Endothelial deletion of EPH receptor A4 alters single-cell profile and Tie2/Akap12 signaling to preserve blood-brain barrier integrity.

Neurobiological consequences of traumatic brain injury (TBI) result from a complex interplay of secondary injury responses and sequela that mediates chronic disability. Endothelial cells are important regulators of the cerebrovascular response to TBI. Our work demonstrates that genetic deletion of endothelial cell (EC)-specific EPH receptor A4 (EphA4) using conditional EphA4f/f/Tie2-Cre and EphA4f/f/VE-Cadherin-CreERT2 knockout (KO) mice promotes blood-brain barrier (BBB) integrity and tissue protection, which correlates with improved motor function and cerebral blood flow recovery following controlled cortical impact (CCI) injury. scRNAseq of capillary-derived KO ECs showed increased differential gene expression of BBB-related junctional and actin cytoskeletal regulators, namely, A-kinase anchor protein 12, Akap12, whose presence at Tie2 clustering domains is enhanced in KO microvessels. Transcript and protein analysis of CCI-injured whole cortical tissue or cortical-derived ECs suggests that EphA4 limits the expression of Cldn5, Akt, and Akap12 and promotes Ang2. Blocking Tie2 using sTie2-Fc attenuated protection and reversed Akap12 mRNA and protein levels cortical-derived ECs. Direct stimulation of Tie2 using Vasculotide, angiopoietin-1 memetic peptide, phenocopied the neuroprotection. Finally, we report a noteworthy rise in soluble Ang2 in the sera of individuals with acute TBI, highlighting its promising role as a vascular biomarker for early detection of BBB disruption. These findings describe a contribution of the axon guidance molecule, EphA4, in mediating TBI microvascular dysfunction through negative regulation of Tie2/Akap12 signaling.

STING-Dependent Signaling in Microglia or Peripheral Immune Cells Orchestrates the Early Inflammatory Response and Influences Brain Injury Outcome.

While originally identified as an antiviral pathway, recent work has implicated that cyclic GMP-AMP-synthase-Stimulator of Interferon Genes (cGAS-STING) signaling is playing a critical role in the neuroinflammatory response to traumatic brain injury (TBI). STING activation results in a robust inflammatory response characterized by the production of inflammatory cytokines called interferons, as well as hundreds of interferon stimulated genes (ISGs). Global knock-out (KO) mice inhibiting this pathway display neuroprotection with evidence that this pathway is active days after injury; yet, the early neuroinflammatory events stimulated by STING signaling remain understudied. Furthermore, the source of STING signaling during brain injury is unknown. Using a murine controlled cortical impact (CCI) model of TBI, we investigated the peripheral immune and microglial response to injury utilizing male chimeric and conditional STING KO animals, respectively. We demonstrate that peripheral and microglial STING signaling contribute to negative outcomes in cortical lesion volume, cell death, and functional outcomes postinjury. A reduction in overall peripheral immune cell and neutrophil infiltration at the injury site is STING dependent in these models at 24 h. Transcriptomic analysis at 2 h, when STING is active, reveals that microglia drive an early, distinct transcriptional program to elicit proinflammatory genes including interleukin 1-ß (IL-1ß), which is lost in conditional knock-out mice. The upregulation of alternative innate immune pathways also occurs after injury in these animals, which supports a complex relationship between brain-resident and peripheral immune cells to coordinate the proinflammatory response and immune cell influx to damaged tissue after injury.

Immunoregulatory and neutrophil-like monocyte subsets with distinct single-cell transcriptomic signatures emerge following brain injury.

Monocytes represent key cellular elements that contribute to the neurological sequela following brain injury. The current study reveals that trauma induces the augmented release of a transcriptionally distinct CD115+/Ly6Chi monocyte population into the circulation of mice pre-exposed to clodronate depletion conditions. This phenomenon correlates with tissue protection, blood-brain barrier stability, and cerebral blood flow improvement. Uniquely, this shifted the innate immune cell profile in the cortical milieu and reduced the expression of pro-inflammatory Il6, IL1r1, MCP-1, Cxcl1, and Ccl3 cytokines. Monocytes that emerged under these conditions displayed a morphological and gene profile consistent with a subset commonly seen during emergency monopoiesis. Single-cell RNA sequencing delineated distinct clusters of monocytes and revealed a key transcriptional signature of Ly6Chi monocytes enriched for Apoe and chitinase-like protein 3 (Chil3/Ym1), commonly expressed in pro-resolving immunoregulatory monocytes, as well as granule genes Elane, Prtn3, MPO, and Ctsg unique to neutrophil-like monocytes. The predominate shift in cell clusters included subsets with low expression of transcription factors involved in monocyte conversion, Pou2f2, Na4a1, and a robust enrichment of genes in the oxidative phosphorylation pathway which favors an anti-inflammatory phenotype. Transfer of this monocyte assemblage into brain-injured recipient mice demonstrated their direct role in neuroprotection. These findings reveal a multifaceted innate immune response to brain injury and suggest targeting surrogate monocyte subsets may foster tissue protection in the brain.

Ordered Van der Waals Hetero-nanoribbon from Pressure-Induced Topochemical Polymerization of Azobenzene.

Pressure-induced topochemical polymerization of molecular crystals with various stackings is a promising way to synthesize materials with different co-existing sub-structures. Here, by compressing the azobenzene crystal containing two kinds of intermolecular stacking, we synthesized an ordered van der Waals carbon nanoribbon (CNR) heterostructure in one step. Azobenzene polymerizes via a [4 + 2] hetero-Diels-Alder (HDA) reaction of phenylazo-phenyl in layer A and a para-polymerization reaction of phenyl in layer B at 18 GPa, as evidenced by in situ Raman and IR spectroscopies, X-ray diffraction, as well as gas chromatography-mass spectrometry and the solid-state nuclear magnetic resonance of the recovered products. The theoretical calculation shows that the obtained CNR heterostructure has a type II (staggered) band gap alignment. Our work highlights a high-pressure strategy to synthesize bulk CNR heterostructures.

Efferocytosis is restricted by axon guidance molecule EphA4 via ERK/Stat6/MERTK signaling following brain injury.

BACKGROUND: Efferocytosis is a process that removes apoptotic cells and cellular debris. Clearance of these cells alleviates neuroinflammation, prevents the release of inflammatory molecules, and promotes the production of anti-inflammatory cytokines to help maintain tissue homeostasis. The underlying mechanisms by which this occurs in the brain after injury remain ill-defined. METHODS: We used GFP bone marrow chimeric knockout (KO) mice to demonstrate that the axon guidance molecule EphA4 receptor tyrosine kinase is involved in suppressing MERTK in the brain to restrict efferocytosis of resident microglia and peripheral-derived monocyte/macrophages. RESULTS: Single-cell RNAseq identified MERTK expression, the primary receptor involved in efferocytosis, on monocytes, microglia, and a subset of astrocytes in the damaged cortex following brain injury. Loss of EphA4 on infiltrating GFP-expressing immune cells improved functional outcome concomitant with enhanced efferocytosis and overall protein expression of p-MERTK, p-ERK, and p-Stat6. The percentage of GFP+ monocyte/macrophages and resident microglia engulfing NeuN+ or TUNEL+ cells was significantly higher in KO chimeric mice. Importantly, mRNA expression of Mertk and its cognate ligand Gas6 was significantly elevated in these mice compared to the wild-type. Analysis of cell-specific expression showed that p-ERK and p-Stat6 co-localized with MERTK-expressing GFP + cells in the peri-lesional area of the cortex following brain injury. Using an in vitro efferocytosis assay, co-culturing pHrodo-labeled apoptotic Jurkat cells and bone marrow (BM)-derived macrophages, we demonstrate that efferocytosis efficiency and mRNA expression of Mertk and Gas6 was enhanced in the absence of EphA4. Selective inhibitors of ERK and Stat6 attenuated this effect, confirming that EphA4 suppresses monocyte/macrophage efferocytosis via inhibition of the ERK/Stat6 pathway. CONCLUSIONS: Our findings implicate the ERK/Stat6/MERTK axis as a novel regulator of apoptotic debris clearance in brain injury that is restricted by peripheral myeloid-derived EphA4 to prevent the resolution of inflammation.

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 (Pr2+) and ATP4-. Under isotonic conditions, we detected with quantitative fluorescence microscopy, a continuous low-rate uptake of Pr (∼150 × 10-21 moles (zmol)/h/cell, [Pr]o = 150 µM, 32°C). It was stimulated transiently but strongly by 66% hypotonic cell swelling reaching an influx amplitude of ∼1500 (zmol/h)/cell. The progressive Pr uptake with increasing [Pr]o (30, 150, and 750 µM) suggested a permeation mechanism by simple diffusion. We quantified separately ATP release with custom wide-field-of-view chemiluminescence imaging. The strong proportionality between ATP efflux and Pr2+ influx during hypotonic challenge, and the absence of stimulation of transmembrane transport following 300% hypertonic shock, indicated that ATP and Pr travel the same conductive pathway. The fluorescence images revealed a homogeneously distributed intracellular uptake of Pr not consistent with high-conductance channels expressed at low density on the plasma membrane. We hypothesized that the pathway consists of transiently formed water pores evenly spread across the plasma membrane. The abolition of cell swelling-induced Pr uptake with 500 µM gadolinium, a known modulator of membrane fluidity, supported the involvement of water pores whose formation depends on the membrane fluidity. Our study suggests an alternative model of a direct permeation of ATP (and other molecules) through the phospholipid bilayer, which may have important physiological implications.

Observing a Zeolite Nucleus (Subcrystal) with a Uniform Framework Structure and Its Oriented Attachment without Single-Molecule Addition.

Multiple and complex crystallization process of zeolite including complementary single-molecule condensation and particle assembly, and alternately dominant nucleation and growth behavior, plays the critical role in zeolite crystallization but meanwhile makes us hard to study the respective effects. Herein, we strip nuclei from the synthetic solution and find that high-ordered nucleus (subcrystal) is the premise to ignite high-speed growth of zeolite crystal. The high-ordered subcrystals with the size of only 6-10 nm possess regular aperture structure and microporous area similar to zeolite nanocrystal. Interestingly, a unitary oriented aggregation process of the subcrystals towards nanosheets is well observed and characterized where single-molecule addition process is greatly repressed. If a wider range of zeotype nuclei can be expanded, a new synthetic strategy of zeotype materials with heterogeneous framework and active sites may be expected, which may novelize zeolite catalytic properties.

Distance-Selected Topochemical Dehydro-Diels-Alder Reaction of 1,4-Diphenylbutadiyne toward Crystalline Graphitic Nanoribbons.

Solid-state topochemical polymerization (SSTP) is a promising method to construct functional crystalline polymeric materials, but in contrast to various reactions that happen in solution, only very limited types of SSTP reactions are reported. Diels-Alder (DA) and dehydro-DA (DDA) reactions are textbook reactions for preparing six-membered rings in solution but are scarcely seen in solid-state synthesis. Here, using multiple cutting-edge techniques, we demonstrate that the solid 1,4-diphenylbutadiyne (DPB) undergoes a DDA reaction under 10-20 GPa with the phenyl as the dienophile. The crystal structure at the critical pressure shows that this reaction is "distance-selected". The distance of 3.2 Å between the phenyl and the phenylethynyl facilitates the DDA reaction, while the distances for other DDA and 1,4-addition reactions are too large to allow the bonding. The obtained products are crystalline armchair graphitic nanoribbons, and hence our studies open a new route to construct the crystalline carbon materials with atomic-scale control.

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. Stretch (1 s, 5-35%) induced ATP release from AT2/AT1 cell cultures, and it was highest on days 2 and 3 but declined in older cultures. ATP release tightly correlated with the number of remaining AT2 cells in culture, consistent with ~10-fold lower ATP release by AT1 than AT2 cells. ATP release was unaffected by inhibitors of putative ATP channels carbenoxolone and probenecid but was significantly diminished in cells loaded with calcium chelator BAPTA. These pharmacological modulators had similar effects on stretch-induced intracellular Ca2+ responses measured by Fura2 fluorescence. The study revealed that AT2 cells are the primary source of stretch-induced ATP release in heterocellular AT2/AT1 cell cultures, suggesting similar contribution in intact alveoli. Our results support a role for calcium-regulated mechanism but not ATP-conducting channels in ATP release by alveolar epithelial cells.

Effect of icariin on early ß-defensin-2 and T cell subsets in rats after tracheotomy. / ·ARTICLES··è®º 著·.

OBJECTIVES: To investigate the effect of icariin (ICA) on early ß-defensin-2 and T cell subsets in rats after tracheotomy. METHODS: A total of 54 SPF male Sprague-Dawley rats were randomly divided into a normal control group (group A), a model group (group B), and a model+ICA treatment group (group C), with 18 rats in each group. A tracheotomy intubation model of the B and C group was prepared. After 6 h of surgery, ICA intervention was given to group C. Groups A and B were given the same amount of normal saline. Lung tissue, alveolar lavage fluid and peripheral blood were taken at 24 h, 72 h and 168 h, respectively. The expression of rat ß-defensin-2 mRNA in lung tissue was detected by RT-PCR. The content of ß-defensin-2 in alveolar lavage fluid and peripheral blood serum was detected by ELISA. The content of peripheral blood T cell subsets (CD3+, CD4+, CD8+) was detected by flow cytometry, and the ratio of CD4+/CD8+ was calculated. RESULTS: After tracheotomy, the levels of ß-defensin-2 mRNA and ß-defensin-2 in lung tissue from the group B were increased significantly at 24 h, then they were decreased gradually, and decreased most significantly at 168 h (P<0.05). The content of ß-defensin-2 in peripheral blood of group B decreased gradually, and the content of ß-defensin-2 in 168 h was significantly lower than that in 24 h (P<0.05), but there was no significant difference between group B and group A (P>0.05). The level of CD3+ T cells in peripheral blood was significantly lower than that in the group A (P<0.05), but their was no significant difference in CD4+ and CD8+ T cells compared with group A (P>0.05). After ICA intervention in group C: lung tissue, alveolar lavage fluid, peripheral blood serum ß-defensin-2 content, and peripheral blood CD3+ and CD4+ T cell levels were gradually increased, significantly higher than those in the group B (P<0.05). CD8+ T cell level was significantly lower than that in the group A at 24 h (P<0.05), the CD4+/CD8+ ratio was significantly higher at 168 h than those in the group A or B (both P<0.01). CONCLUSIONS: ICA can improve the early lung immune function in rats with tracheotomy, which might be related to up-regulation of ß-defensin-2 in lung tissue and alveolar lavage fluid, concomitant with increases in CD3+ and CD4+ T cells and CD4+/CD8+ ratio in peripheral blood while reduction in CD8+ cells.

Facile Synthesis of Hierarchical Nanosized Single-Crystal Aluminophosphate Molecular Sieves from Highly Homogeneous and Concentrated Precursors.

The synthesis of hierarchical nanosized zeolite materials without growth modifiers and mesoporogens remains a substantial challenge. Herein, we report a general synthetic approach to produce hierarchical nanosized single-crystal aluminophosphate molecular sieves by preparing highly homogeneous and concentrated precursors and heating at elevated temperatures. Accordingly, aluminophosphate zeotypes of LTA (8-rings), AEL (10-rings), AFI (12-rings), and -CLO (20-rings) topologies, ranging from small to extra-large pores, were synthesized. These materials show exceptional properties, including small crystallites (30-150 nm), good monodispersity, abundant mesopores, and excellent thermal stability. A time-dependent study revealed a non-classical crystallization pathway by particle attachment. This work opens a new avenue for the development of hierarchical nanosized zeolite materials and understanding their crystallization mechanism.

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-1) or swelled by hypotonic shock. The amount of ATP secreted in response to a series of five graded stretch pulses (5-37% linear deformation, 1-s duration at 25°C) changed nonmonotonically with respect to strain amplitude and was inhomogeneous across the cell monolayer. In a typical experiment, extracellular ATP density averaged 250 fmol/mm2, but the area of detectable signal covered only ∼40% of the cells. In some areas, ATP accumulation peaked around 900 fmol/mm2, which corresponded to an estimated concentration of 4.5 µM. The total amount of ATP released from the combined stretch pulses reached 384 ± 224 pmol/million cells (n = 4). Compared with stretch, hypotonic shock (50%, 30°C) elicited a more homogeneous ATP secretion from the entire cell population but at a lower yield totaling 28 ± 12 pmol/million cells (n = 4). The quantitative extracellular ATP mapping of several thousand cells at once, with this wide field of view imaging system, will help identify ATP release pathways by providing unique insights on the dynamics and inhomogeneities of the cellular ATP secretion that are otherwise difficult to assess within the smaller field of view of a microscope.

Two-dimensional Acetate-based Light Lanthanide Fluoride Nanomaterials (F-Ln, Ln = La, Ce, Pr, and Nd): Morphology, Structure, Growth Mechanism, and Stability.

Discovery of novel two-dimensional (2D) materials is of fundamental importance but remains challenging. In this work, we design a simple and facile bottom-up approach to fabricate a new family of 2D acetate-based light lanthanide fluoride nanomaterials (F-Ln, Ln = La, Ce, Pr, Nd) at room temperature and atmosphere pressure, for the first time. Various characterization techniques confirm that as-synthesized F-Ln exhibit an ultrathin morphology with thickness up to 1.45 nm and lateral dimensions up to several hundred nanometers. Microstructure analysis demonstrates that F-Ln are a series of defect-rich 2D nanomaterials, which consist of nanocrystals with sub-10 nm domains. Structure characterization of F-Ce, a typical example, infers that BN-like F-Ce one-atom-layers sandwiched by intercalated acetate anions stack alternately along [001] direction to form nanocrystal building blocks of F-Ce. The study of growth mechanism suggests that three procedures are involved in the formation of F-Ce: hydrolysis reaction of cerium(III) acetate, structure transformation induced by fluorine ions, and assembly process guided by acetate anions. The as-prepared nanosheets show excellent stability with respect to environment stimuli such as air, heat, solvent, and high-energy electron beam. This study enriches the library of 2D materials and paves the way for future application of such 2D materials in areas such as catalysis, adsorption, separation, and energy storage/conversion.

Assessment of prior opioid tolerance among new users of fentanyl transdermal system in FDA's Sentinel System.

PURPOSE: Fentanyl transdermal system (FTS) is intended only for patients with prior opioid tolerance. The purpose of this study is to identify the proportion of new FTS users who had evidence of prior opioid tolerance, by dosage strength, in FDA's Sentinel System. METHODS: We identified new FTS episodes (183-day washout) from 2009 through 2013. Members were <65 years and enrolled in medical and pharmacy coverage for 183 days prior to initial FTS dispensing (index). We assessed the proportion of users with prior tolerance stratified by dosage strength of FTS using four definitions of opioid tolerance: ≥30-mg oxycodone equivalents/day in each of 7 consecutive days immediately prior to index; ≥30-mg oxycodone equivalents/day for any 7 days in the 30 days prior to index (secondary); any dose in each of 7 days in the 7 consecutive days immediately prior to index (tertiary); and any dose for any 7 days in the 30 days prior to index (quaternary). RESULTS: Of 44 450 episodes of 25 mcg/hr FTS, 37% met the primary definition, and 77% met the quaternary definition. Of 3507 episodes of 100 mcg/hr FTS, 57% and 74% met the primary and quaternary definitions, respectively. Those aged 25 to 34 years had the highest proportion of episodes with prior tolerance; those aged 55 to 64 accounted for more of the episodes overall. CONCLUSIONS: In Sentinel, many new users of FTS did not have evidence of prior opioid tolerance by the primary definition, ie, the product label definition, which is the minimum standard for the lowest FTS dose (12 mcg/hr), especially at the highest strength (100 mcg/hr). Validation of this metric is warranted, but our findings suggest the need for further prescriber education regarding appropriate prescribing of FTS.

A new analytic tool developed to assess safe use recommendations.

PURPOSE: Develop a flexible analytic tool for the Food and Drug Administration's (FDA's) Sentinel System to assess adherence to safe use recommendations with two capabilities: characterize adherence to patient monitoring recommendations for a drug, and characterize concomitant medication use before, during, and/or after drug therapy. METHODS: We applied the tool in the Sentinel Distributed Database to assess adherence to the labeled recommendation that patients treated with dronedarone undergo electrocardiogram (ECG) testing no less often than every 3 months. Measures of length of treatment, time to first ECG, number of ECGs, and time between ECGs were assessed. We also assessed concomitant use of contraception among female users of mycophenolate per label recommendations (concomitancy 4 weeks before through 6 weeks after discontinuation of mycophenolate). Unadjusted results were stratified by age, month-year, and sex. RESULTS: We identified 21 457 new episodes of dronedarone use of greater than or equal to 90 days (July 2009 to September 2015); 86% had greater than or equal to one ECG, and 22% met the recommendation of an ECG no less often than every 3 months. We identified 21 942 new episodes of mycophenolate use among females 12 to 55 years (January 2016 to September 2015); 16% had greater than or equal to 1 day of concomitant contraception dispensed, 12% had concomitant contraception use for greater than or equal to 50% of the 4 weeks before initiation through 6 weeks after mycophenolate; younger females had more concomitancy. These results may be underestimates as the analyses are limited to claims data. CONCLUSIONS: We developed a tool for use in databases formatted to the Sentinel Common Data Model that can assess adherence to safe use recommendations involving patient monitoring and concomitant drug use over time.

Salidroside protected against MPP+ -induced Parkinson's disease in PC12 cells by inhibiting inflammation, oxidative stress and cell apoptosis.

The present study aimed to investigate the protective effects of salidroside (SAL) on 1-methyl-4-phenylpyridinium (MPP+ )-induced PC12 cell model for Parkinson's disease. PC12 cells were pretreated with SAL in different concentrations and then exposed to MPP+ . To evaluate the effects of SAL on cytotoxicity, the survival rate was tested by the 3-(4,5-dimethylthiazol-2-yl)-2,5-dimethyltetrazolium bromide (MTT) assay and the apoptosis was tested via flow cytometry and Western blot. Reactive oxygen species (ROS), glutathione (GSH), and malondialdehyde (MDA) were detected to analyze the effects of SAL on oxidative stress. The mRNA and protein levels of inflammatory factors TNF-α and IL-1ß were also determined by real-time quantitative polymerase chain reaction and Western blot. Pretreatment with SAL effectively relieved the MPP+ cytotoxic effects and decreased the release of ROS production and inflammatory cytokines. SAL also inhibited apoptosis, suppressed MDA activity, and increased GSH levels in MPP+ -treated PC12 cells. Moreover, the expression levels of caspase-9, caspase-3, and Bax were significantly decreased in the SAL treatment groups compared with the MPP+ group, whereas Bcl-2 expression was significantly increased in the SAL treatment groups. In summary, the overall results suggested that SAL have neuroprotective effects on the MPP+ -induced PC12 cell model by inhibiting inflammation, oxidative stress, and cell apoptosis. SAL may be a potential active product to protect against Parkinson's disease.

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. The present review intends to provide the current state of knowledge of the mechanisms of ATP release in the lung. Putative pathways of the release, including the contribution of cell membrane injury and cell lysis are discussed addressing their strength, weaknesses and missing evidence that requires future study. We also provide an overview of the recent technical advances in studying cellular ATP release in vitro and ex vivo. Special attention is given to new insights into lung ATP release obtained with the real-time luminescence ATP imaging. This includes recent data on stretch-induced mechanosensitive ATP release in a model and primary cells of lung alveoli in vitro as well as inflation-induced ATP release in airspaces and pulmonary blood vessels of lungs, ex vivo.

Transcriptomic and Co-Expression Network Profiling of Shoot Apical Meristem Reveal Contrasting Response to Nitrogen Rate between Indica and Japonica Rice Subspecies.

Reducing nitrogen (N) input is a key measure to achieve a sustainable rice production in China, especially in Jiangsu Province. Tiller is the basis for achieving panicle number that plays as a major factor in the yield determination. In actual production, excessive N is often applied in order to produce enough tillers in the early stages. Understanding how N regulates tillering in rice plants is critical to generate an integrative management to reduce N use and reaching tiller number target. Aiming at this objective, we utilized RNA sequencing and weighted gene co-expression network analysis (WGCNA) to compare the transcriptomes surrounding the shoot apical meristem of indica (Yangdao6, YD6) and japonica (Nipponbare, NPB) rice subspecies. Our results showed that N rate influenced tiller number in a different pattern between the two varieties, with NPB being more sensitive to N enrichment, and YD6 being more tolerant to high N rate. Tiller number was positively related to N content in leaf, culm and root tissue, but negatively related to the soluble carbohydrate content, regardless of variety. Transcriptomic comparisons revealed that for YD6 when N rate enrichment from low (LN) to medium (MN), it caused 115 DEGs (LN vs. MN), from MN to high level (HN) triggered 162 DEGs (MN vs. HN), but direct comparison of low with high N rate showed a 511 DEGs (LN vs. HN). These numbers of DEG in NPB were 87 (LN vs. MN), 40 (MN vs. HN), and 148 (LN vs. HN). These differences indicate that continual N enrichment led to a bumpy change at the transcription level. For the reported sixty-five genes which affect tillering, thirty-six showed decent expression in SAM at tiller starting phase, among them only nineteen being significantly influenced by N level, and two genes showed significant interaction between N rate and variety. Gene ontology analysis revealed that the majority of the common DEGs are involved in general stress responses, stimulus responses, and hormonal signaling process. WGCNA network identified twenty-two co-expressing gene modules and ten candidate hubgenes for each module. Several genes associated with tillering and N rate fall on the related modules. These indicate that there are more genes participating in tillering regulation in response to N enrichment.

Ca2 PbGa8 O15 : Rational Design, Synthesis, and Structure Determination of a Purely Tetrahedra-Based Intergrowth Oxide.

Intergrowth oxides, like Aurivillius, Ruddlesden-Popper phase, comprise functional layers and exhibit interesting physical properties. The hitherto known intergrowth structures mainly were composed of closed-packing of oxygen ions, and it is very challenging to develop new types of intergrowth structures. We proposed the possible match between the tridymite and grossite, both of which are purely tetrahedra-based structures. We synthesized Ca2 PbGa8 O15 ((Ca0.5 Pb0.5 Ga2 O4 )2 (CaGa4 O7 )) and its structure was solved by ab-initio method. Pb2+ is vitally important to stabilize this first example of tetrahedra-based intergrowth oxide. The appropriate size difference between Pb2+ and Ca2+ causes the layered type cationic ordering, and reduced the thermodynamic potential, in addition, the high hybridization between Pb 6s6p and O 2p orbitals further consolidate the covalency of the tetrahedra-base framework.

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 Ca2+ through activation of the store-operated calcium entry (SOCE) involving an inositol-3-phosphate receptor (IP3R)-coupled depletion of endoplasmic reticular Ca2+ and a subsequent activation of the stromal interaction molecule 1 (STIM-1)/Orai-1 complex. However, the involvement of IP3R/STIM-1/Orai-1-Ca2+ -dependent signaling in Egr-1 expression in VSMC remains unexplored. Therefore, in the present studies, we have examined the role of Ca2+ signaling in Ang-II-induced Egr-1 expression in VSMC and investigated the contribution of STIM-1 or Orai-1 in mediating this response. 2-aminoethoxydiphenyl borate (2-APB), a dual non-competitive antagonist of IP3R and inhibitor of SOCE, decreased Ang-II-induced Ca2+ release and attenuated Ang-II-induced enhanced expression of Egr-1 protein and mRNA levels. Egr-1 upregulation was also suppressed following blockade of calmodulin and CaMKII. Furthermore, RNA interference-mediated depletion of STIM-1 or Orai-1 attenuated Ang-II-induced Egr-1 expression as well as Ang-II-induced phosphorylation of ERK1/2 and CREB. In addition, siRNA-induced silencing of CREB resulted in a reduction in the expression of Egr-1 stimulated by Ang-II. In summary, our data demonstrate that Ang-II-induced Egr-1 expression is mediated by STIM-1/Orai-1/Ca2+ -dependent signaling pathways in A-10 VSMC.