Establishment of the deuterium oxide dilution method as a new possibility for determining the transendothelial water permeability.

Increase in transendothelial water permeability is an essential etiological factor in a variety of diseases like edema and shock. Despite the high clinical relevance, there has been no precise method to detect transendothelial water flow until now. The deuterium oxide (DO) dilution method, already established for measuring transepithelial water transport, was used to precisely determine the transendothelial water permeability.

Serially passaged, conditionally reprogrammed nasal epithelial cells as a model to study epithelial functions and SARS-CoV-2 infection.

Primary airway epithelial cells (pAECs) cultivated at air-liquid interface (ALI) conditions are widely used as surrogates for human in vivo epithelia. To extend the proliferative capacity and to enable serially passaging of pAECs, conditional reprogramming (cr) has been employed in recent years. However, ALI epithelia derived from cr cells often display functional changes with increasing passages.

TGF-β1 increases permeability of ciliated airway epithelia via redistribution of claudin 3 from tight junction into cell nuclei.

TGF-β1 is a major mediator of airway tissue remodelling during atopic asthma and affects tight junctions (TJs) of airway epithelia. However, its impact on TJs of ciliated epithelia is sparsely investigated. Herein we elaborated effects of TGF-β1 on TJs of primary human bronchial epithelial cells.

Retinoic acid signalling adjusts tight junction permeability in response to air-liquid interface conditions.

The pulmonary epithelium separates the gaseous intraluminal space of the airways and the aqueous interstitium. This compartimentalization is required for appropriate lung function, it is established during perinatal periods and can be disturbed in lung edema. Herein we elaborated the impact of the air-liquid interface (ALI) on the function of the pulmonary epithelium.

IL-13 Impairs Tight Junctions in Airway Epithelia.

Interleukin-13 (IL-13) drives symptoms in asthma with high levels of T-helper type 2 cells (T2-cells). Since tight junctions (TJ) constitute the epithelial diffusion barrier, we investigated the effect of IL-13 on TJ in human tracheal epithelial cells. We observed that IL-13 increases paracellular permeability, changes claudin expression pattern and induces intracellular aggregation of the TJ proteins zonlua occludens protein 1, as well as claudins.

Pharmacological cholesterol depletion disturbs ciliogenesis and ciliary function in developing zebrafish.

Patients with an inherited inability to synthesize sufficient amounts of cholesterol develop congenital malformations of the skull, toes, kidney and heart. As development of these structures depends on functional cilia we investigated whether cholesterol regulates ciliogenesis through inhibition of hydroxymethylglutaryl-Coenzyme A reductase (HMG-CoA-R), the rate-limiting enzyme in cholesterol synthesis. HMG-CoA-R is efficiently inhibited by statins, a standard medication for hyperlipidemia.

Aquaporins in the lung.

The lung is the interface between air and blood where the exchange of oxygen and carbon dioxide occurs. The surface liquid that is directly exposed to the gaseous compartment covers both conducting airways and respiratory zone and forms the air-liquid interface. The barrier that separates this lining fluid of the airways and alveoli from the extracellular compartment is the pulmonary epithelium.

Inflammation-induced upregulation of P2X expression augments mucin secretion in airway epithelia.

Mucus clearance provides an essential innate defense mechanism to keep the airways and lungs free of particles and pathogens. Baseline and stimulated mucin secretion from secretory airway epithelial cells need to be tightly regulated to prevent mucus hypersecretion and mucus plugging of the airways. It is well established that extracellular ATP is a potent stimulus for regulated mucus secretion.

P2X receptor re-sensitization depends on a protonation/deprotonation cycle mediated by receptor internalization and recycling.

Re-sensitization of P2X receptors depends on a protonation/de-protonation cycle Protonation and de-protonation of the receptors is achieved by internalization and recycling of P2X receptors via acidic compartments Protonation and de-protonation occurs at critical histidine residues within the extracellular loop of P2X receptors Re-sensitization is blocked in the presence of the receptor agonist ATP ABSTRACT: P2X receptors are members of the P2X receptor family of cation-permeable, ligand-gated ion channels that open in response to the binding of extracellular ATP. P2X receptors are implicated in a variety of biological processes, including cardiac function, cell death, pain sensation and immune responses. These physiological functions depend on receptor activation on the cell surface.

Tight junctions in pulmonary epithelia during lung inflammation.

Inflammatory lung diseases like asthma bronchiale, chronic obstructive pulmonary disease and allergic airway inflammation are widespread public diseases that constitute an enormous burden to the health systems. Mainly classified as inflammatory diseases, the treatment focuses on strategies interfering with local inflammatory responses by the immune system. Inflammatory lung diseases predispose patients to severe lung failures like alveolar oedema, respiratory distress syndrome and acute lung injury.

Glucocorticoids Regulate Tight Junction Permeability of Lung Epithelia by Modulating Claudin 8.

The lung epithelium constitutes a selective barrier that separates the airways from the aqueous interstitial compartment. Regulated barrier function controls water and ion transport across the epithelium and is essential for maintaining lung function. Tight junctions (TJs) seal the epithelial barrier and determine the paracellular transport.

Amiloride-sensitive fluid resorption in NCI-H441 lung epithelia depends on an apical Cl(-) conductance.

Proper apical airway surface hydration is essential to maintain lung function. This hydration depends on well-balanced water resorption and secretion. The mechanisms involved in resorption are still a matter of debate, especially as the measurement of transepithelial water transport remains challenging.

Deuterium oxide dilution: a novel method to study apical water layers and transepithelial water transport.

Lung epithelia regulate the water flux between gas filled airways and the interstitial compartment in order to maintain organ function. Current methodology to assess transepithelial water transport is limited. We present a D2O dilution method to quantify submicroliter volumes of aqueous solutions on epithelial cell layers.

Fusion-activated cation entry (FACE) via P2X₄ couples surfactant secretion and alveolar fluid transport.

Two fundamental mechanisms within alveoli are essential for lung function: regulated fluid transport and secretion of surfactant. Surfactant is secreted via exocytosis of lamellar bodies (LBs) in alveolar type II (ATII) cells. We recently reported that LB exocytosis results in fusion-activated cation entry (FACE) via P2X₄ receptors on LBs.

Benzimidazolones enhance the function of epithelial Na⁺ transport.

Background And Purpose: Pharmacological enhancement of vectorial Na⁺ transport may be useful to increase alveolar fluid clearance. Herein, we investigated the influence of the benzimidazolones 1-ethyl-1,3-dihydro-2-benzimidazolone (1-EBIO), 5,6-dichloro-1-EBIO (DC-EBIO) and chlorzoxazone on vectorial epithelial Na⁺ transport.

Experimental Approach: Effects on vectorial Na⁺ transport and amiloride-sensitive apical membrane Na⁺ permeability were determined by measuring short-circuit currents (I(SC)) in rat fetal distal lung epithelial (FDLE) monolayers.

Combined atomic force microscopy-fluorescence microscopy: analyzing exocytosis in alveolar type II cells.

Hybrid atomic force microscopy (AFM)-fluorescence microscopy (FM) investigation of exocytosis in lung epithelial cells (ATII cells) allows the detection of individual exocytic events by FM, which can be simultaneously correlated to structural changes in individual cells by AFM. Exocytosis of lamellar bodies (LBs) represents a slow form of exocytosis found in many non-neuronal cells. Exocytosis of LBs, following stimulation with adenosine-5'-triphosphate (ATP) and phorbol 12-myristate 13-acetate (PMA), results in a cation influx via P2X(4) receptors at the site of LB fusion with the plasma membrane (PM), which should induce a temporary increase in cell height/volume.

Actin coating and compression of fused secretory vesicles are essential for surfactant secretion--a role for Rho, formins and myosin II.

Secretion of vesicular contents by exocytosis is a fundamental cellular process. Increasing evidence suggests that post-fusion events play an important role in determining the composition and quantity of the secretory output. In particular, regulation of fusion pore dilation and closure is considered a key regulator of the post-fusion phase.

An ultra fast detection method reveals strain-induced Ca(2+) entry via TRPV2 in alveolar type II cells.

A commonly used technique to investigate strain-induced responses of adherent cells is culturing them on an elastic membrane and globally stretching the membrane. However, it is virtually impossible to acquire microscopic images immediately after the stretch with this method. Using a newly developed technique, we recorded the strain-induced increase of the cytoplasmic Ca(2+) concentration ([Ca(2+)](c)) in rat primary alveolar type II (ATII) cells at an acquisition rate of 30ms and without any temporal delay.

Molecular basis of early epithelial response to streptococcal exotoxin: role of STIM1 and Orai1 proteins.

Streptolysin O (SLO) is a cholesterol-dependent cytolysin (CDC) from Streptococcus pyogenes. SLO induces diverse types of Ca(2+) signalling in host cells which play a key role in membrane repair and cell fate determination. The mechanisms behind SLO-induced Ca(2+) signalling remain poorly understood.

Fusion-activated Ca2+ entry via vesicular P2X4 receptors promotes fusion pore opening and exocytotic content release in pneumocytes.

Ca(2+) is considered a key element in multiple steps during regulated exocytosis. During the postfusion phase, an elevated cytoplasmic Ca(2+) concentration ([Ca(2+)])(c) leads to fusion pore dilation. In neurons and neuroendocrine cells, this results from activation of voltage-gated Ca(2+) channels in the plasma membrane.

Atomic force microscopy of microvillous cell surface dynamics at fixed and living alveolar type II cells.

Scanning probe techniques enable direct imaging of morphology changes associated with cellular processes at life specimen. Here, glutaraldehyde-fixed and living alveolar type II (ATII) cells were investigated by atomic force microscopy (AFM), and the obtained topographical data were correlated with results obtained by scanning electron microscopy (SEM) and confocal microscopy (CM). We show that low-force contact mode AFM at glutaraldehyde-fixed cells provides complementary results to SEM and CM.

Modulation of calcium-activated potassium channels induces cardiogenesis of pluripotent stem cells and enrichment of pacemaker-like cells.

Background: Ion channels are key determinants for the function of excitable cells, but little is known about their role and involvement during cardiac development. Earlier work identified Ca(2+)-activated potassium channels of small and intermediate conductance (SKCas) as important regulators of neural stem cell fate. Here we have investigated their impact on the differentiation of pluripotent cells toward the cardiac lineage.