Modeling Fertilization Outcome in a Changing World.

Marine organisms have complex life histories. For broadcast spawners, successful continuation of the population requires their small gametes to make contact in the water column for sufficiently long periods for fertilization to occur. Anthropogenic climate change has been shown to impact fertilization success in various marine invertebrates, including sea urchins which are key grazers in their habitats. Gamete performance of both sexes declined when exposed to elevated temperature and/or pCO2 levels. Examples of reduced performance included slower sperm swimming speed and thinning egg jelly coat. However, such responses to climate change stress were not uniform between individuals. Such variations could serve as the basis for selection. Fertilization kinetics has long been modeled as a particle collision process. Here, we present a modified fertilization kinetics model that incorporates individual variations in performance in a more environmentally-relevant regime, and which the performance of groups with different traits can be separately tracked in a mixture. Numerical simulations highlight that fertilization outcome is influenced by changes in gametes traits as they age in sea water and the presence of competition groups (multiple dams or sires). These results highlight the importance of considering multiple individuals and at multiple time points during in-vivo assays. We also applied our model to show that interspecific variation in climate stress vulnerabilities elevates the risk of hybridization. By making a numerical model open-source, we aim to help us better understand the fate of organisms in the face of climate change by enabling the community to consider the mean and variance of the response to capture adaptive potential.

Demonstration of optically-assisted reconfigurable average of two 20-Gbaud 4-phase-encoded data channels using nonlinear wave mixing.

Networks can play a key role in high-speed and reconfigurable arithmetic computing. However, two performance bottlenecks may arise when: (i) relying solely on electronics to handle computation for multiple data channels at high data rates, and (ii) the data streams input to a processing node (PN) are transmitted as phase-encoded signals over an optical network. We experimentally demonstrate the operation of optically-assisted reconfigurable average of two 4-phase-encoded data channels at 10- and 20-Gbaud rates. Our input signals are two streams of 2-bit numbers representing a binary floating-point format, and the operation results in 7-phase-encoded output signals represented by 3-bit numbers. The average operation is achieved in three stages: (1) phase encoding and division-using an optical modulator to encode the data streams; (2) summation-using a highly nonlinear fiber (HNLF); and (3) multicast-using a periodically poled lithium niobate (PPLN) waveguide to multicast back the result into the original signal wavelengths. The experimental results validate the concept, and the measured penalties indicate that: (i) the error vector magnitudes (EVMs) of optical signals increase at each stage and reach ∼18-21% for the final multicast results, and (ii) compared to the inputs, the optical signal-to-noise ratio (OSNR) penalty of output is ∼6.7 dB for the 10-Gbaud rate and ∼6.9 dB for the 20-Gbaud rate at a bit error rate (BER) of 3.8e-3.

Simultaneous interrogation of multiple cores in a shape sensor fiber with a graded index fiber micro-turnaround.

A critical limitation for optical fiber sensor technology is the complexity of the interrogators used in such measurements, which has driven continued interest in enhanced optical fibers and fiber assemblies that will simplify interrogator design. In this work, we report on a novel multicore fiber shape sensor utilizing a distal graded index (GRIN) fiber micro-turnaround. We show that four offset cores of this fiber can be interrogated simultaneously with a single high performance optical frequency domain reflectometry measurement. The GRIN turnaround is 498 µm in length and reflects signal from one offset core to an opposite core with a 2 dB roundtrip attenuation. We show that the bend sensing accuracy of our single measurement system is similar to the accuracy of sequential measurements of four individual cores. We also demonstrate fiber shape reconstruction with a single measurement over 0.55 m with 80 µm spatial resolution when the fiber is wrapped around two posts.

Role of Carbon Monoxide in Oxidative Stress-Induced Senescence in Human Bronchial Epithelium.

Prolonged or excessive stimulation from inhaled toxins may cause oxidative stress and DNA damage that can lead to stress-induced senescence in epithelial cells, which can contribute to several airway diseases. Mounting evidence has shown carbon monoxide (CO) confers cytoprotective effects. We investigated the effects of CO on oxidative stress-induced senescence in human airway epithelium and elucidated the underlying molecular mechanisms. Here, CO pretreatment reduced H2O2-mediated increases in total reactive oxygen species (ROS) production and mitochondrial superoxide in a human bronchial epithelial cell line (BEAS-2B). H2O2 treatment triggered a premature senescence-like phenotype with enlarged and flattened cell morphology accompanied by increased SA-ß-gal activity, cell cycle arrest in G0/G1, reduced cell viability, and increased transcription of senescence-associated secretory phenotype (SASP) genes. Additionally, exposure to H2O2 increased protein levels of cellular senescence markers (p53 and p21), reduced Sirtuin 3 (SIRT3) and manganese superoxide dismutase (MnSOD) levels, and increased p53 K382 acetylation. These H2O2-mediated effects were attenuated by pretreatment with a CO-containing solution. SIRT3 silencing induced mitochondrial superoxide production and triggered a senescence-like phenotype, whereas overexpression decreased mitochondrial superoxide production and alleviated the senescence-like phenotype. Air-liquid interface (ALI) culture of primary human bronchial cells, which becomes a fully differentiated pseudostratified mucociliary epithelium, was used as a model. We found that apical and basolateral exposure to H2O2 induced a vacuolated structure that impaired the integrity of ALI cultures, increased goblet cell numbers, decreased SCGB1A1+ club cell numbers, increased p21 protein levels, and increased SASP gene transcription, consistent with our observations in BEAS-2B cells. These effects were attenuated in the apical presence of a CO-containing solution. In summary, we revealed that CO has a pivotal role in epithelial senescence by regulating ROS production via the SIRT3/MnSOD/p53/p21 pathway. This may have important implications in the prevention and treatment of age-associated respiratory pathologies.

TM9SF4 Is a Crucial Regulator of Inflammation and ER Stress in Inflammatory Bowel Disease.

BACKGROUND & AIMS: Inflammatory bowel disease (IBD) is a major intestinal disease. Excessive inflammation and increased endoplasmic reticulum (ER) stress are the key events in the development of IBD. Search of a genome-wide association study database identified a remarkable correlation between a TM9SF4 single-nucleotide polymorphism and IBD. Here, we aimed to resolve its underlying mechanism. METHODS: The role of TM9SF4 was determined with experimental mouse models of IBD. ER stress cascades, barrier functions, and macrophage polarization in colonic tissues and cells were assessed in vivo and in vitro. The expression of TM9SF4 was compared between inflamed regions of ulcerative colitis patients and normal colon samples. RESULTS: In mouse models of IBD, genetic knockout of the TM9SF4 gene aggravated the disease symptoms. In colonic epithelial cells, short hairpin RNA-mediated knockdown of TM9SF4 expression promoted inflammation and increased ER stress. In macrophages, TM9SF4 knockdown promoted M1 macrophage polarization but suppressed M2 macrophage polarization. Genetic knockout/knockdown of TM9SF4 also disrupted epithelial barrier function. Mechanistically, TM9SF4 deficiency may act through Ca2+ store depletion and cytosolic acidification to induce an ER stress increase. Furthermore, the expression level of TM9SF4 was found to be much lower in the inflamed colon regions of human ulcerative colitis patients than in normal colon samples. CONCLUSIONS: Our study identified a novel IBD-associated protein, TM9SF4, the reduced expression of which can aggravate intestinal inflammation. Deficiency of TM9SF4 increases ER stress, promotes inflammation, and impairs the intestinal epithelial barrier to aggravate IBD.

Meteorin-ß/Meteorin like/IL-41 attenuates airway inflammation in house dust mite-induced allergic asthma.

We sought to examine the regulatory effect of Meteorin-ß (Metrnß)/Meteorin like (Metrnl)/IL-41 on lung inflammation in allergic asthma. We found that Metrnß was elevated significantly in asthmatic patients and in mice with allergic asthma induced by house dust mite (HDM) extract. Upon exposure to HDM, Metrnß was secreted predominantly by airway epithelial cells and inflammatory cells, including macrophages and eosinophils. The increased Metrnß effectively blocked the development of airway hyperreactivity (AHR) and decreased inflammatory cell airway infiltration and type 2 cytokine production, which was associated with downregulated DC-mediated adaptive immune responses. Moreover, Metrnß impaired the maturation and function of bone marrow-derived dendritic cells in vitro. Asthmatic mice adoptively transferred with dendritic cells isolated from Metrnß-treated allergic mice displayed decreased AHR, airway inflammation, and lung injury. Metrnß also displayed anti-inflammatory properties in immunodeficient SCID mice with allergic asthma and in in vitro 3D ALI airway models. Moreover, blockade of Metrnß by anti-Metrnß antibody treatment promoted the development of allergic asthma. These results revealed the unappreciated protective roles of Metrnß in alleviating DC-mediated Th2 inflammation in allergic asthma, providing the novel treatment strategy of therapeutic targeting of Metrnß in allergic asthma.

ß2-Adrenoceptor Activation Stimulates IL-6 Production via PKA, ERK1/2, Src, and Beta-Arrestin2 Signaling Pathways in Human Bronchial Epithelia.

OBJECTIVE: ß2-Adrenoceptor agonists are widely used to treat asthma because of their bronchial-dilation effects. We previously reported that isoprenaline, via the apical and basolateral ß2-adrenoceptor, induced Cl- secretion by activating cyclic AMP (cAMP)-dependent pathways in human bronchial epithelia. Despite these results, whether and how the ß2-adrenoceptor-mediated cAMP-dependent pathway contributes to pro-inflammatory cytokine release in human bronchial epithelia remains poorly understood. METHODS: We investigated ß2-adrenoceptor-mediated signaling pathways involved in the production of two pro-inflammatory cytokines, interleukin (IL)-6 and IL-8, in 16HBE14o- human bronchial epithelia. The effects of isoprenaline or formoterol were assessed in the presence of protein kinase A (PKA), exchange protein directly activated by cAMP (EPAC), Src, and extracellular signal-regulated protein kinase (ERK)1/2 inhibitors. The involvement of ß-arrestin2 was examined using siRNA knockdown. RESULTS: Isoprenaline and formoterol (both ß2 agonists) induced IL-6, but not IL-8, release, which could be inhibited by ICI 118,551 (ß2 antagonist). The PKA-specific inhibitor, H89, partially inhibited IL-6 release. Another intracellular cAMP receptor, EPAC, was not involved in IL-6 release. Isoprenaline-mediated IL-6 secretion was attenuated by dasatinib, a Src inhibitor, and PD98059, an ERK1/2 inhibitor. Isoprenaline treatment also led to ERK1/2 phosphorylation. In addition, knockdown of ß-arrestin2 by siRNA specifically suppressed cytokine release when a high concentration of isoprenaline (1 mM) was used. CONCLUSION: Our results suggest that activation of the ß2-adrenoceptor in 16HBE14o- cells stimulated the PKA/Src/ERK1/2 and/or ß-arrestin2 signaling pathways, leading to IL-6 release. Therefore, our data reveal that ß2-adrenoceptor signaling plays a role in the immune regulation of human airway epithelia.

TRPM2 promotes autophagic degradation in vascular smooth muscle cells.

Transient receptor potential channel M2 (TRPM2) is a Ca2+-permeable channel that is activated by reactive oxygen species (ROS). In many cell types, ROS activate TRPM2 to induce excessive Ca2+ influx, resulting in Ca2+ overload and consequent cell death. Recent studies suggest that TRPM2 may also regulate autophagy in pericytes and cancer cells by acting on the early step of autophagy, i.e. autophagic induction. However, there is no report on the role of TRPM2 in autophagic degradation, which is the late stage of autophagy. In the present study, we found abundant TRPM2 expression in lysosomes/autolysosomes in the primary cultured mouse aortic smooth muscle cells (mASMCs). Nutrient starvation stimulated autophagic flux in mASMCs mainly by promoting autophagic degradation. This starvation-induced autophagic degradation was reduced by TRPM2 knockout. Importantly, starvation-induced lysosomal/autolysosomal acidification and cell death were also substantially reduced by TRPM2 knockout. Taken together, the present study uncovered a novel mechanism that lysosomal TRPM2 facilitates lysosomal acidification to stimulate excessive autolysosome degradation and consequent cell death.

Measuring the shape of microbends in optical fibers.

We report on the distributed shape measurement of small deformations produced along the length of an optical fiber. The fiber contains multiple waveguiding cores, each inscribed with weak continuous Bragg gratings. The distributed Bragg-reflectivity data for the fiber cores, obtained from the optical backscatter reflectometry, are used to estimate the local curvature and the position of the fiber. We successfully demonstrate the sensing of periodic microdeformations-approximately 1 µm or less in amplitude and a few hundred µm in length. Such microbends are known to cause attenuation in optical fibers, and the approach presented here can enable a detailed measurement of these microbends in applications ranging from telecommunications cable design to biotechnology, robotics, manufacturing, aerospace, and security.

ß2 adrenoceptor signaling regulates ion transport in 16HBE14o- human airway epithelial cells.

We investigated the regulation of Cl- secretion by adrenoceptors in polarized 16HBE14o- human bronchial epithelial cells. Treatment with the nonselective ß adrenoceptor agonist isoprenaline stimulated an increase in short-circuit current (ISC ), which was inhibited by the ß adrenoceptor blocker propranolol. Treatment with procaterol, an agonist specific for the ß2 adrenoceptor subtype, stimulated a similar increase in ISC , which was inhibited by the ß2 adrenoceptor antagonist ICI 118551. Inhibitors of cystic fibrosis transmembrane conductance regulator (CFTR) and calcium-activated Cl- channel (CaCC), but not K+ channel blockers, were able to inhibit the increase in ISC . "Trimultaneous" recording of ISC and intracellular cyclic adenosine monophosphate (cAMP) and Ca2+ levels in 16HBE14o- epithelia confirmed that the ISC induced by isoprenaline or procaterol involved both cAMP and Ca2+ signaling. Our results demonstrate that ß2 adrenoceptors regulate Cl- secretion in the human airway epithelium by activating apical CFTRs and CaCCs via cAMP-dependent and intracellular Ca2+ -dependent mechanisms, respectively.

Anti-inflammatory action of HO-1/CO in human bronchial epithelium in response to cationic polypeptide challenge.

Carbon monoxide (CO) is an anti-inflammatory gaseous molecule produced endogenously by heme oxygenases (HOs) HO-1 and HO-2. However, the mechanisms underlying the anti-inflammatory effects of CO in the human bronchial epithelium are still not fully understood. In this study, the cationic peptide poly-l-arginine (PLA) was utilized to induce bronchial epithelial damage and subsequent pro-inflammatory cytokine release in the human bronchial epithelial cell line 16HBE14o-. Expression of both HO-1 and HO-2 after PLA exposure was examined. The polarized secretion of two pro-inflammatory cytokines, interleukin (IL)-6 and IL-8, was determined by ELISA. The anti-inflammatory effects of CO liberated from CO-releasing molecules (CORMs) were examined by both ELISA and western blot analysis. Our results indicate that PLA exposure leads to upregulation of HO-1 expression and p65 NF-κB phosphorylation, as well as IL-6 and IL-8 release. HO-1 induction by hemin or CORMs significantly suppressed IL-6 and IL-8 release. In addition, HO-1 knockdown further increased IL-6 and IL-8 release under basal and PLA-stimulated conditions. Our results thereby demonstrate that the HO-1/CO axis exerts significant anti-inflammatory activity during bronchial epithelial damage caused by cationic protein.

Carbon Monoxide Inhibits Cytokine and Chloride Secretion in Human Bronchial Epithelia.

BACKGROUND/AIMS: Carbon monoxide (CO) is an important gas produced endogenously by heme oxygenase (HO) that functions as an anti-inflammatory and in ion channel modulation, but the effects of CO on airway inflammation and ion transport remains unclear. METHODS: The effect of CO on cell damage- and nucleotide-induced pro-inflammatory cytokine release in primary human bronchial epithelia cells (HBE) and in the 16HBE14o- human bronchial epithelial cell line were investigated. The effects of CO on calcium- and cAMP-dependent chloride (Cl-) secretion were examined using a technique that allowed the simultaneous measurement and quantification of real-time changes in signalling molecules (cAMP and Ca2+) and ion transport in a polarised epithelium. RESULTS: CO suppressed the release of interleukin (IL)-6 and IL-8 and decreased the phosphorylation of ERK1/2 and NF-κB p65. Furthermore, CO inhibited UTP-induced increases in calcium and Cl- secretion, and forskolin-induced increases in cAMP and Cl- secretion. CONCLUSIONS: These findings suggest a novel anti-inflammatory role of CO in human bronchial epithelia via interactions with purinergic signalling pathways. Further, CO modulated both the Ca2+- and cAMP-dependent secretion of Cl-.

Increased intracellular Cl- concentration promotes ongoing inflammation in airway epithelium.

Airway epithelial cells harbor the capacity of active Cl- transepithelial transport and play critical roles in modulating innate immunity. However, whether intracellular Cl- accumulation contributes to relentless airway inflammation remains largely unclear. This study showed that, in airway epithelial cells, intracellular Cl- concentration ([Cl-]i) was increased after Pseudomonas aeruginosa lipopolysaccharide (LPS) stimulation via nuclear factor-κB (NF-κB)-phosphodiesterase 4D (PDE4D)-cAMP signaling pathways. Clamping [Cl-]i at high levels or prolonged treatment with LPS augmented serum- and glucocorticoid-inducible protein kinase 1 (SGK1) phosphorylation and subsequently triggered NF-κB activation in airway epithelial cells, whereas inhibition of SGK1 abrogated airway inflammation in vitro and in vivo. Furthermore, Cl--SGK1 signaling pathway was pronouncedly activated in patients with bronchiectasis, a chronic airway inflammatory disease. Conversely, hydrogen sulfide (H2S), a sulfhydryl-containing gasotransmitter, confers anti-inflammatory effects through decreasing [Cl-]i via activation of cystic fibrosis transmembrane conductance regulator (CFTR). Our study confirms that intracellular Cl- is a crucial mediator of sustained airway inflammation. Medications that abrogate excessively increased intracellular Cl- may offer novel targets for the management of airway inflammatory diseases.

Regulation of Intracellular Calcium by Carbon Monoxide in Human Bronchial Epithelial Cells.

BACKGROUND/AIMS: Carbon monoxide (CO) is an important autocrine/paracrine messenger involved in a variety of physiological and pathological processes. This study aimed to investigate the regulatory role of CO released by CO-releasing molecule-2 (CORM-2) in a P2Y receptor-mediated calcium-signaling pathway in the human bronchial epithelial cell line, 16HBE14o-. METHODS: Intracellular calcium ([Ca2+]i) was measured by fura-2 microspectrofluorimetry. D-myo-inositol-1-phosphate (IP1) levels and cGMP-dependent protein kinase activity (PKG) were also quantified. RESULTS: The exogenous application of CORM-2 increased both intracellular Ca2+ and IP1, which are inhibited by U73122, a phospholipase C (PLC) inhibitor. In contrast, the P2Y2/P2Y4 receptor-mediated intracellular Ca2+ release and influx induced by UTP were inhibited in the presence of CORM-2. However, CORM-2 did not affect the store-operated Ca2+ entry (SOCE) induced by thapsigargin (Tg). Moreover, the inhibitory effect of CORM-2 on UTP-induced calcium increase could be attenuated by a soluble guanylyl cyclase (sGC) inhibitor, 1H-[1,2,4]oxadiazolo[4,3,-a]quinoxalin-1-one (ODQ), or a Protein Kinase G (PKG) inhibitor, KT5823, suggesting the involvement of sGC/PKG signaling in this process. CONCLUSION: CORM-2 serves a dual role in modulating [Ca2+]i in 16HBE14o- cells. Thus, CO released by CORM-2 may act as a regulator of calcium homeostasis in human airway epithelia. These findings help further elucidate the function of CO in many physiological and pathological conditions.

G protein-coupled estrogen receptor inhibits the P2Y receptor-mediated Ca(2+) signaling pathway in human airway epithelia.

P2Y receptor activation causes the release of inflammatory cytokines in the bronchial epithelium, whereas G protein-coupled estrogen receptor (GPER), a novel estrogen (E2) receptor, may play an anti-inflammatory role in this process. We investigated the cellular mechanisms underlying the inhibitory effect of GPER activation on the P2Y receptor-mediated Ca(2+) signaling pathway and cytokine production in airway epithelia. Expression of GPER in primary human bronchial epithelial (HBE) or 16HBE14o- cells was confirmed on both the mRNA and protein levels. Stimulation of HBE or 16HBE14o- cells with E2 or G1, a specific agonist of GPER, attenuated the nucleotide-evoked increases in [Ca(2+)]i, whereas this effect was reversed by G15, a GPER-specific antagonist. G1 inhibited the secretion of two proinflammatory cytokines, interleukin (IL)-6 and IL-8, in cells stimulated by adenosine 5'-(γ-thio)triphosphate (ATPγS). G1 stimulated a real-time increase in cAMP levels in 16HBE14o- cells, which could be inhibited by adenylyl cyclase inhibitors. The inhibitory effects of E2 or G1 on P2Y receptor-induced increases in Ca(2+) were reversed by treating the cells with a protein kinase A (PKA) inhibitor. These results demonstrated that the inhibitory effects of G1 or E2 on P2Y receptor-mediated Ca(2+) mobilization and cytokine secretion were due to GPER-mediated activation of a cAMP-dependent PKA pathway. This study has reported, for the first time, the expression and function of GPER as an anti-inflammatory component in human bronchial epithelia, which may mediate through its opposing effects on the pro-inflammatory pathway activated by the P2Y receptors in inflamed airway epithelia.

A small synthetic molecule functions as a chloride-bicarbonate dual-transporter and induces chloride secretion in cells.

A C2 symmetric small molecule composed of l-phenylalanine and isophthalamide was found to function as a Cl(-)/HCO3(-) dual transporter and self-assemble into chloride channels. In Ussing-chamber based short-circuit current measurements, this molecule elicited chloride-dependent short-circuit current (Isc) increase in both Calu-3 cell and CFBE41o-cell (with F508del mutant CFTR) monolayers.

Nobiletin Stimulates Chloride Secretion in Human Bronchial Epithelia via a cAMP/PKA-Dependent Pathway.

BACKGROUND/AIMS: Nobiletin, a citrus flavonoid isolated from tangerines, alters ion transport functions in intestinal epithelia, and has antagonistic effects on eosinophilic airway inflammation of asthmatic rats. The present study examined the effects of nobiletin on basal short-circuit current (I(SC)) in a human bronchial epithelial cell line (16HBE14o-), and characterized the signal transduction pathways that allowed nobiletin to regulate electrolyte transport. METHODS: The I(SC) measurement technique was used for transepithelial electrical measurements. Intracellular calcium ([Ca(2+)]i) and cAMP were also quantified. RESULTS: Nobiletin stimulated a concentration-dependent increase in I(SC), which was due to Cl- secretion. The increase in I(SC) was inhibited by a cystic fibrosis transmembrane conductance regulator inhibitor (CFTR(inh)-172), but not by 4,4'-diisothiocyano-stilbene-2,2'-disulphonic acid (DIDS), Chromanol 293B, clotrimazole, or TRAM-34. Nobiletin-stimulated I(SC) was also sensitive to a protein kinase A (PKA) inhibitor, H89, and an adenylate cyclase inhibitor, MDL-12330A. Nobiletin could not stimulate any increase in I(SC) in a cystic fibrosis (CF) cell line, CFBE41o-, which lacked a functional CFTR. Nobiletin stimulated a real-time increase in cAMP, but not [Ca(2+)]i. CONCLUSION: Nobiletin stimulated transepithelial Cl- secretion across human bronchial epithelia. The mechanisms involved activation of adenylate cyclase- and cAMP/PKA-dependent pathways, leading to activation of apical CFTR Cl- channels.

P2Y6 receptor-mediated proinflammatory signaling in human bronchial epithelia.

P2Y receptors are expressed in virtually all epithelia and are responsible for the control of fluid and electrolyte transport. In asthmatic inflammation, the bronchial epithelia are damaged by eosinophil-derived, highly toxic cationic proteins, such as major basic protein (MBP). Consequently, extracellular nucleotides are released into the extracellular space from airway epithelial cells, and act in an autocrine or paracrine fashion to regulate immune functions. Our data show damage to the human bronchial epithelial cell line, 16HBE14o-, by poly-L-arginine-induced UDP release into the extracellular medium. Activation of P2Y6 receptor by its natural ligand, UDP, or its specific agonist, MRS 2693, led to the production of two proinflammatory cytokines, interleukin (IL)-6 and IL-8. This may have resulted from increased IL-6 and IL-8 mRNA expression, and activation of p38 and ERK1/2 MAPK, and NF-κB pathways. Our previous study demonstrated that UDP stimulated transepithelial Cl- secretion via both Ca2+- and cAMP-dependent pathways in 16HBE14o- epithelia. This was further confirmed in this study by simultaneous imaging of Ca2+ and cAMP levels in single cells using the Fura-2 fluorescence technique and a FRET-based approach, respectively. Moreover, the P2Y6 receptor-mediated production of IL-6 and IL-8 was found to be dependent on Ca2+, but not the cAMP/PKA pathway. Together, these studies show that nucleotides released during the airway inflammatory processes will activate P2Y6 receptors, which will lead to further release of inflammatory cytokines. The secretion of cytokines and the formation of such "cytokine networks" play an important role in sustaining the airway inflammatory disease.

Purinergic P2Y receptors in airway epithelia: from ion transport to immune functions.

The regulated transport of salt and water is essential to the integrated function of many organ systems, including the respiratory, reproductive, and digestive tracts. Airway epithelial fluid secretion is a passive process that is driven by osmotic forces, which are generated by ion transport. The main determinant of a luminally-directed osmotic gradient is the mucosal transport of chloride ions (Cl(-)) into the lumen. As with many epithelial cells, a number of classic signal transduction cascades are involved in the regulation of ion transport. There are two well-known intracellular signaling systems: an increase in intracellular Ca(2+) concentration ([Ca(2+)]i) and an increase in the rate of synthesis of cyclic nucleotides, such as cyclic adenosine monophosphate (cAMP). Therefore, Cl(-) secretion is primarily activated via the opening of apical Ca(2+)- or cAMP-dependent Cl(-) channels at the apical membrane. The opening of basolateral Ca(2+)- or cAMP-activated K(+) channels, which hyperpolarizes the cell to maintain the driving force for Cl(-) exit through apical Cl(-) channels that are constitutively open, is also important in regulating transepithelial ion transport. P2Y receptors are expressed in the apical and/or basolateral membranes of virtually all polarized epithelia to control the transport of fluid and electrolytes. Human airway epithelial cells express multiple nucleotide receptors. Extracellular nucleotides, such as UTP and ATP, are calcium-mobilizing secretagogues. They are released into the extracellular space from airway epithelial cells and act on the same cell in an autocrine fashion to stimulate transepithelial ion transport. In addition, recent data support the role of P2Y receptors in releasing inflammatory cytokines in the bronchial epithelium and other immune cells.

GmSAL1 hydrolyzes inositol-1,4,5-trisphosphate and regulates stomatal closure in detached leaves and ion compartmentalization in plant cells.

Inositol polyphosphatases are important regulators since they control the catabolism of phosphoinositol derivatives, which are often signaling molecules for cellular processes. Here we report on the characterization of one of their members in soybean, GmSAL1. In contrast to the substrate specificity of its Arabidopsis homologues (AtSAL1 and AtSAL2), GmSAL1 only hydrolyzes inositol-1,4,5-trisphosphate (IP3) but not inositol-1,3,4-trisphosphate or inositol-1,4-bisphosphate.The ectopic expression of GmSAL1 in transgenic Arabidopsis thaliana led to a reduction in IP3 signals, which was inferred from the reduction in the cytoplasmic signals of the in vivo biomarker pleckstrin homology domain-green florescent protein fusion protein and the suppression of abscisic acid-induced stomatal closure. At the cellular level, the ectopic expression of GmSAL1 in transgenic BY-2 cells enhanced vacuolar Na(+) compartmentalization and therefore could partially alleviate salinity stress.