Propidium uptake and ATP release in A549 cells share similar transport mechanisms.

The lipid bilayer of eukaryotic cells' plasma membrane is almost impermeable to small ions and large polar molecules, but its miniscule basal permeability in intact cells is poorly characterized. This report describes the intrinsic membrane permeability of A549 cells toward the charged molecules propidium (Pr) and ATP. Under isotonic conditions, we detected with quantitative fluorescence microscopy, a continuous low-rate uptake of Pr (∼150 × 10 moles (zmol)/h/cell, [Pr] = 150 μM, 32°C).

Activation of Subcutaneous Mast Cells in Acupuncture Points Triggers Analgesia.

This review summarizes experimental evidence indicating that subcutaneous mast cells are involved in the trigger mechanism of analgesia induced by acupuncture, a traditional oriental therapy, which has gradually become accepted worldwide. The results are essentially based on work from our laboratories. Skin mast cells are present at a high density in acupuncture points where fine needles are inserted and manipulated during acupuncture intervention.

Lytic Release of Cellular ATP: Physiological Relevance and Therapeutic Applications.

The lytic release of ATP due to cell and tissue injury constitutes an important source of extracellular nucleotides and may have physiological and pathophysiological roles by triggering purinergic signalling pathways. In the lungs, extracellular ATP can have protective effects by stimulating surfactant and mucus secretion. However, excessive extracellular ATP levels, such as observed in ventilator-induced lung injury, act as a danger-associated signal that activates NLRP3 inflammasome contributing to lung damage.

P2Y and P2X receptors modulate mechanically induced adenosine triphosphate release from mast cells.

Subcutaneous mast cells (MCs) are vulnerable to mechanical stimulation from external environment. Thus, MCs immune function could be modulated by their mechanosensitivity. This property has been identified as the trigger mechanism of needling acupuncture, a traditional oriental therapy.

Type 2 secretory cells are primary source of ATP release in mechanically stretched lung alveolar cells.

Extracellular ATP and its metabolites are potent paracrine modulators of lung alveolar cell function, including surfactant secretion and fluid transport, but the sources and mechanism of intra-alveolar ATP release remain unclear. To determine the contribution of gas-exchanging alveolar type 1 (AT1) and surfactant-secreting type 2 (AT2) cells to stretch-induced ATP release, we used quantitative real-time luminescence ATP imaging and rat primary alveolar cells cultured on silicon substrate for 2-7 days. When cultured on solid support, primary AT2 cells progressively transdifferentiated into AT1-like cells with ~20% of cells showing AT1 phenotype by day 2-3 (AT2:AT1 ≈ 4:1), while on day 7, the AT2:AT1 cell ratio was reversed with up to 80% of the cells displaying characteristics of AT1 cells.

Elevation of Intracellular Na Contributes to Expression of Early Response Genes Triggered by Endothelial Cell Shrinkage.

Background/aims: Prolonged hyperosmotic shrinkage evokes expression of osmoprotective genes via nuclear factor NFAT5-mediated pathway and activates Na influx via hypertonicity-induced cation channels (HICC). In human umbilical vein endothelial cells (HUVEC) elevation of intracellular sodium concentration ([Na]) triggers transcription of dozens of early response genes (ERG). This study examined the role of monovalent cations in the expression of Na-sensitive ERGs in iso- and hyperosmotically shrunken HUVEC.

Dissociation of natriuresis and diuresis by oxytocin molecular forms in rats.

In the rat, oxytocin (OT) produces dose-dependent diuretic and natriuretic responses. Post-translational enzymatic conversion of the OT biosynthetic precursor forms both mature and C-terminally extended peptides. The plasma concentrations of these C-terminally extended peptides (OT-G; OT-GK and OT-GKR) are elevated in newborns and pregnant rats.

Wide field of view quantitative imaging of cellular ATP release.

Although several mechanical stressors promote ATP secretion from eukaryotic cells, few mechanosensitive pathways for ATP release have been precisely characterized and none have been clearly identified. To facilitate progress, we report here a wide field of view (∼20 × 20 mm sample area) imaging technique paired with a quantitative image analysis to accurately map the dynamics of ATP release from a cell population. The approach has been tested on A549 cells stretched at high initial strain rate (2-5 s) or swelled by hypotonic shock.

Mechanosensitive ATP release in the lungs: New insights from real-time luminescence imaging studies.

Extracellular ATP and other nucleotides are important autocrine/paracrine mediators that stimulate purinergic receptors and regulate diverse processes in the normal lungs. They are also associated with pathogenesis of a number of respiratory diseases and clinical complications including acute respiratory distress syndrome and ventilator induced lung injury. Mechanical forces are major stimuli for cellular ATP release but precise mechanisms responsible for this release are still debated.

Search for Upstream Cell Volume Sensors: The Role of Plasma Membrane and Cytoplasmic Hydrogel.

The plasma membrane plays a prominent role in the regulation of cell volume by mediating selective transport of extra- and intracellular osmolytes. Recent studies show that upstream sensors of cell volume changes are mainly located within the cytoplasm that displays properties of a hydrogel and not in the plasma membrane. Cell volume changes occurring in anisosmotic medium as well as in isosmotic environment affect properties of cytoplasmic hydrogel that, in turn, trigger rapid regulatory volume increase and decrease (RVI and RVD).

Effects of Hypoxia on Erythrocyte Membrane Properties-Implications for Intravascular Hemolysis and Purinergic Control of Blood Flow.

Intravascular hemolysis occurs in hereditary, acquired, and iatrogenic hemolytic conditions but it could be also a normal physiological process contributing to intercellular signaling. New evidence suggests that intravascular hemolysis and the associated release of adenosine triphosphate (ATP) may be an important mechanism for local purinergic signaling and blood flow regulation during exercise and hypoxia. However, the mechanisms that modulate hypoxia-induced RBC membrane fragility remain unclear.

Imaging viscosity of intragranular mucin matrix in cystic fibrosis cells.

Abnormalities of mucus viscosity play a critical role in the pathogenesis of several respiratory diseases, including cystic fibrosis. Currently, there are no approaches to assess the rheological properties of mucin granule matrices in live cells. This is the first example of the use of a molecular rotor, a BODIPY dye, to quantitatively visualize the viscosity of intragranular mucin matrices in a large population of individual granules in differentiated primary bronchial epithelial cells using fluorescence lifetime imaging microscopy.

STIM-1 and ORAI-1 channel mediate angiotensin-II-induced expression of Egr-1 in vascular smooth muscle cells.

An upregulation of Egr-1 expression has been reported in models of atherosclerosis and intimal hyperplasia and, various vasoactive peptides and growth promoting stimuli have been shown to induce the expression of Egr-1 in vascular smooth muscle cells (VSMC). Angiotensin-II (Ang-II) is a key vasoactive peptide that has been implicated in the pathogenesis of vascular diseases. Ang-II elevates intracellular Ca through activation of the store-operated calcium entry (SOCE) involving an inositol-3-phosphate receptor (IP3R)-coupled depletion of endoplasmic reticular Ca and a subsequent activation of the stromal interaction molecule 1 (STIM-1)/Orai-1 complex.

Real-time imaging of inflation-induced ATP release in the ex vivo rat lung.

Extracellular ATP and other nucleotides are important autocrine/paracrine mediators that regulate diverse processes critical for lung function, including mucociliary clearance, surfactant secretion, and local blood flow. Cellular ATP release is mechanosensitive; however, the impact of physical stimuli on ATP release during breathing has never been tested in intact lungs in real time and remains elusive. In this pilot study, we investigated inflation-induced ATP release in rat lungs ex vivo by real-time luciferin-luciferase (LL) bioluminescence imaging coupled with simultaneous infrared tissue imaging to identify ATP-releasing sites.

Calcium is not required for triggering volume restoration in hypotonically challenged A549 epithelial cells.

Maintenance of cell volume is a fundamental housekeeping function in eukaryotic cells. Acute cell swelling activates a regulatory volume decrease (RVD) process with poorly defined volume sensing and intermediate signaling mechanisms. Here, we analyzed the putative role of Ca signaling in RVD in single substrate-adherent human lung epithelial A549 cells.

Deoxygenation Affects Composition of Membrane-Bound Proteins in Human Erythrocytes.

Background/aims: ATP release from erythrocyte plays a key role in hypoxia-induced elevation of blood flow in systematic circulation. We have previously shown that hemolysis contributes to erythrocyte ATP release triggered by several stimuli, including hypoxia, but the molecular mechanisms of hypoxia-increased membrane fragility remain unknown.

Methods: In this study, we compared the action of hypoxia on hemolysis, ATP release and the composition of membrane-bound proteins in human erythrocytes.

Complementary roles of KCa3.1 channels and β1-integrin during alveolar epithelial repair.

Background: Extensive alveolar epithelial injury and remodelling is a common feature of acute lung injury and acute respiratory distress syndrome (ARDS) and it has been established that epithelial regeneration, and secondary lung oedema resorption, is crucial for ARDS resolution. Much evidence indicates that K(+) channels are regulating epithelial repair processes; however, involvement of the KCa3.1 channels in alveolar repair has never been investigated before.

Heat induces adenosine triphosphate release from mast cells in vitro: a putative mechanism for moxibustion.

Objective: To investigate the role of adenosine tri-phosphate (ATP) purinergic signaling in mast cells (MCs) modulated by heat to further understand the molecular mechanisms of moxibustion.

Methods: Skin temperatures induced by monkshood cake moxibustion were evaluated by measuring the Neiguan acupoint (PC 6) from 31 participants with a digital thermocouple thermometer. Temperatures of 43 °C and 52 °C were applied to cultured human leukemia mast cell line HMC-1 in vitro.

Role of cytoskeleton network in anisosmotic volume changes of intact and permeabilized A549 cells.

Recently we found that cytoplasm of permeabilized mammalian cells behaves as a hydrogel displaying intrinsic osmosensitivity. This study examined the role of microfilaments and microtubules in the regulation of hydrogel osmosensitivity, volume-sensitive ion transporters, and their contribution to volume modulation of intact cells. We found that intact and digitonin-permeabilized A549 cells displayed similar rate of shrinkage triggered by hyperosmotic medium.

Modulation of extracellular ATP content of mast cells and DRG neurons by irradiation: studies on underlying mechanism of low-level-laser therapy.

Low-level-laser therapy (LLLT) is an effective complementary treatment, especially for anti-inflammation and wound healing in which dermis or mucus mast cells (MCs) are involved. In periphery, MCs crosstalk with neurons via purinergic signals and participate in various physiological and pathophysiological processes. Whether extracellular ATP, an important purine in purinergic signaling, of MCs and neurons could be modulated by irradiation remains unknown.

Salt and osmosensing: role of cytoplasmic hydrogel.

Osmotic perturbations, occurring frequently under physiological and pathological conditions, alter cell size/volume and function. To protect cellular homeostasis, cell osmo- and volume-sensing mechanisms activate volume compensatory processes. The plasma membrane plays a prominent role in cell volume regulation by mediating the selective transport of extra- and intracellular osmolytes.

Hemolysis is a primary ATP-release mechanism in human erythrocytes.

The hypothesis that regulated ATP release from red blood cells (RBCs) contributes to nitric oxide-dependent control of local blood flow has sparked much interest in underlying release mechanisms. Several stimuli, including shear stress and hypoxia, have been found to induce significant RBC ATP release attributed to activation of ATP-conducting channels. In the present study, we first evaluated different experimental approaches investigating stimulated RBC ATP release and quantifying hemolysis.