Biology subjects

Villanueva, S.

Publications and source records attributed to Villanueva, S..

7 recordsLinked to original sources

bioRxiv2026

Functional differences in electrolyte transport between the mouse proximal and distal trachea

AimThe mammalian tracheal epithelium is composed by different cell types unevenly distributed along the proximal-distal axis. Nevertheless, variations in expression and function of ion channels and transporters participating in fluid absorption and secretion had never been studied separately in proximal and distal sections of the mouse trachea. In this work, we aim to characterize basal and stimulated absorption and secretion of fluid obtained from proximal and distal trachea from the same animal. MethodsUssing chamber experiments were performed using a custom-made tissue slider that allowed the mounting small tracheal sections, where response to agonists and blockers was recorded. The role of the NKCC1 co-transporter was studied using the Slc12a2-/- mouse. A genetically tomato-induced mouse model was used to assess co-expression of NKCC1 and ASCL3 by immunofluorescence. Animals were instilled with different interleukins (ILs) to determine changes in absorption, secretion and mucus properties. ResultsProximal trachea didnt participate in sodium absorption but exhibited higher cAMP- and succinate-induced anion secretion than the distal section. NBCe1-dependent bicarbonate and TMEM16A-driven chloride secretion was significantly higher in the distal section. NKCC1+ cells were found in the submucosal glands (SMGs) and abundant patches of NKCC1+ cells in the distal region. Isolated NKCC1+ cells co-expressing ASCL3 were also detected. ILs treatment changed the electrophysiological properties of the distal but not the proximal trachea. ConclusionsOur experiments determined that the mouse trachea organizes its functions differentially in the proximal and distal sections, based in the functional distribution of channels, transporters and receptors. While the distal trachea drastically changed its responses to agonists inducing anion secretion the proximal trachea was unperturbed by the action of ILs.

physiology↗

bioRxiv2024

Airway succinate chemosensing induces CFTR-dependent anion secretion and mucus clearance which is impaired in cystic fibrosis.

The respiratory tract possesses a highly regulated innate defense system which includes efficient cilia-mediated mucus transport or mucociliary clearance (MCC). This essential process relies on appropriate hydration of airway surfaces which is controlled by a blend of transepithelial sodium and liquid absorption via the epithelial sodium channel (ENaC), and anion and liquid secretion, primarily regulated by the cystic fibrosis transmembrane conductance regulator (CFTR) channel. MCC is tightly regulated by second messenger signalling pathways. Succinate is derived from parasites, microorganisms and inflammatory cells, and its concentration increases in the airway surface liquid (ASL) during infections. Increases in ASL succinate activates the G-protein coupled succinate receptor (SUCNR1), which acts as a succinate sensor. Here, we tested the hypothesis that succinate signalling was linked to CFTR activity, ASL hydration and increased MCC. We observed that SUCNR1 activation stimulated anion secretion, increased mucus transport and induced bronchoconstriction in mouse airways. In parallel, stimulation of human bronchial epithelial cells (HBEC) with succinate activated anion secretion and increased ASL height. All functions activated by succinate/SUCNR1 were impeded when working with tissues and cells isolated from animal models or individuals affected cystic fibrosis (CF) or when CFTR was inhibited. Moreover, when HBECs derived from {Delta}F508 individuals were incubated with the triple drug combination of elexacaftor/tezacaftor/ivacaftor (ETI), succinate-induced anion secretion was restored, confirming the tight relationship between SUCNR1 signalling and CFTR function. Our results identify a novel activation pathway for CFTR that participates in the defence response of the airways, which is defective in CF. We propose that succinate acts as a danger molecule that alerts the airways to the presence of pathogens leading to a flushing out of the airways.

physiology↗

bioRxiv2021

Inhibition of the sodium-dependent HCO3 -transporter SLC4A4, produces a cystic fibrosis-like airway disease phenotype.

Bicarbonate secretion is a fundamental process involved in maintaining acid-base homeostasis. Disruption of bicarbonate entry into airway lumen, as has been observed in cystic fibrosis, produces several defects in lung function due to thick mucus accumulation. Bicarbonate is critical for correct mucin deployment and there is increasing interest in understanding its role in airway physiology, particularly in the initiation of lung disease in children affected by cystic fibrosis, in the absence of detectable bacterial infection. The current model of anion secretion in mammalian airways consists of CFTR and TMEM16A as apical anion exit channels, with limited capacity for bicarbonate transport compared to chloride. However, both channels can couple to SLC26A4 anion exchanger to maximise bicarbonate secretion. Nevertheless, current models lack any details about the identity of the basolateral protein(s) responsible for bicarbonate uptake into airway epithelial cells. We report herein that the electrogenic, sodium-dependent, bicarbonate cotransporter, SLC4A4, is expressed in the basolateral membrane of human and mouse airways, and that its pharmacological inhibition or genetic silencing reduces bicarbonate secretion. In fully differentiated primary human airway cells, SLC4A4 inhibition induced an acidification of the airways surface liquid and markedly reduced the capacity of cells to recover from an acid load. Studies in the Slc4a4-null mice revealed a previously unreported lung phenotype, characterized by mucus accumulation and reduced mucociliary clearance. Collectively, our results demonstrate that the reduction of SLC4A4 function induced a CF-like phenotype, even when chloride secretion remained intact, highlighting the important role SLC4A4 plays in bicarbonate secretion and mammalian airway function.

physiology↗

bioRxiv2021

Effect of reduced graphene oxide (rGO) compaction degree and concentration on rGO-polymer composites printability and cell interactions

Graphene derivatives combined with polymers have attracted enormous attention for bone tissue engineering applications. Among others, reduced graphene oxide (rGO) is one of the preferred graphene-based fillers for the preparation of composites via melt compounding, and their further processing into 3D scaffolds, due to its established large-scale production method, thermal stability, and electrical conductivity. In this study, rGO (low bulk density 10g/L) was compacted by densification using a solvent (either acetone or water) prior to melt compounding, to simplify its handling and dosing into a twin-screw extrusion system. The effects of rGO bulk density (medium and high), densification solvent, and rGO concentration (3, 10 and 15% in weight) on rGO dispersion within the composite, electrical conductivity, printability and cell-material interactions were studied. High bulk density rGO (90 g/L) occupied a low volume fraction within polymer composites, offering poor electrical properties but a reproducible printability up to 15 wt% rGO. On the other hand, the volume fraction within the composites of medium bulk density rGO (50 g/L) was higher for a given concentration, enhancing rGO particle interactions and leading to enhanced electrical conductivity, but compromising the printability window. For a given bulk density (50 g/L), rGO densified in water was more compacted and offered poorer dispersability within the polymer than rGO densified in acetone, and resulted in scaffolds with poor layer bonding or even lack of printability at high rGO percentages. A balance in printability and electrical properties was obtained for composites with medium bulk density rGO densified in acetone. Here, increasing rGO concentration led to more hydrophilic composites with a noticeable increase in protein adsorption. Moreover, scaffolds prepared with such composites presented antimicrobial properties even at low rGO contents (3 wt%). In addition, the viability and proliferation of human mesenchymal stromal cells (hMSCs) was maintained on scaffolds with up to 15% rGO and with enhanced osteogenic differentiation on 3% rGO scaffolds.

bioengineering↗

bioRxiv2021

Additive manufactured scaffolds for bone tissue engineering: physical characterization of thermoplastic composites with functional fillers

Thermoplastic polymer - filler composites are excellent materials for bone tissue engineering (TE) scaffolds, combining the functionality of fillers with suitable load bearing ability, biodegradability, and additive manufacturing (AM) compatibility of the polymer. Two key determinants of their utility are their rheological behavior in the molten state, determining AM processability, and their mechanical load-bearing properties. We report here the characterization of both these physical properties for four bone TE relevant composite formulations with poly(ethylene oxide terephthalate) / poly(butylene terephthalate (PEOT/PBT) as a base polymer, which is often used to fabricate TE scaffolds. The fillers used were reduced graphene oxide (rGO), hydroxyapatite (HA), gentamycin intercalated in zirconium phosphate (ZrP-GTM) and ciprofloxacin intercalated in MgAl layered double hydroxide (MgAl-CFX). The rheological assessment showed that generally the viscous behavior dominated the elastic behavior (G > G) for the studied composites, at empirically determined extrusion temperatures. Coupled rheological-thermal characterization of ZrP-GTM and HA composites showed that the fillers increased the solidification temperatures of the polymer melts during cooling. Both these findings have implications for the required extrusion temperatures and bonding between layers. Mechanical tests showed that the fillers generally made the polymer stiffer but more brittle in proportion to the filler fractions. Furthermore, the elastic moduli of scaffolds did not directly correlate with the corresponding bulk material properties, implying composite-specific AM processing effects on the mechanical properties. Lastly, we show computational models to predict multi-material scaffold elastic moduli using measured single material scaffold and bulk moduli. The reported characterizations are essential for assessing the AM processability and ultimately the suitability of the manufactured scaffolds for the envisioned bone regeneration application.

bioengineering↗

bioRxiv2020

Neuroanatomy of the grey seal brain: bringing pinnipeds into the neurobiological study of vocal learning

Comparative studies of vocal learning and vocal non-learning animals can increase our understanding of the neurobiology and evolution of vocal learning and human speech. Mammalian vocal learning is understudied: most research has either focused on vocal learning in songbirds or its absence in non-human primates. Here we focus on a highly promising model species for the neurobiology of vocal learning: grey seals. We provide a neuroanatomical atlas (based on dissected brain slices and magnetic resonance images), a labelled MRI template, a 3D model with volumetric measurements of brain regions, and histological cortical stainings. Four main features of the grey seal brain stand out. (1) It is relatively big and highly convoluted. (2) It hosts a relatively large temporal lobe and cerebellum, structures which could support developed timing abilities and acoustic processing. (3) The cortex is similar to humans in thickness and shows the expected six-layered mammalian structure. (4) Expression of FoxP2 - a gene involved in vocal learning and spoken language - is present in deeper layers of the cortex. Our results could facilitate future studies targeting the neural and genetic underpinnings of mammalian vocal learning, thus bridging the research gap from songbirds to humans and non-human primates.

animal behavior and cognition↗

bioRxiv2019

Urinary glucocorticoids in harbour seal (Phoca vitulina) pups during rehabilitation

The glucocorticoid (GC) hormone cortisol is often measured in animals to indicate their welfare and stress levels. However, the levels of other naturally occurring GCs are usually overlooked. We aimed to investigate whether aspects of the care and conditions of harbour seal (Phoca vitulina) pups in rehabilitation centres are reflected in urinary concentrations of four endogenous GCs. Urine samples were collected non-invasively from pups taken in as "orphans" at five different rehabilitation centres: three on the Irish Sea and two in the southern North Sea. Concentrations of urinary cortisol, cortisone, prednisolone and prednisone were analysed by mass spectrometry. Urinary concentrations of endogenous prednisolone and prednisone occurred in similar magnitude to cortisol, for the first time in any mammal species. The levels of all GC concentrations decreased as pups gained mass, but the most significant effect was for prednisone. Pups with mass less than 11kg, i.e. healthy average birth mass, had significantly higher levels of prednisone (but not of the other GCs) than pups of 11kg or more. Cortisol, cortisone and prednisolone concentrations were slightly higher for pups without access to water than those with water; however, we found no significant effect of social group on GC levels. Based on these findings, we tentatively suggest that the GCs may be elevated in harbour seal pups during rehabilitation in response to some physiological factors deviating from the norm of free-living pups. Our findings highlight the importance of measuring other GCs, in addition to cortisol, for understanding stressors affecting the welfare of seal pup in rehabilitation.

zoology↗