levels. These data together with a significant dissimilar substrate specificity, support the idea of distinct roles for these proteins in peptide recognition and transport.

SLC genes display different expression profiles along the rostral-caudal axis of the gut depending on species, digestive requirements and specialization of the sequential domains. Here, we report on the longitudinal expression profiles of SLC genes in the intestine of two simple-gutted teleost fish, zebrafish and Ballan wrasse (L. bergylta), using a subset of publicly available data from the GEO repository (zebrafish: [GSE20884]; Ballan wrasse: [GSE93191]). We explored intra-species/intra-dataset concordances/discordances in the expression profiles through Spearman simple correlation, which coefficients, even if heavily affected by the compositionality of the data, may be informative of which genes are co- or counter-expressed. Through hierarchical clustering on the correlation coefficients we defined groups of concordant and discordant profiles of SLC genes. An online app (freely available at https://gianmarcopiccinno.shinyapps.io/zebrafish_wrasse_v2/) was developed that allows rapid display of the longitudinal patterns of the genes differentially expressed in the two datasets. All the analyses were performed through R programming language. To define differentially expressed genes, we performed multiple One-Way ANOVA tests, whose p-values were subjected to Benjamini-Hochberg correction. Genes were considered differentially expressed if the corresponding corrected p-value from the ANOVA test was lower than 0.05. The plots were prepared using the R package ggplot2. The online app was produced through the R package shiny, and is hosted by free servers of RStudio. Our tools allow comparative analysis of the longitudinal profiles of the genes expressed in simple guts, and can be extended to implement any new similar datasets.

Endocrine Disruptors (EDs) are exogenous chemicals that interfere with the endocrine system and may cause adverse health effects. The difficulties encountered so far to identify EDs and define the risks to humans and wildlife strongly suggest that investigations must be performed with a new focus, based on main features of endocrine physiology: i) effects of hormones (Hs) are exerted at very low concentration through specific receptors and coupled pathways; ii) some Hs may act through more than one receptor; iii) Hs may act through different/multiple mechanisms in different physiological systems; iv) Hs produce different effects during animal life cycle. Same properties are shared by EDs and us such have to be investigated. Our laboratories demonstrated that natural and environmental estrogens affect embryo development in mussels and zebrafish, and impair cardiac activity leading to expression of cellular stress markers in seabass. BPA in particular is known to impair many steps of amphibians metamorphosis. We showed that it induces lipid accumulation in rat liver, and mimics the effect of E2 reducing ER intracellular levels and activating ER - dependent gene transcription in human cell lines; it also induces breast and trophoblast cell proliferation. Although the involvement of intracellular receptors was expected, the different models showed that membrane receptors, including Gprotein coupled receptors, are also targets, and MAPK, PI3K, calcium and cAMP – dependent pathways are involved. Data highlighted action mechanisms of EDs, suggested the taxonomic conservation of some routes, and provided the basis to establish key-events shared by Hs and EDs. Further work addressing comparative aspects from invertebrates to humans is needed to develop full understanding.

Programme and Abstracts of the 68th National Congress of the Italian Physiological Society (Società Italiana di Fisiologia) (Abstract) Carnosine (β-Ala-L-his, CAR) is an endogenous dipeptide acknowledged to contribute to maintenance of intracellular homeostasis in vertebrate excitable tissues. CAR biochemical properties (e.g. antioxidant and antiglycating activity) account for CAR homeostatic function(s) in many dysregulated states, such as hyperglycaemia upsetting diabetes-targeted tissue/organ districts. In this view, CAR hypoglycaemic effects have been shown to be physiologically regulated in the peripheral nervous system. Previously, we reported responsiveness of CAR-regulating genes in peripheral nerves of hyperglycaemic mice; remarkably, this responsiveness is strongly enhanced in the nerve’s fibroblast component. Here, we identify variations in expression of genes responsible for biosynthesis, degradation and transmembrane transport of CAR in the NIH/3T3 model of murine fibroblasts exposed to a hyperglycaemic stress, in the presence or absence of extracellular CAR. In parallel, we detect functional protection against the stress induced by glucose excess due to CAR, which inhibits ROS production and apoptotic events, and preserves fibroblast proliferation. Overall, the in vitro cellular model mirrors the ex vivo evidence in the peripheral nerves of hyperglycaemic mice, highlighting the (patho)physiological impact of CAR homeostatic variations in diabetes-targeted tissues and, remarkably, in fibroblasts, for which a functional role of ‘sensors’ of hyperglycaemic impairment in tissues could be reckoned.

Programme and Abstracts of the 68th National Congress of the Italian Physiological Society (Società Italiana di Fisiologia)

Programme and Abstracts of the 68th National Congress of the Italian Physiological Society (Società Italiana di Fisiologia)

Allograft Inflammatory Factor1 (AIF1), alias ionized calcium-binding adapter molecule 1 (IBA1), is a highly conserved Ca2+-binding cytokine that has been identified as a key regulator of the immune response in vertebrates. AIF1 is highly expressed in activated macrophages during inflammatory responses, thus representing an accurate indicator of macrophage activation in the body and a pathogenic factor in several inflammatory diseases. Proteins of the AIF1 superfamily are also present in invertebrates, from sponges to echinoderms. Here, we describe the Paracentrotus lividus Aif-1, which encodes a predicted protein of 151 amino acids with high similarity to vertebrate AIF1. In the common sea urchin, molecular and immunocytochemical analyses showed the constitutive expression of Aif-1 in the coelomocytes. Aif-1 localizes in the perinuclear area of amoebocytes and inside the granules of red cells, but it is not present in vibratile cells and colorless spherula cells. Moreover, significant increase of P. lividus Aif-1 expression, at both mRNA and protein level, are observed in coelomocytes after Gram+ bacterial challenge. BLAST searches across Echinoderm databases resulted in identification of orthologous proteins from 24 species (8 sea urchins, 1 brittle star, 12 starfishes and 3 sea cucumbers). Among these, P. lividus Aif-1 shared a high identity with several species, e.g., 85.4% with the sea urchin Strongylocentrotus purpuratus, 60.9% with the brittle star Ophiocoma echinata, 59.6% with the starfish Achantaster planci, and 52.3% with the sea cucumber Apostichopus japonicus. Our study on P. lividus Aif-1 will contribute to elucidate AIF1 function along the evolutionary scale and to consolidate the key evolutionary position of echinoderms throughout metazoans with respect to the common immune paths.

Programme and Abstracts of the 68th National Congress of the Italian Physiological Society (Società Italiana di Fisiologia) (Abstract) The SLC15A4 gene codes for the peptide/histidine transporter 1 (PHT1), a carrier of di/tripeptides and histidine. In vertebrates, the SLC15A4 genes are mainly expressed by the immune and nervous systems, in which they are acknowledged to undergo splicing events of unknown physiological meaning. After the early generic detection of the canonical SLC15A4/PHT1 isoforms on plasma and subcellular membranes, recent studies have pointed out that the human SLC15A4 gene codes for a late endosome/lysosome transmembrane carrier taking part in TLR7-, TLR9-, and NOD1-mediated pathways, thus suggesting that its gene products may act as regulators of inflammatory/immune/autoimmune onsets and diseases. Accordingly, SLC15A4/PHT1 altered activity has been associated to conditions such as type 2 diabetes, inflammatory bowel diseases (IBD) and systemic lupus erythematosus. Here, we report the description of a unique alternative splicing-mediated regulation of the SLC15A4 gene, conserved from zebrafish to human. We demonstrate that this regulation is physiologically facilitated by the Nonsense-Mediated mRNA Decay (NMD) surveillance system, in human immune-derived cells too, and that it is sensitive to inflammatory molecular triggers. Furthermore, in an IBD murine model induced with the inflammatory challenge by dextran sulfate sodium, we describe its impact on the expression of the alternative SLC15A4 gene products, revealing their differential modulations in different GI tracts.

In agreement with all authors of this paper the order of the authors and the contribution “Paola Lunetti[+], Alessandro Romano[+], Chiara Carrisi, Daniela Antonucci, Tiziano Verri, Giuseppe E. De Benedetto, Vincenza Dolce, Francesco P. Fanizzi, Michele Benedetti,* and Loredana Capobianco.* [+] These authors contributed equally to this paper. * Corresponding authors: Michele Benedetti and Loredana Capobianco, Department of Biological and Environmental Sciences and Technologies, University of Salento, 73100 Lecce.” is corrected to read the original order of the first submitted version of the paper: “Chiara Carrisi[+], Alessandro Romano[+], Paola Lunetti, Daniela Antonucci, Tiziano Verri, Giuseppe E. De Benedetto, Vincenza Dolce, Francesco P. Fanizzi, Michele Benedetti,* and Loredana Capobianco.* [+] These authors contributed equally to this paper. * Corresponding authors: Loredana Capobianco and Michele Benedetti, Department of Biological and Environmental Sciences and Technologies, University of Salento, 73100 Lecce.” For completeness the following individual contributions of the authors were added in the Supporting Information: “Author contributions: L.C., M.B., C.C., A.R. designed research; C.C., A.R., P.L. and D.A. performed research; L.C., M.B., C.C., A.R., V.D. and P.L. analysed data; F.P.F., A.R., L.C. and M.B. wrote the paper; L.C., M.B., T.V., G.D.B. and F.P.F. active discussion paper revision.”.

Submitted by Paulo Gavaia (pgavaia@ualg.pt) on 2014-06-06T09:52:10Z No. of bitstreams: 1 PepT1 in a coldwater marine teleost larvae atlantic cod cloning and preliminary studies of expression and phylogeny.pdf: 92089 bytes, checksum: 29d7c838f8ab20cd979bf7fa808abca9 (MD5) Approved for entry into archive by Pedro Sousa (a19661@ualg.pt) on 2014-06-25T12:43:54Z (GMT) No. of bitstreams: 1 PepT1 in a coldwater marine teleost larvae atlantic cod cloning and preliminary studies of expression and phylogeny.pdf: 92089 bytes, checksum: 29d7c838f8ab20cd979bf7fa808abca9 (MD5) Made available in DSpace on 2014-06-25T12:43:54Z (GMT). No. of bitstreams: 1 PepT1 in a coldwater marine teleost larvae atlantic cod cloning and preliminary studies of expression and phylogeny.pdf: 92089 bytes, checksum: 29d7c838f8ab20cd979bf7fa808abca9 (MD5) Previous issue date: 2006

Sepsis outcome is determined by a balance between inflammation and immune suppression. We aimed to evaluate monocytes polarization and reprogramming during these processes. We analyzed 93 patients with procalcitonin level >0.5 ng/mL (hPCT) and suspected/confirmed sepsis, and 84 controls by analysis of CD14, CD16 and HLA-DR expression on blood monocytes using fluorescent labeled monoclonal antibodies and BD FACS CANTO II. Complete blood cell count, procalcitonin and other biochemical markers were evaluated. Intermediate monocytes CD14++CD16+ increased in hPCT patients (including both positive and negative culture) compared to controls (13.6% ± 0.8 vs 6.2% ± 0.3, p

Il compito della fisiologia animale è quello di descrivere, analizzare e comparare, qualitativamente e quantitativamente, le funzioni che caratterizzano il mondo animale, dagli organismi unicellulari che possiedono tutte le funzioni a quelli pluricellulari più complessi la cui integrità è garantita dalle funzioni di cellule specializzate e organizzate in tessuti e organi. Il libro inizia con la presentazione delle caratteristiche chimico-fisiche dei diversi ambienti terrestri, a cui fare riferimento per capire o giustificare la comparsa di un determinato adattamento che ha reso possibile la vita in condizioni apparentemente impossibili. Viene proposta una fisiologia animale integrata con le strategie adottate per l’adattamento all’ambiente di vita, che esamina attraverso strumenti moderni e sofisticati non solo le differenze fisiologiche tra gli organismi, ma anche i fini processi e i geni che sottendono alle differenze esistenti. Lo scopo di questo libro è di offrire agli studenti universitari che seguono vari corsi di Fisiologia fonti utili di informazioni sui sistemi fisiologici e spiegazioni chiare dei concetti fisiologici facendo riferimento agli animali (vertebrati e invertebrati) nel loro ambiente naturale, non escludendo principi e concetti legati ai vari processi fisiologici per poter capire e valutare l’origine dei vari processi e considerando argomenti di attualità come la percezione degli stimoli esterni, le strategie respiratorie (la fisiologia dell’immersione, i pigmenti respiratori) o le migrazioni. Per catturare e mantenere l’interesse degli studenti sono state inserite fotografie, figure e schemi originali per illustrare e soprattutto chiarire i vari aspetti discussi nel testo. Il risultato è un testo dai contenuti molto aggiornati, con approfondimenti che originano da ricerche scientifiche molto recenti.

Alamandine is a peptide of the Renin Angiotensin System (RAS), either generated from Angiotensin A via the Angiotensin Converting Enzyme 2 (ACE2), or directly from Ang-(1–7). In mammals, it elicits cardioprotection via Mas-related G-protein-coupled receptor D (MrgD), and the NOS/NO system. In teleost fish, RAS is known to modulate heart performance. However, no information is available on the presence of a cardioactive ACE2/Alamandine axis. To fill this gap, we used the cyprinid teleost Carassius auratus (goldfish) for in silico and in vitro analyses. Via the NCBI Blast P suite we found that in cyprinids ace2 is phylogenetically detectable in a subcluster of proteins including ace2-like isoforms, and is correlated with a hypoxia-dependent pathway. By real-time PCR, Western Blotting, and HPLC, ACE2 and Alamandine were identified in goldfish heart and plasma, respectively. Both increased after chronic exposure to low O2 (2.6 mg O2 L−1). By using an ex-vivo working goldfish-heart preparation, we observed that in vitro administration of exogenous Alamandine dose-dependently stimulates myocardial contractility starting from 10−11 M. The effect that involved Mas-related receptors and PKA occurred via the NOS/NO system. This was shown by exposing the perfused heart to the NOS inhibitor L-NMMA (10−5 M) that abolished the cardiac effect of Alamandine and was supported by the increased expression of the phosphorylated NOS enzyme in the extract from goldfish heart exposed to 10−10 M Alamandine. Our data are the first to show that an ACE2/Alamandine axis is present in the goldfish C. auratus and, to elicit cardiac modulation, requires the obligatory involvement of the NOS/NO system.

Background: Molecular changes associated with the initiation of the epithelial to mesenchymal transition (EMT) program involve alterations of large proteome-based networks. The role of protein products mapping to non-coding genomic regions is still unexplored. Objective: The goal of this study was the identification of an alternative protein signature in breast cancer cellular models with a distinct expression of EMT markers. Methods: We profiled MCF-7 and MDA-MB-231 cells using liquid-chromatography mass/spectrometry (LCMS/ MS) and interrogated the OpenProt database to identify novel predicted isoforms and novel predicted proteins from alternative open reading frames (AltProts). Results: Our analysis revealed an AltProt and isoform protein signature capable of classifying the two breast cancer cell lines. Among the most highly expressed alternative proteins, we observed proteins potentially associated with inflammation, metabolism and EMT. Conclusion: Here, we present an AltProts signature associated with EMT. Further studies will be needed to define their role in cancer progression.

Nucleos(t)ide analogues (NA) belong to a family of compounds widely used in anticancer/antiviral treatments. They generally exhibit a cell toxicity limited by cellular uptake levels and the resulting nucleos(t)ides metabolism modifications, interfering with the cell machinery for nucleic acids synthesis. We previously synthesized purine nucleos(t)ide analogues N7-coordinated to a platinum centre with unaltered sugar moieties of the type: [Pt(dien)(N7-dGuo)]2+ (1; dien = diethylenetriamine; dGuo = 2'-deoxy-guanosine), [Pt(dien)(N7-dGMP)] (2; dGMP = 5'-(2'-deoxy)-guanosine monophosphate), and [Pt(dien)(N7-dGTP)]2- (3; dGTP = 5'-(2'-deoxy)-guanosine triphosphate), where the indicated electric charge is calculated at physiological pH (7.4). In this work, we specifically investigated the uptake of these complexes (1-3) at the plasma membrane level. Specific experiments on HeLa cervical cancer cells indicated a relevant cellular uptake of the model platinated deoxynucleos(t)ide 1 and 3 while complex 2 appeared unable to cross the cell plasma membrane. Obtained data buttress an uptake mechanism involving Na+-dependent concentrative transporters localized at the plasma membrane level. Consistently, 1 and 3 showed higher cytotoxicity with respect to complex 2 also suggesting selective possible applications as antiviral/antitumor drugs among the used model compounds.

Hydroxytyrosol (HT), a peculiar olive and olive oil phenolic antioxidant, plays a significant role in the endothelial and cardiovascular protection associated with olive oil consumption. However, studies examining the effects of HT on the whole-genome expression of endothelial cells, which are prominent targets for vasculo-protective effects of olive oil polyphenols, have been lacking. This study aims to comprehensively evaluate the genomic effects exerted by HT, at the transcriptional level, in endothelial cells under resting or proinflammatory conditions. Human umbilical vein endothelial cells (HUVECs) were treated with 10 µmol/L HT for 1 h and then stimulated with 5 ng/mL interleukin (IL)-1β for 3 h. Total RNA was extracted, and gene expression profile assessed with microarray analysis. Functional enrichment analysis and pathway analysis were performed by Ingenuity Pathways Analysis. Microarray data were validated by qRT-PCR. Fixing a significance threshold at 1.5-fold change, HT affected the expression of 708 and 599 genes, respectively, in HUVECs under resting and IL-1β-stimulated conditions, among these, 190 were common to both conditions. Unfolded protein response (UPR) and endoplasmic reticulum stress resulted from the two top canonical pathways common between HT and HT-IL-1β affected genes. IL-17F/A signaling was found in the top canonical pathways of HT modified genes under resting unstimulated conditions, whereas cardiac hypertrophy signaling was identified among the pathways affected by HT-IL-1β. The transcriptomic analysis allowed pinpointing immunological, inflammatory, proliferative, and metabolic-related pathways as the most affected by HT in endothelial cells. It also revealed previously unsuspected genes and related gene pathways affected by HT, thus broadening our knowledge of its biological properties. The unbiased identification of novel genes regulated by HT improves our understanding of mechanisms by which olive oil prevents or attenuates inflammatory diseases and identifies new genes to be enquired as potential contributors to the inter-individual variation in response to functional food consumption.

The Concise Guide to PHARMACOLOGY 2021/22 is the fifth in this series of biennial publications. The Concise Guide provides concise overviews, mostly in tabular format, of the key properties of nearly 1900 human drug targets with an emphasis on selective pharmacology (where available), plus links to the open access knowledgebase source of drug targets and their ligands (www.guidetopharmacology.org), which provides more detailed views of target and ligand properties. Although the Concise Guide constitutes over 500 pages, the material presented is substantially reduced compared to information and links presented on the website. It provides a permanent, citable, point-in-time record that will survive database updates. The full contents of this section can be found at http://onlinelibrary.wiley.com/doi/bph.15543. Transporters are one of the six major pharmacological targets into which the Guide is divided, with the others being: G protein-coupled receptors, ion channels, nuclear hormone receptors, catalytic receptors and enzymes. These are presented with nomenclature guidance and summary information on the best available pharmacological tools, alongside key references and suggestions for further reading. The landscape format of the Concise Guide is designed to facilitate comparison of related targets from material contemporary to mid-2021, and supersedes data presented in the 2019/20, 2017/18, 2015/16 and 2013/14 Concise Guides and previous Guides to Receptors and Channels. It is produced in close conjunction with the Nomenclature and Standards Committee of the International Union of Basic and Clinical Pharmacology (NC-IUPHAR), therefore, providing official IUPHAR classification and nomenclature for human drug targets, where appropriate.

Food intake is a vital process that supplies necessary energy and essential nutrients to the body. Information regarding luminal composition in the gastrointestinal tract (GIT) collected through mechanical and nutrient sensing mechanisms are generally conveyed, in both mammals and fish, to the hypothalamic neurocircuits. In this context, ghrelin, the only known hormone with an orexigenic action, and the intestinal peptide transporters 1 and 2, involved in absorption of dietary di- and tripeptides, exert important and also integrated roles for the nutrient uptake. Together, both are potentially involved in signaling pathways that control food intake originating from different segments of the GIT. However, little is known about the role of different paralogs and their response to fasting. Therefore, after 3 weeks of acclimatization, 12 Atlantic salmon (Salmo salar) post-smolt were fasted for 4 days to explore the gastrointestinal response in comparison with fed control (n = 12). The analysis covered morphometric (weight, length, condition factor, and wet content/weight fish %), molecular (gene expression variations), and correlation analyses. Such short-term fasting is a common and recommended practice used prior to any handling in commercial culture of the species. There were no statistical differences in length and weight but a significant lower condition factor in the fasted group. Transcriptional analysis along the gastrointestinal segments revealed a tendency of downregulation for both paralogous genes slc15a1a and slc15a1b and with significant lowered levels in the pyloric ceca for slc15a1a and in the pyloric ceca and midgut for slc15a1b. No differences were found for slc15a2a and slc15a2b (except a higher expression of the fasted group in the anterior midgut), supporting different roles for slc15 paralogs. This represents the first report on the effects of fasting on slc15a2 expressed in GIT in teleosts. Transcriptional analysis of ghrelin splicing variants (ghrl-1 and ghrl-2) showed no difference between treatments. However, correlation analysis showed that the mRNA expression for all genes (restricted to segment with the highest levels) were affected by the residual luminal content. Overall, the results show minimal effects of 4 days of induced fasting in Atlantic salmon, suggesting that more time is needed to initiate a large GIT response.

The feasibility to obtain highly porous collagen-based scaffolds (SCs) integrated with uniformly dispersed poly (lactide-co-glycolide)(PLGA) microparticles (MPs) was evaluated. PLGA-MPs were prepared by double emulsion technique and SCs with different amounts of MPs (SC/PLGA-MPs) were produced. Results showed that SC/PLGA-MPs could be successfully manufactured, the PLGA-MPs being physically retained in the SCs, without affecting the porous structure. PLGA-MPs also increased the SC hydrophilicity, acted as a mechanical reinforcement and retarded the degradation rate. In addition, the interactions between MPs and Caco-2 cells did not interfere with the correct morphological, adhesion and proliferation pathways.

To the SLC6 family belong 20 human transporters that utilize the sodium electrochemical gradient to move biogenic amines, osmolytes, amino acids and related compounds into cells. They are classified into two functional groups, the Neurotransmitter transporters (NTT) and Nutrient amino acid transporters (NAT). Here we summarize how since their first cloning in 1998, the insect (Lepidopteran) Orthologs of the SLC6 family transporters have represented very important tools for investigating functional–structural relationships, mechanism of transport, ion and pH dependence and substate interaction of the mammalian (and human) counterparts. Supplementary Information The online version contains supplementary material available at 10.1007/s11064-021-03410-1.

Fiber photometry is widely used in neuroscience labs for in vivo detection of functional fluorescence from optical indicators of neuronal activity with a simple optical fiber. The fiber is commonly placed next to the region of interest to both excite and collect the fluorescence signal. However, the path of both excitation and fluorescence photons is altered by the uneven optical properties of the brain, due to local variation of the refractive index, different cellular types, densities and shapes. Nonetheless, the effect of the local anatomy on the actual shape and extent of the volume of tissue that interfaces with the fiber has received little attention so far. To fill this gap, we measured the size and shape of fiber photometry efficiency field in the primary motor and somatosensory cortex, in the hippocampus and in the striatum of the mouse brain, highlighting how their substructures determine the detected signal and the depth at which photons can be mined. Importantly, we show that the information on the spatial expression of the fluorescent probes alone is not sufficient to account for the contribution of local subregions to the overall collected signal, and it must be combined with the optical properties of the tissue adjacent to the fiber tip.

The increasing access to antihypertensive medications has improved longevity and quality of life in hypertensive patients. Nevertheless, hypertension still remains a major risk factor for stroke and myocardial infarction, suggesting the need to implement management of pre- and hypertensive patients. In addition to antihypertensive medications, lifestyle changes, including healthier dietary patterns, such as the Dietary Approaches to Stop Hypertension (DASH) and the Mediterranean diet, have been shown to favorably affect blood pressure and are now recommended as integrative tools in hypertension management. An analysis of the effects of nutritional components of the Mediterranean diet(s) on blood pressure has therefore become mandatory. After a literature review of the impact of Mediterranean diet(s) on cardiovascular risk factors, we here analyze the effects of olive oil and its major components on blood pressure in healthy and cardiovascular disease individuals and examine underlying mechanisms of action. Both experimental and human studies agree in showing anti-hypertensive effects of olive oil. We conclude that due to its high oleic acid and antioxidant polyphenol content, the consumption of olive oil may be advised as the optimal fat choice in the management protocols for hypertension in both healthy and cardiovascular disease patients.

A label-free localized surface plasmon resonance (LSPR)-based biosensor exploiting gold nanorods (GNRs) is proposed and demonstrated. For this aim, 35±5 nm long and 20±4 thick GNRs spaced by a few nanometers thick polyelectrolytes (PE) from a gold thin film was analyzed and synthesized. The morphology of the GNRs, the plasmon properties of GNRs, swelling of PE layers and the wettability of the surfaces were characterized by transmission and scanning electron microscopy, spectroscopic reflectivity and contact angle measurements, respectively. Indeed, when immersed in a phosphate buffer saline solution, the GNRs-PE-gold system shows an optical shift of the LSPR wavelength. This shift was found to correspond to a vertical swelling of about 2 nm, demonstrating the extreme sensitivity of the biosensor. Finally, we show that LSPR measurements can be used to detect dynamic resonance changes in response to both thickness and buffer solution, while the hydrophobic behavior of the surface can be exploited for reducing the number of liquid analytes in clinical biosensing application.

Peptide transporter 1 (PepT1) mediates the uptake of dietary di-/tripeptides in vertebrates. However, in teleost fish gut, more than one PepT1-type transporter might operate, because of teleost-specific whole gen(om)e duplication event(s) that occurred during evolution. Here, we describe a novel teleost di-/tripeptide transporter, i.e., the Atlantic salmon ( Salmo salar) peptide transporter 1a [PepT1a; or solute carrier family 15 member 1a (Slc15a1a)], which is a paralog (77% similarity and 64% identity at the amino acid level) of the well-described Atlantic salmon peptide transporter 1b [PepT1b, alias PepT1; or solute carrier family 15 member 1b (Slc15a1b)]. Comparative analysis and evolutionary relationships of gene/protein sequences were conducted after ad hoc database mining. Tissue mRNA expression analysis was performed by quantitative real-time PCR, whereas transport function analysis was accomplished by heterologous expression in Xenopus laevis oocytes and two-electrode voltage-clamp measurements. Atlantic salmon pept1a is highly expressed in the proximal intestine (pyloric ceca ≈ anterior midgut > midgut >> posterior midgut), in the same gut regions as pept1b but notably ~5-fold less abundant. Like PepT1b, Atlantic salmon PepT1a is a low‐affinity/high‐capacity system. Functional analysis showed electrogenic, Na+-independent/pH-dependent transport and apparent substrate affinity ( K0.5) values for Gly-Gln of 1.593 mmol/L at pH 7.6 and 0.076 mmol/L at pH 6.5. In summary, we show that a piscine PepT1a-type transporter is functional. Defining the role of Atlantic salmon PepT1a in the gut will help to understand the evolutionary and functional relationships among peptide transporters. Its functional characterization will contribute to elucidate the relevance of peptide transporters in Atlantic salmon nutritional physiology.

Simple Summary Current research on nanocomposite materials with tailored physical–chemical properties is increasingly advancing in biomedical applications for bone regeneration. In this study, occurrence of differential responsiveness to dextran-grafted iron oxide (DM) nanoparticles and to their hybrid nano-hydroxyapatite (DM/n-HA) counterpart was investigated in human-derived, osteoblast-like cells. Sensitivity of cells in the presence of DMs or DM/n-HAs was evaluated in terms of cytoskeletal dynamics. Remarkably, it was shown that effects triggered by the DM are no more retained when DM is embedded onto DM/n-HA nanocomposites. In parallel, analyses on the expression of genes involved in (a) intracellular signaling pathways triggered by ligands or cell interactions with elements of the extracellular matrix, (b) modulation of processes such as cell cycle arrest, apoptosis, senescence, DNA repair, metabolism changes, and (c) iron homeostasis and absorption through cell membranes, indicated that the DM/n-HA-treated cells retain tracts of physiological responsiveness unlike DM-treated cells. Overall, a shielding effect by the n-HA was assumed (masking the DM’s cytotoxicity), and a modular biomimicry of the DM/n-HA nanocomposites. On these bases, the biocompatibility of n-HA associated to DM’s magnetic responsiveness offer a combination of structural/functional features of these nano-tools for bone tissue engineering, for finely acting within physiological ranges. Abstract The development of nanocomposites with tailored physical–chemical properties, such as nanoparticles containing magnetic iron oxides for manipulating cellular events at distance, implies exciting prospects in biomedical applications for bone tissue regeneration. In this context, this study aims to emphasize the occurrence of differential responsiveness in osteoblast-like cells to different nanocomposites with diverse features: dextran-grafted iron oxide (DM) nanoparticles and their hybrid nano-hydroxyapatite (DM/n-HA) counterpart. Here, responsiveness of cells in the presence of DMs or DM/n-HAs was evaluated in terms of cytoskeletal features. We observed that effects triggered by the DM are no more retained when DM is embedded onto the DM/n-HA nanocomposites. Also, analysis of mRNA level variations of the focal adhesion kinase (FAK), P53 and SLC11A2/DMT1 human genes showed that the DM/n-HA-treated cells retain tracts of physiological responsiveness compared to the DM-treated cells. Overall, a shielding effect by the n-HA component can be assumed, masking the DM’s cytotoxic potential, also hinting a modular biomimicry of the nanocomposites respect to the physiological responses of osteoblast-like cells. In this view, the biocompatibility of n-HA together with the magnetic responsiveness of DMs represent an optimized combination of structural with functional features of the DM/n-HA nano-tools for bone tissue engineering, for finely acting within physiological ranges.

In this work, zebrafish (Danio rerio) embryos were subjected to different physical stresses, consisting of a static and two low frequency magnetic fields, and two radio frequencies. Specifically, embryos were exposed to static (Bo, 0 Hz), very low frequency (VLF, 0.2 Hz), low frequency (LF, 270 kHz), very high frequency (VHF, 100 MHz) and ultra-high frequency (UHF, 900 MHz) magnetic field irradiations for up to 5 days. The field intensities were 40 mT, 40 mT, 470 μT, 240 nT and 240 nT, respectively. Untreated embryos were used as control (n = 10 for each condition).

Background Peptide transporter 1 (PepT1, alias Slc15a1) mediates the uptake of dietary di/tripeptides in all vertebrates. However, in teleost fish, more than one PepT1-type transporter might function, due to specific whole genome duplication event(s) that occurred during their evolution leading to a more complex paralogue gene repertoire than in higher vertebrates (tetrapods). Results Here, we describe a novel di/tripeptide transporter in the zebrafish (Danio rerio), i.e., the zebrafish peptide transporter 1a (PepT1a; also known as Solute carrier family 15 member a1, Slc15a1a), which is a paralogue (78% similarity, 62% identity at the amino acid level) of the previously described zebrafish peptide transporter 1b (PepT1b, alias PepT1; also known as Solute carrier family 15 member 1b, Slc15a1b). Also, we report a basic analysis of the pept1a (slc15a1a) mRNA expression levels in zebrafish adult tissues/organs and embryonic/early larval developmental stages. As assessed by expression in Xenopus laevis oocytes and two-electrode voltage clamp measurements, zebrafish PepT1a, as PepT1b, is electrogenic, Na+-independent, and pH-dependent and functions as a low-affinity system, with K0.5 values for Gly-Gln at − 60 mV of 6.92 mmol/L at pH 7.6 and 0.24 mmol/L at pH 6.5 and at − 120 mV of 3.61 mmol/L at pH 7.6 and 0.45 mmol/L at pH 6.5. Zebrafish pept1a mRNA is highly expressed in the intestine and ovary of the adult fish, while its expression in early development undergoes a complex trend over time, with pept1a mRNA being detected 1 and 2 days post-fertilization (dpf), possibly due to its occurrence in the RNA maternal pool, decreasing at 3 dpf (~ 0.5-fold) and increasing above the 1–2 dpf levels at 4 to 7 dpf, with a peak (~ 7-fold) at 6 dpf. Conclusions We show that the zebrafish PepT1a-type transporter is functional and co-expressed with pept1b (slc15a1b) in the adult fish intestine. Its expression is also confirmed during the early phases of development when the yolk syncytial layer is present and yolk protein resorption processes are active. While completing the missing information on PepT1-type transporters function in the zebrafish, these results open to future investigations on the similar/differential role(s) of PepT1a/PepT1b in zebrafish and teleost fish physiology.

Zebrafish (Danio rerio) embryos were subjected to different physical stresses, consisting of a static and two low frequency magnetic fields, and two radiofrequencies. Embryos were exposed to static (Bo, 0 Hz), very low frequency (VLF, 0.2 Hz), low frequency (LF, 270 kHz), very high frequency (VHF, 100 MHz), and ultra-high frequency (UHF, 900 MHz) magnetic field irradiation for up to 5 days. The field intensities were 40 mT, 40 mT, 470 µT, 240 nT and 240 nT, respectively. Untreated embryos were used as control.(n= 10 for each condition).

Chronic inflammation of the adipose tissue (AT) is a major contributor to obesity-associated cardiometabolic complications. The olive oil polyphenol hydroxytyrosol (HT) contributes to Mediterranean diet cardiometabolic benefits through mechanisms still partially unknown. We investigated HT (1 and 10 &mu, mol/L) effects on gene expression (mRNA and microRNA) related to inflammation induced by 10 ng/mL tumor necrosis factor (TNF)-&alpha, in human Simpson&ndash, Golabi&ndash, Behmel Syndrome (SGBS) adipocytes. At real-time PCR, HT significantly inhibited TNF-&alpha, induced mRNA levels, of monocyte chemoattractant protein-1, C-X-C Motif Ligand-10, interleukin (IL)-1&beta, IL-6, vascular endothelial growth factor, plasminogen activator inhibitor-1, cyclooxygenase-2, macrophage colony-stimulating factor, matrix metalloproteinase-2, Cu/Zn superoxide dismutase-1, and glutathione peroxidase, as well as surface expression of intercellular adhesion molecule-1, and reverted the TNF-&alpha, mediated inhibition of endothelial nitric oxide synthase, peroxisome proliferator-activated receptor coactivator-1&alpha, and glucose transporter-4. We found similar effects in adipocytes stimulated by macrophage-conditioned media. Accordingly, HT significantly counteracted miR-155-5p, miR-34a-5p, and let-7c-5p expression in both cells and exosomes, and prevented NF-&kappa, B activation and production of reactive oxygen species. HT can therefore modulate adipocyte gene expression profile through mechanisms involving a reduction of oxidative stress and NF-&kappa, B inhibition. By such mechanisms, HT may blunt macrophage recruitment and improve AT inflammation, preventing the deregulation of pathways involved in obesity-related diseases.