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Sample records for intercellular junction assembly

  1. Intercellular junctions in myriapods.

    PubMed

    Dallai, R; Bigliardi, E; Lane, N J

    1990-01-01

    Tissue from the intestinal tract of myriapods, including millipedes, centipedes and pauropods were examined in tracer-impregnated sections and freeze-fracture replicas. The foregut and hindgut of all three classes exhibit pleated septate junctions; these display undulating intercellular ribbons in thin sections. In replicas they show discrete intramembranous particle (IMP) arrays aligned in rows in parallel; with one another. The tissues of the hindgut also possess scalariform junctions, characterized by cross-striated intercellular clefts in sections and IMP-enriched membranes in replicas. Gap junctions occur in all groups, but they are atypical in replicas in that their component IMPs do not always fracture onto the E face, as is characteristic of other arthropods; some IMPs cleave to the P face and others to the E face. The midgut of these organisms exhibits smooth septate junctions with conventional straight septal ribbons and occasional interseptal columns. However the intramembranous appearance in replicas is variable, particularly in centipedes, in that the rows of IMPs in chemically-unfixed propanecryofixed tissues, are prominent and adhere preferentially to the E face, with complementary P face grooves, while in fixed tissues the IMPs are much less distinct and fracture to either P face or E face. They tend not to protrude far beyond the mid-plane of the membrane bilayer and lie in rows which commonly take on the form of a network. Individual rows of the network sometimes curve to run beside a second row, over a short distance, before bending away into another part of the network. The aligned particle rows, which are much more prominent in millipedes, where they frequently lie in close parallel appositions, do not fuse into ridges as often occurs in insect tissues. The myriapod junctions, therefore, are of the same general kind as are found in the gut tract of other arthropod groups, but differ with respect to the subtleties of their intramembranous

  2. Terbinafine inhibits gap junctional intercellular communication.

    PubMed

    Lee, Ju Yeun; Yoon, Sei Mee; Choi, Eun Ju; Lee, Jinu

    2016-09-15

    Terbinafine is an antifungal agent that selectively inhibits fungal sterol synthesis by blocking squalene epoxidase. We evaluated the effect of terbinafine on gap junctional intercellular communication (GJIC). Fluorescence recovery after photobleaching (FRAP) and I-YFP GJIC assays revealed that terbinafine inhibits GJIC in a reversible and dose-dependent manner in FRT-Cx43 and LN215 cells. Treatment with terbinafine did not affect Cx43 phosphorylation status or intracellular Ca(2+) concentration, well-known action mechanisms of various GJIC blockers. While a structurally related chemical, naftifine, attenuated GJIC, epigallocatechin gallate, another potent squalene epoxidase inhibitor with a different structure, did not. These results suggest that terbinafine inhibits GJIC with a so far unknown mechanism of action.

  3. Altered patterns of cardiac intercellular junction distribution in hypertrophic cardiomyopathy.

    PubMed Central

    Sepp, R.; Severs, N. J.; Gourdie, R. G.

    1996-01-01

    OBJECTIVE: To examine the distribution pattern of intercellular junctions (the mechanically coupling desmosomes and the electrically coupling gap junctions) in hypertrophic cardiomyopathy (HCM) hearts showing myofibre disarray. DESIGN: Samples from six necropsied hearts were studied, representing the interventricular septum and the free walls of the left and right ventricles. Immunohistochemical labelling of desmoplakin was used as a marker for desmosomes, and of connexin43 as a marker for gap junctions, in single and double stainings. The slides were examined by confocal laser scanning microscopy. RESULTS: Marked disorganisation of intercalated discs was observed in areas featuring myofibre disarray. Besides overall derangement, localised abnormalities in desmosome organisation were evident, which included: (1) the formation of abnormally enlarged megadiscs; (2) the presence of intersecting disc structures; and (3) aberrant side to side desmosomal connections. Gap junctional abnormalities included: (1) random distribution of gap junctions over the surface of myocytes, rather than localisation to intercalated discs; (2) abundant side to side gap junction connections between adjacent myocytes; and (3) formation of abnormally shaped gap junctions. Circles of myocytes continuously interconnected by gap junctions were also observed. Regions of the diseased hearts lacking myofibre disarray, and control hearts of normal patients and patients with other cardiac diseases, did not show these alterations. CONCLUSIONS: The disorganisation of the intercellular junctions associated with myofibre disarray in HCM may play an important role in the pathophysiological manifestations of the disease. The remodelling of gap junction distribution may underlie the formation of an arrhythmogenic substrate, thereby contributing to the generation and maintenance of cardiac arrhythmias associated with HCM. Images PMID:8944586

  4. Gap Junction Intercellular Communication in Bone Marrow Failure

    DTIC Science & Technology

    2012-10-01

    osteogenesis, osteoblast function, or mesenchymal-derived hematopoietic support. This project postulates that the loss of gap junction ( GJ )-mediated...intercellular communication (IC) in the osteogenic HM is one of the mechanisms involved in dysfunctional mesenchymal hematopoietic support. GJ are cell...bone (30-39). GJ are formed by hexamers (hemichannels) of a family of proteins called connexins, with cell specific expression and functions(40, 41

  5. Treponema pallidum Invades Intercellular Junctions of Endothelial Cell Monolayers

    NASA Astrophysics Data System (ADS)

    Thomas, D. Denee; Navab, Mahamad; Haake, David A.; Fogelman, Alan M.; Miller, James N.; Lovett, Michael A.

    1988-05-01

    The pathogenesis of syphilis reflects invasive properties of Treponema pallidum, but the actual mode of tissue invasion is unknown. We have found two in vitro parallels of treponemal invasiveness. We tested whether motile T. pallidum could invade host cells by determining the fate of radiolabeled motile organisms added to a HeLa cell monolayer; 26% of treponemes associated with the monolayer in a trypsin-resistant niche, presumably between the monolayer and the surface to which it adhered, but did not attain intracellularity. Attachment of T. pallidum to cultured human and rabbit aortic and human umbilical vein endothelial cells was 2-fold greater than to HeLa cells. We added T. pallidum to aortic endothelial cells grown on membrane filters under conditions in which tight intercellular junctions had formed. T. pallidum was able to pass through the endothelial cell monolayers without altering tight junctions, as measured by electrical resistance. In contrast, heat-killed T. pallidum and the nonpathogen Treponema phagedenis biotype Reiter failed to penetrate the monolayer. Transmission electron micrographs of sections of the monolayer showed T. pallidum in intercellular junctions. Our in vitro observations suggest that these highly motile spirochetes may leave the circulation by invading the junctions between endothelial cells.

  6. F-actin binding protein, anillin, regulates integrity of intercellular junctions in human epithelial cells

    PubMed Central

    Feygin, Alex; Ivanov, Andrei I.

    2015-01-01

    Tight junctions (TJ) and adherens junctions (AJ) are key morphological features of differentiated epithelial cells that regulate the integrity and permeability of tissue barriers. Structure and remodeling of epithelial junctions depends on their association with the underlying actomyosin cytoskeleton. Anillin is a unique scaffolding protein interacting with different cytoskeletal components, including actin filaments and myosin motors. Its role in the regulation of mammalian epithelial junctions remains unexplored. Downregulation of anillin expression in human prostate, colonic, and lung epithelial cells triggered AJ and TJ disassembly without altering the expression of junctional proteins. This junctional disassembly was accompanied by dramatic disorganization of the perijunctional actomyosin belt; while the general architecture of the actin cytoskeleton, and activation status of non-muscle myosin II, remained unchanged. Furthermore, loss of anillin disrupted the adducin-spectrin membrane skeleton at the areas of cell-cell contact, selectively decreased γ-adducin expression, and induced cytoplasmic aggregation of αII-spectrin. Anillin knockdown activated c-Jun N-terminal kinase (JNK), and JNK inhibition restored AJ and TJ integrity and cytoskeletal organization in anillin-depleted cells. These findings suggest a novel role for anillin in regulating intercellular adhesion in model human epithelia by mechanisms involving the suppression of JNK activity and controlling the assembly of the perijunctional cytoskeleton. PMID:25809162

  7. Gap Junction Intercellular Communication Mediates Ammonia-Induced Neurotoxicity.

    PubMed

    Bobermin, Larissa Daniele; Arús, Bernardo Assein; Leite, Marina Concli; Souza, Diogo Onofre; Gonçalves, Carlos-Alberto; Quincozes-Santos, André

    2016-02-01

    Astrocytes are important brain targets of ammonia, a neurotoxin implicated in the development of hepatic encephalopathy. During hyperammonemia, the pivotal role of astrocytes in brain function and homeostasis is impaired. These cells are abundantly interconnected by gap junctions (GJ), which are intercellular channels that allow the exchange of signaling molecules and metabolites. This communication may also increase cellular vulnerability during injuries, while GJ uncoupling could limit the extension of a lesion. Therefore, the current study was performed to investigate whether astrocyte coupling through GJ contributes to ammonia-induced cytotoxicity. We found that carbenoxolone (CBX), an effective GJ blocker, prevented the following effects induced by ammonia in astrocyte primary cultures: (1) decrease in cell viability and membrane integrity; (2) increase in reactive oxygen species production; (3) decrease in GSH intracellular levels; (4) GS activity; (5) pro-inflammatory cytokine release. On the other hand, CBX had no effect on C6 astroglial cells, which are poorly coupled via GJ. To our knowledge, this study provides the first evidence that GJ play a role in ammonia-induced cytotoxicity. Although more studies in vivo are required to confirm our hypothesis, our data suggest that GJ communication between astrocytes may transmit damage signals and excitotoxic components from unhealthy to normal cells, thereby contributing to the propagation of the neurotoxicity of ammonia.

  8. Benzalkonium Chloride Suppresses Rabbit Corneal Endothelium Intercellular Gap Junction Communication

    PubMed Central

    Zhang, Zhenhao; Huang, Yue; Xie, Hui; Pan, Juxin; Liu, Fanfei; Li, Xuezhi; Chen, Wensheng; Hu, Jiaoyue; Liu, Zuguo

    2014-01-01

    Purpose Gap junction intercellular communication (GJIC) plays a critical role in the maintenance of corneal endothelium homeostasis. We determined if benzalkonium chloride (BAK) alters GJIC activity in the rabbit corneal endothelium since it is commonly used as a drug preservative in ocular eyedrop preparations even though it can have cytotoxic effects. Methods Thirty-six adult New Zealand albino rabbits were randomly divided into three groups. BAK at 0.01%, 0.05%, and 0.1% was applied twice daily to one eye of each of the rabbits in one of the three groups for seven days. The contralateral untreated eyes were used as controls. Corneal endothelial morphological features were observed by in vivo confocal microscopy (IVCM). Immunofluorescent staining resolved changes in gap junction integrity and localization. Western blot analysis and RT-PCR evaluated changes in levels of connexin43 (Cx43) and tight junction zonula occludens-1 (ZO-1) gene and protein expression, respectively. Cx43 and ZO-1 physical interaction was detected by immunoprecipitation (IP). Primary rabbit corneal endothelial cells were cultured in Dulbecco's Modified Eagle Medium (DMEM) containing BAK for 24 hours. The scrape-loading dye transfer technique (SLDT) was used to assess GJIC activity. Results Topical administration of BAK (0.05%, 0.1%) dose dependently disrupted corneal endothelial cell morphology, altered Cx43 and ZO-1 distribution and reduced Cx43 expression. BAK also markedly induced increases in Cx43 phosphorylation status concomitant with decreases in the Cx43-ZO-1 protein-protein interaction. These changes were associated with marked declines in GJIC activity. Conclusions The dose dependent declines in rabbit corneal endothelial GJIC activity induced by BAK are associated with less Cx43-ZO-1 interaction possibly arising from increases in Cx43 phosphorylation and declines in its protein expression. These novel changes provide additional evidence that BAK containing eyedrop preparations

  9. The modulation of gap-junctional intercellular communication by lipid rafts.

    PubMed

    Defamie, Norah; Mesnil, Marc

    2012-08-01

    Lipid rafts are specific microdomains of plasma membrane which are enriched in cholesterol and sphingolipids. These domains seem to favour the interactions of particular proteins and the regulation of signalling pathways in the cells. Recent data have shown that among the proteins, which are preferentially localized in lipid rafts, are connexins that are the structural proteins of gap junctions. Since gap junctional intercellular communication is involved in various cellular processes and pathologies such as cancer, we were interested to review the various observations concerning this specific localization of connexins in lipid rafts and its consequences on gap junctional intercellular communication capacity. In particular, we will focus our discussion on the role of the lipid raft-connexin connection in cancer progression. This article is part of a Special Issue entitled: The Communicating junctions, composition, structure and characteristics.

  10. Effect of sound on gap-junction-based intercellular signaling: Calcium waves under acoustic irradiation

    NASA Astrophysics Data System (ADS)

    Deymier, P. A.; Swinteck, N.; Runge, K.; Deymier-Black, A.; Hoying, J. B.

    2015-11-01

    We present a previously unrecognized effect of sound waves on gap-junction-based intercellular signaling such as in biological tissues composed of endothelial cells. We suggest that sound irradiation may, through temporal and spatial modulation of cell-to-cell conductance, create intercellular calcium waves with unidirectional signal propagation associated with nonconventional topologies. Nonreciprocity in calcium wave propagation induced by sound wave irradiation is demonstrated in the case of a linear and a nonlinear reaction-diffusion model. This demonstration should be applicable to other types of gap-junction-based intercellular signals, and it is thought that it should be of help in interpreting a broad range of biological phenomena associated with the beneficial therapeutic effects of sound irradiation and possibly the harmful effects of sound waves on health.

  11. Effect of sound on gap-junction-based intercellular signaling: Calcium waves under acoustic irradiation.

    PubMed

    Deymier, P A; Swinteck, N; Runge, K; Deymier-Black, A; Hoying, J B

    2015-01-01

    We present a previously unrecognized effect of sound waves on gap-junction-based intercellular signaling such as in biological tissues composed of endothelial cells. We suggest that sound irradiation may, through temporal and spatial modulation of cell-to-cell conductance, create intercellular calcium waves with unidirectional signal propagation associated with nonconventional topologies. Nonreciprocity in calcium wave propagation induced by sound wave irradiation is demonstrated in the case of a linear and a nonlinear reaction-diffusion model. This demonstration should be applicable to other types of gap-junction-based intercellular signals, and it is thought that it should be of help in interpreting a broad range of biological phenomena associated with the beneficial therapeutic effects of sound irradiation and possibly the harmful effects of sound waves on health.

  12. Chlorpromazine reduces the intercellular communication via gap junctions in mammalian cells

    SciTech Connect

    Orellana, Juan A.; Palacios-Prado, Nicolas; Saez, Juan C. . E-mail: jsaez@bio.puc.cl

    2006-06-15

    In the work presented herein, we evaluated the effect of chlorpromazine (CPZ) on gap junctions expressed by two mammalian cell types; Gn-11 cells (cell line derived from mouse LHRH neurons) and rat cortical astrocytes maintained in culture. We also attempted to elucidate possible mechanisms of action of CPZ effects on gap junctions. CPZ, in concentrations comparable with doses used to treat human diseases, was found to reduce the intercellular communication via gap junctions as evaluated with measurements of dye coupling (Lucifer yellow). In both cell types, maximal inhibition of functional gap junctions was reached within about 1 h of treatment with CPZ, an recovery was almost complete at about 5 h after CPZ wash out. In both cell types, CPZ treatment increased the phosphorylation state of connexin43 (Cx43), a gap junction protein subunit. Moreover, CPZ reduced the reactivity of Cx43 (immunofluorescence) at cell interfaces and concomitantly increased its reactivity in intracellular vesicles, suggesting an increased retrieval from and/or reduced insertion into the plasma membrane. CPZ also caused cellular retraction reducing cell-cell contacts in a reversible manner. The reduction in contact area might destabilize existing gap junctions and abrogate formation of new ones. Moreover, the CPZ-induced reduction in gap junctional communication may depend on the connexins (Cxs) forming the junctions. If Cx43 were the only connexin expressed, MAPK-dependent phosphorylation of this connexin would induce closure of gap junction channels.

  13. The Role of Chemical Inhibition of Gap Junctional Intercellular Communication in Toxicology.

    DTIC Science & Technology

    1988-02-14

    cell communication, tumor promoters, terato- 0620 gens, neurotoxins, protein kinase C, chemical toxicity. 19 ABSTRACT (Continue on reverse if necessary...hypothesis that chemical modulation of gap junctional intercellular communication can lead to many toxic endpoints, such as teratogenesis, tumor promotion... tumor promotion, reproductive-, immune- and neurotoxicities. To date, after two years into the project, we have initiated work on all of the specific aims

  14. The Role of Chemical Inhibition of Gap-Junctional Intercellular Communication in Toxicology

    DTIC Science & Technology

    1991-03-31

    Florida. Inhibition of gap junctional intercellular communication (GJIC has been implicated as an important epigenetic modulation during...Annual Meeting of the Society of Toxicology, February 1991, Dallas, Texas. A major epigenetic modulation induced by many tumor promoters both in vivo...GJIC in rat pancreatic epithelial cells. The results indicated that many chlorinated pesticides , the phorbol ester tumor promoter, TPA, and a number of

  15. A fine structural analysis of intercellular junctions in the mouse liver.

    PubMed

    Goodenough, D A; Revel, J P

    1970-05-01

    Zonulae occludentes and gap junctions were examined both in the intact mouse liver and in a junction-rich membrane fraction from homogenized mouse liver. These preparations were visualized with the techniques of uranyl acetate staining en bloc, staining with colloidal lanthanum, negative staining with phosphotungstate, and freeze-cleaving. The zonula occludens is arranged as a meshwork of branching and anastomosing threadlike contacts sealing the lumen of the bile canaliculus from the liver intercellular space. The gap junction is characterized in section by a 20 A gap between the apposed junctional membrane outer leaflets, and permeation of this space with lanthanum or phosphotungstate reveals a polygonal lattice of subunits with a center-to-center spacing of 90-100 A. Freeze-cleaved gap junctions show a similar lattice. Extraction of junction-rich fractions with 60% aqueous acetone results in a disappearance of the 20 A gap in sectioned pellets and an inability to demonstrate the polygonal lattice with either the freeze-cleave or negative staining techniques. Extraction of the membranes with 50% acetone does not produce this effect. Thin-layer chromatography of the acetone extracts reveals a group of phospholipids in the 60% extract that are not detectable in the 50% extract. Acetone does not cause any detectable change in the structure of the zonula occludens, but the occluding junction becomes leaky to lanthanum following acetone treatment. The effects of other reagents on the junctions are reported.

  16. Reversible Opening of Intercellular Junctions of Intestinal Epithelial and Brain Endothelial Cells With Tight Junction Modulator Peptides.

    PubMed

    Bocsik, Alexandra; Walter, Fruzsina R; Gyebrovszki, Andrea; Fülöp, Lívia; Blasig, Ingolf; Dabrowski, Sebastian; Ötvös, Ferenc; Tóth, András; Rákhely, Gábor; Veszelka, Szilvia; Vastag, Monika; Szabó-Révész, Piroska; Deli, Mária A

    2016-02-01

    The intercellular junctions restrict the free passage of hydrophilic compounds through the paracellular clefts. Reversible opening of the tight junctions of biological barriers is investigated as one of the ways to increase drug delivery to the systemic circulation or the central nervous system. Six peptides, ADT-6, HAV-6, C-CPE, 7-mer (FDFWITP, PN-78), AT-1002, and PN-159, acting on different integral membrane and linker junctional proteins were tested on Caco-2 intestinal epithelial cell line and a coculture model of the blood-brain barrier. All peptides tested in nontoxic concentrations showed a reversible tight junctions modulating effect and were effective to open the paracellular pathway for the marker molecules fluorescein and albumin. The change in the structure of cell-cell junctions was verified by immunostaining for occludin, claudin-4,-5, ZO-1, β-catenin, and E-cadherin. Expression levels of occludin and claudins were measured in both models. We could demonstrate a selectivity of C-CPE, ADT-6, and HAV-6 peptides for epithelial cells and 7-mer and AT-1002 peptides for brain endothelial cells. PN-159 was the most effective modulator of junctional permeability in both models possibly acting via claudin-1 and -5. Our results indicate that these peptides can be effectively and selectively used as potential pharmaceutical excipients to improve drug delivery across biological barriers.

  17. Kinetic Measurements Reveal Enhanced Protein-Protein Interactions at Intercellular Junctions

    PubMed Central

    Shashikanth, Nitesh; Kisting, Meridith A.; Leckband, Deborah E.

    2016-01-01

    The binding properties of adhesion proteins are typically quantified from measurements with soluble fragments, under conditions that differ radically from the confined microenvironment of membrane bound proteins in adhesion zones. Using classical cadherin as a model adhesion protein, we tested the postulate that confinement within quasi two-dimensional intercellular gaps exposes weak protein interactions that are not detected in solution binding assays. Micropipette-based measurements of cadherin-mediated, cell-cell binding kinetics identified a unique kinetic signature that reflects both adhesive (trans) bonds between cadherins on opposing cells and lateral (cis) interactions between cadherins on the same cell. In solution, proposed lateral interactions were not detected, even at high cadherin concentrations. Mutations postulated to disrupt lateral cadherin association altered the kinetic signatures, but did not affect the adhesive (trans) binding affinity. Perturbed kinetics further coincided with altered cadherin distributions at junctions, wound healing dynamics, and paracellular permeability. Intercellular binding kinetics thus revealed cadherin interactions that occur within confined, intermembrane gaps but not in solution. Findings further demonstrate the impact of these revealed interactions on the organization and function of intercellular junctions. PMID:27009566

  18. INHIBITION OF GAP JUNCTIONAL INTERCELLULAR COMMUNICATION BY PERFLUORINATED COMPOUNDS IN RAT LIVER AND DOLPHIN KIDNEY EPITHELIAL CELL LINES IN VITRO AND SPRAGUE-DAWLEY RATS IN VIVO

    EPA Science Inventory

    Abstract

    Gap Junctional Intercellular Communication (GJIC) is the major pathway of intercellular signal transduction, and is, thus, important for normal cell growth and function. Recent studies have revealed a global distribution of some perfluorinated organic compounds e...

  19. Requirement of gap junctional intercellular communication for human villous trophoblast differentiation.

    PubMed

    Cronier, Laurent; Frendo, Jean-Louis; Defamie, Norah; Pidoux, Guillaume; Bertin, Gladys; Guibourdenche, Jean; Pointis, Georges; Malassine, Andre

    2003-11-01

    During pregnancy, the villous trophoblast develops from the fusion of cytotrophoblastic cells (CT) into a syncytiotrophoblast (ST), supporting the main physiological functions of the human placenta. Connexin43 (Cx43) is demonstrated in situ and in vitro in the villous trophoblast between CT and between CT and ST. Moreover, the presence of a gap junctional intercellular communication (GJIC) during in vitro trophoblast differentiation was previously demonstrated. Because the exchange of molecules through gap junctions is considered to play a major role in the control of cell and tissue differentiation, we studied the effects of a gap junctional uncoupler, heptanol, on morphological and functional trophoblast differentiation and on GJIC measured by the fluorescence recovery after photobleaching method. We found that when the GJIC was interrupted, CT still aggregated but fused poorly. This morphological effect was associated with a significant decrease of trophoblastic-specific gene expression (beta human chorionic gonadotropin and human chorionic somatomammotropin). This blocking action was reversible as demonstrated by recovery of GJIC and trophoblast differentiation process after heptanol removal. Moreover, the inhibition of the trophoblast differentiation did not affect Cx43 transcript expression and Cx43 protein expression. These data suggest that the molecular exchanges through gap junctions preceding cellular fusion are essential for trophoblast differentiation generating the multifunctional syncytiotrophoblast.

  20. A rapid and sensitive assay of intercellular coupling by voltage imaging of gap junction networks

    PubMed Central

    2013-01-01

    Background A variety of mechanisms that govern connexin channel gating and permeability regulate coupling in gap junction networks. Mutations in connexin genes have been linked to several pathologies, including cardiovascular anomalies, peripheral neuropathy, skin disorders, cataracts and deafness. Gap junction coupling and its patho–physiological alterations are commonly assayed by microinjection experiments with fluorescent tracers, which typically require several minutes to allow dye transfer to a limited number of cells. Comparable or longer time intervals are required by fluorescence recovery after photobleaching experiments. Paired electrophysiological recordings have excellent time resolution but provide extremely limited spatial information regarding network connectivity. Results Here, we developed a rapid and sensitive method to assay gap junction communication using a combination of single cell electrophysiology, large–scale optical recordings and a digital phase–sensitive detector to extract signals with a known frequency from Vf2.1.Cl, a novel fluorescent sensor of plasma membrane potential. Tests performed in HeLa cell cultures confirmed that suitably encoded Vf2.1.Cl signals remained confined within the network of cells visibly interconnected by fluorescently tagged gap junction channels. We used this method to visualize instantly intercellular connectivity over the whole field of view (hundreds of cells) in cochlear organotypic cultures from postnatal mice. A simple resistive network model reproduced accurately the spatial dependence of the electrical signals throughout the cellular network. Our data suggest that each pair of cochlear non−sensory cells of the lesser epithelial ridge is coupled by ~1500 gap junction channels, on average. Junctional conductance was reduced by 14% in cochlear cultures harboring the T5M mutation of connexin30, which induces a moderate hearing loss in connexin30T5M/T5M knock–in mice, and by 91% in cultures from

  1. Regulation of gap junctional intercellular communication by TCDD in HMEC and MCF-7 breast cancer cells

    SciTech Connect

    Gakhar, Gunjan Schrempp, Diane Nguyen, Thu Annelise

    2009-03-01

    Previous studies suggest that many neoplastic tissues exhibit a decrease in gap junctional intercellular communication (GJIC). Many hydrocarbons and organochlorine compounds are environmental pollutants known to be carcinogenic. The effect of an organochlorine compound, TCDD, on GJIC in human breast cell lines has not been established. In the present study, we showed that TCDD causes an inhibition in the gap junctional activity in MCF-7 (breast cancer cells). In MCF-7 cells, an increase in the phosphorylated form of gap junctional protein, connexin 43 (Cx43), and PKC {alpha} was seen in the presence of TCDD. Gap junctional plaque formation was significantly decreased in MCF-7 cells in the presence of TCDD. Immunoprecipitation studies of PKC {alpha} showed that TCDD caused a significant 40% increase in the phosphorylated Cx43 in MCF-7 cells. TCDD also modulated the translocation of PKC {alpha} from the cytosol to the membrane and caused a 2-fold increase in the PKC {alpha} activity at 50 nM TCDD in MCF-7 cells. Calphostin C, an inhibitor of PKC {alpha}, showed a significant inhibition of PKC {alpha} activity in the presence of TCDD. Furthermore, TCDD also caused a decrease in the gap junctional activity and Cx43 protein in human mammary epithelial cells (HMEC). However, we observed a shift in the Cx43 plaques towards the perinuclear membrane in the presence of TCDD by confocal microscopy and Western blot. Overall, these results conclude that TCDD decreases GJIC by phosphorylating Cx43 via PKC {alpha} signaling pathway in MCF-7 cells; however, TCDD decreases the GJIC by affecting the localization of Cx43 in HMEC. These new findings elucidate the differential mode of effect of TCDD in the downregulation of GJIC in HMEC and MCF-7 cells.

  2. [Gap junctional intercellular communication: a new mechanism in pathophysiology of migraine with aura. Therapeutic applications].

    PubMed

    Sarrouilhe, D; Dejean, C

    2012-12-01

    Migraine is a common, recurrent and disabling primary headache disorder, which affects up to 20% of the population. About a third of patients with migraine have attacks with aura, a focal neurological disturbance that manifests itself as visual, sensitive or motor symptoms. Cortical spreading depression, a wave of electrical activity that moves across the cerebral cortex through neuronal-glial cell gap junctions, would be involved in the triggering of migraine aura. Moreover, cortical spreading depression activates perivascular trigeminal afferents in the neocortex, that through central and peripheral reflex, cause inflammatory reaction in the meninges to generate the headache. Tonabersat, a novel benzopyran compound, was selected for clinical trial on the basis of its inhibitory activity on cortical spreading depression and neurogenic inflammation in animal models of migraine. Moreover, tonabersat inhibited trigeminal ganglion neuronal-glial cell gap junctions, suggesting that this compound could prevent peripheral sensitization within the ganglion. In clinical trial, tonabersat showed a preventive effect on attacks of migraine with aura but had no efficacy on non-aura attacks and in the acute treatment of migraine. In conclusion, neuronal-glial cell gap junctional intercellular communication seems to be involved in the pathophysiology of migraine with aura and is emerging as a new promising therapeutic target for prophylactic treatment of patients with chronic attacks.

  3. Intercellular signaling via cyclic GMP diffusion through gap junctions restarts meiosis in mouse ovarian follicles.

    PubMed

    Shuhaibar, Leia C; Egbert, Jeremy R; Norris, Rachael P; Lampe, Paul D; Nikolaev, Viacheslav O; Thunemann, Martin; Wen, Lai; Feil, Robert; Jaffe, Laurinda A

    2015-04-28

    Meiosis in mammalian oocytes is paused until luteinizing hormone (LH) activates receptors in the mural granulosa cells of the ovarian follicle. Prior work has established the central role of cyclic GMP (cGMP) from the granulosa cells in maintaining meiotic arrest, but it is not clear how binding of LH to receptors that are located up to 10 cell layers away from the oocyte lowers oocyte cGMP and restarts meiosis. Here, by visualizing intercellular trafficking of cGMP in real-time in live follicles from mice expressing a FRET sensor, we show that diffusion of cGMP through gap junctions is responsible not only for maintaining meiotic arrest, but also for rapid transmission of the signal that reinitiates meiosis from the follicle surface to the oocyte. Before LH exposure, the cGMP concentration throughout the follicle is at a uniformly high level of ∼2-4 μM. Then, within 1 min of LH application, cGMP begins to decrease in the peripheral granulosa cells. As a consequence, cGMP from the oocyte diffuses into the sink provided by the large granulosa cell volume, such that by 20 min the cGMP concentration in the follicle is uniformly low, ∼100 nM. The decrease in cGMP in the oocyte relieves the inhibition of the meiotic cell cycle. This direct demonstration that a physiological signal initiated by a stimulus in one region of an intact tissue can travel across many layers of cells via cyclic nucleotide diffusion through gap junctions could provide a general mechanism for diverse cellular processes.

  4. microRNAs, Gap Junctional Intercellular Communication and Mesenchymal Stem Cells in Breast Cancer Metastasis

    PubMed Central

    Gregory, Larissa A.; Ricart, Rachel A.; Patel, Shyam A.; Lim, Philip K.; Rameshwar, Pranela

    2010-01-01

    The failed outcome of autologous bone marrow transplantation for breast cancer opens the field for investigations. This is particularly important because the bone marrow could be a major source of cancer cells during tertiary metastasis. This review discusses subsets of breast cancer cells, including those that enter the bone marrow at an early period of disease development, perhaps prior to clinical detection. This population of cells evades chemotherapeutic damage even at high doses. An understanding of this population might be crucial for the success of bone marrow transplants for metastatic breast cancer and for the eradication of cancer cells in bone marrow. In vivo and in vitro studies have demonstrated gap junctional intercellular communication (GJIC) between bone marrow stroma and breast cancer cells. This review discusses GJIC in cancer metastasis, facilitating roles of mesenchymal stem cells (MSCs). In addition, the review addresses potential roles for miRNAs, including those already linked to cancer biology. The literature on MSCs is growing and their links to metastasis are beginning to be significant leads for the development of new drug targets for breast cancer. In summary, this review discusses interactions among GJIC, miRNAs and MSCs as future consideration for the development of cancer therapies. PMID:21886602

  5. Chemopreventive Agents Attenuate Rapid Inhibition of Gap Junctional Intercellular Communication Induced by Environmental Toxicants.

    PubMed

    Babica, Pavel; Čtveráčková, Lucie; Lenčešová, Zuzana; Trosko, James E; Upham, Brad L

    2016-07-01

    Altered gap junctional intercellular communication (GJIC) has been associated with chemical carcinogenesis, where both chemical tumor promoters and chemopreventive agents (CPAs) are known to conversely modulate GJIC. The aim of this study was to investigate whether attenuation of chemically inhibited GJIC represents a common outcome induced by different CPAs, which could be effectively evaluated using in vitro methods. Rat liver epithelial cells WB-F344 were pretreated with a CPA for either 30 min or 24 h, and then exposed to GJIC-inhibiting concentration of a selected tumor promoter or environmental toxicant [12-O-tetradecanoylphorbol-13-acetate (TPA), lindane, fluoranthene, 1,1,1-trichloro-2,2-bis(4-chlorophenyl)ethane (DDT), perfluorooctanoic acid (PFOA), or pentachlorophenol]. Out of nine CPAs tested, quercetin and silibinin elicited the most pronounced effects, preventing the dysregulation of GJIC by all the GJIC inhibitors, but DDT. Metformin and curcumin attenuated the effects of three GJIC inhibitors, whereas the other CPAs prevented the effects of two (diallyl sulfide, emodin) or one (indole-3-carbinol, thymoquinone) GJIC inhibitor. Significant attenuation of chemically induced inhibition of GJIC was observed in 27 (50%) out of 54 possible combinations of nine CPAs and six GJIC inhibitors. Our data demonstrate that in vitro evaluation of GJIC can be used as an effective screening tool for identification of chemicals with potential chemopreventive activity.

  6. Caveolin-1 and -2 Interact with Connexin43 and Regulate Gap Junctional Intercellular Communication in Keratinocytes

    PubMed Central

    Langlois, Stéphanie; Cowan, Kyle N.; Shao, Qing; Cowan, Bryce J.

    2008-01-01

    Connexin43 (Cx43) has been reported to interact with caveolin (Cav)-1, but the role of this association and whether other members of the caveolin family bind Cx43 had yet to be established. In this study, we show that Cx43 coimmunoprecipitates and colocalizes with Cav-1 and Cav-2 in rat epidermal keratinocytes. The colocalization of Cx43 with Cav-1 was confirmed in keratinocytes from human epidermis in vivo. Our mutation and Far Western analyses revealed that the C-terminal tail of Cx43 is required for its association with Cavs and that the Cx43/Cav-1 interaction is direct. Our results indicate that newly synthesized Cx43 interacts with Cavs in the Golgi apparatus and that the Cx43/Cavs complex also exists at the plasma membrane in lipid rafts. Using overexpression and small interfering RNA approaches, we demonstrated that caveolins regulate gap junctional intercellular communication (GJIC) and that the presence of Cx43 in lipid raft domains may contribute to the mechanism modulating GJIC. Our results suggest that the Cx43/Cavs association occurs during exocytic transport, and they clearly indicate that caveolin regulates GJIC. PMID:18162583

  7. Bay or baylike regions of polycyclic aromatic hydrocarbons were potent inhibitors of Gap junctional intercellular communication.

    PubMed Central

    Weis, L M; Rummel, A M; Masten, S J; Trosko, J E; Upham, B L

    1998-01-01

    Many polycyclic aromatic hydrocarbons (PAHs) are known carcinogens, and a considerable amount of research has been devoted to predicting the tumor-initiating potential of PAHs based on chemical structure. However, there has been little research into the effects of PAHs on the epigenetic events of tumor promotion and no structural correlation has been made thereof. Gap junctional intercellular communication (GJIC) activity was used in this study as an epigenetic biomarker to determine the structure-activity relationships of twelve different PAHs. The PAHs used were naphthalene, 1-methylnaphthalene, 2-methylnaphthalene, anthracene, 1-methylanthracene, 2-methylanthracene, 9-methylanthracene, 9, 10-dimethylanthracene, phenanthrene, fluorene, 1-methylfluorene, and fluoranthene. Results showed that PAHs containing bay or baylike regions inhibited GJIC more than did the linear PAHs. The nonnaphthalene PAHs were not cytotoxic as determined by a vital dye uptake assay, but the naphthalene compounds were cytotoxic at the higher doses, indicating that the down regulation of GJIC by these naphthalenes could be a consequence of general membrane damage. Inhibition of GJIC by all the inhibitory PAHs was reversed when the cells were refreshed with PAH-free growth medium. Inhibition of GJIC occurred within 0.5-5 min and correlated with the aqueous solubility of the PAHs. The present study revealed that there are structural determinants of epigenetic toxicity as determined by GJIC activity. Images Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Figure 7 Figure 8 Figure 9 Figure 10 PMID:9417772

  8. Extract from the zooxanthellate jellyfish Cotylorhiza tuberculata modulates gap junction intercellular communication in human cell cultures.

    PubMed

    Leone, Antonella; Lecci, Raffaella Marina; Durante, Miriana; Piraino, Stefano

    2013-05-22

    On a global scale, jellyfish populations in coastal marine ecosystems exhibit increasing trends of abundance. High-density outbreaks may directly or indirectly affect human economical and recreational activities, as well as public health. As the interest in biology of marine jellyfish grows, a number of jellyfish metabolites with healthy potential, such as anticancer or antioxidant activities, is increasingly reported. In this study, the Mediterranean "fried egg jellyfish" Cotylorhiza tuberculata (Macri, 1778) has been targeted in the search forputative valuable bioactive compounds. A medusa extract was obtained, fractionated, characterized by HPLC, GC-MS and SDS-PAGE and assayed for its biological activity on breast cancer cells (MCF-7) and human epidermal keratinocytes (HEKa). The composition of the jellyfish extract included photosynthetic pigments, valuable ω-3 and ω-6 fatty acids, and polypeptides derived either from jellyfish tissues and their algal symbionts. Extract fractions showed antioxidant activity and the ability to affect cell viability and intercellular communication mediated by gap junctions (GJIC) differentially in MCF-7 and HEKa cells. A significantly higher cytotoxicity and GJIC enhancement in MCF-7 compared to HEKa cells was recorded. A putative action mechanism for the anticancer bioactivity through the modulation of GJIC has been hypothesized and its nutraceutical and pharmaceutical potential was discussed.

  9. Effect of airborne particles from selected indoor and outdoor environments on gap-junctional intercellular communication.

    PubMed

    Alink, G M; Sjögren, M; Bos, R P; Doekes, G; Kromhout, H; Scheepers, P T

    1998-08-01

    The effect of airborne particles from diesel exhaust, rubber and metal industry, urban air and biological sources (poultry, pig farming, compost industry) on gap-junctional intercellular communication (GJIC) were compared, using HEPA1c1c7 cells. Particles as such were compared with aqueous and organic extracts. Significant inhibition of GJIC by particle suspensions was only observed for the diesel and rubber samples, and for one biological sample (compost). Up to 83% of the inhibition of the whole suspension could be attributed to the particles as such. Washing the particles with organic solvents (aceton, methanol, hexane) did not result in a significant loss of activity from the particles, although the organic fractions showed a significant activity towards GJIC. More active organics was eluted from the rubber industry particles than from the diesel particles by the organic solvent. It is suggested that cancer promoting potential as measured by inhibition of GJIC may vary widely depending on the particle source, and that this effect may be exerted by the particles as such and/or by means of tightly bound bio-active material to the surface.

  10. Extract from the Zooxanthellate Jellyfish Cotylorhiza tuberculata Modulates Gap Junction Intercellular Communication in Human Cell Cultures

    PubMed Central

    Leone, Antonella; Lecci, Raffaella Marina; Durante, Miriana; Piraino, Stefano

    2013-01-01

    On a global scale, jellyfish populations in coastal marine ecosystems exhibit increasing trends of abundance. High-density outbreaks may directly or indirectly affect human economical and recreational activities, as well as public health. As the interest in biology of marine jellyfish grows, a number of jellyfish metabolites with healthy potential, such as anticancer or antioxidant activities, is increasingly reported. In this study, the Mediterranean “fried egg jellyfish” Cotylorhiza tuberculata (Macri, 1778) has been targeted in the search forputative valuable bioactive compounds. A medusa extract was obtained, fractionated, characterized by HPLC, GC-MS and SDS-PAGE and assayed for its biological activity on breast cancer cells (MCF-7) and human epidermal keratinocytes (HEKa). The composition of the jellyfish extract included photosynthetic pigments, valuable ω-3 and ω-6 fatty acids, and polypeptides derived either from jellyfish tissues and their algal symbionts. Extract fractions showed antioxidant activity and the ability to affect cell viability and intercellular communication mediated by gap junctions (GJIC) differentially in MCF-7and HEKa cells. A significantly higher cytotoxicity and GJIC enhancement in MCF-7 compared to HEKa cells was recorded. A putative action mechanism for the anticancer bioactivity through the modulation of GJIC has been hypothesized and its nutraceutical and pharmaceutical potential was discussed. PMID:23697954

  11. Local Oxidative Stress Expansion through Endothelial Cells – A Key Role for Gap Junction Intercellular Communication

    PubMed Central

    Feine, Ilan; Pinkas, Iddo; Salomon, Yoram; Scherz, Avigdor

    2012-01-01

    Background Major circulation pathologies are initiated by oxidative insult expansion from a few injured endothelial cells to distal sites; this possibly involves mechanisms that are important to understanding circulation physiology and designing therapeutic management of myocardial pathologies. We tested the hypothesis that a localized oxidative insult of endothelial cells (ECs) propagates through gap junction inter-cellular communication (GJIC). Methodology/Principal Findings Cultures comprising the bEnd.3 cell line, that have been established and recognized as suitable for examining communication among ECs, were used to study the propagation of a localized oxidative insult to remote cells. Spatially confined near infrared illumination of parental or genetically modified bEnd.3 cultures, pretreated with the photosensitizer WST11, generated O2•− and •OH radicals in the illuminated cells. Time-lapse fluorescence microscopy, utilizing various markers, and other methods, were used to monitor the response of non-illuminated bystander and remote cells. Functional GJIC among ECs was shown to be mandatory for oxidative insult propagation, comprising de-novo generation of reactive oxygen and nitrogen species (ROS and RNS, respectively), activation and nuclear translocation of c-Jun N-terminal kinase, followed by massive apoptosis in all bystander cells adjacent to the primarily injured ECs. The oxidative insult propagated through GJIC for many hours, over hundreds of microns from the primary photogeneration site. This wave is shown to be limited by intracellular ROS scavenging, chemical GJIC inhibition or genetic manipulation of connexin 43 (a key component of GJIC). Conclusion/Significance Localized oxidative insults propagate through GJIC between ECs, while stimulating de-novo generation of ROS and RNS in bystander cells, thereby driving the insult's expansion. PMID:22911831

  12. Effect of DDT on hepatic gap junctional intercellular communication in rats.

    PubMed

    Tateno, C; Ito, S; Tanaka, M; Oyamada, M; Yoshitake, A

    1994-03-01

    The effects of in vivo exposure to DDT on hepatic gap junctional intercellular communication (GJIC) and connexin gene/protein expression in Sprague-Dawley rats were examined by in vivo/in vitro dye-transfer assay, immunohistochemical staining, and by Western and Northern blot analyses. In the dose-response study, three dose levels of DDT (5, 25 and 50 mg/kg/day) were administered orally to rats once a day for 2 weeks. The average size of the dye spread after injection of Lucifer Yellow and the area of Cx32 spots per hepatocyte decreased in a dose-dependent manner, but there was no effect on the number of Cx32 spots per hepatocyte. In the time-course study, DDT (50 mg/kg/day) was administered orally once a day for up to 6 weeks. Hepatic GJIC decreased at week 1 but recovered at week 6. The average area of Cx32 spots per hepatocyte gradually decreased at weeks 2 and 4, and remained at the same level at week 6, correlating with the decreased Cx32 protein level in plasma membranes. The average area of Cx26 spots per hepatocyte in the peripheral zones clearly decreased at week 1, but quickly recovered at week 2 and increased at week 6; however, no clear change of the Cx26 protein level in plasma membranes was observed. No changes of Cx32 and Cx26 mRNA levels were observed in DDT groups. These results suggest that DDT, a liver tumor-promoting agent, inhibits hepatic GJIC in vivo dose-dependently in rats and that aberrant Cx32 and Cx26 protein expression and/or localization may be responsible for this effect.

  13. Activation of L-type calcium channels is required for gap junction-mediated intercellular calcium signaling in osteoblastic cells

    NASA Technical Reports Server (NTRS)

    Jorgensen, Niklas Rye; Teilmann, Stefan Cuoni; Henriksen, Zanne; Civitelli, Roberto; Sorensen, Ole Helmer; Steinberg, Thomas H.

    2003-01-01

    The propagation of mechanically induced intercellular calcium waves (ICW) among osteoblastic cells occurs both by activation of P2Y (purinergic) receptors by extracellular nucleotides, resulting in "fast" ICW, and by gap junctional communication in cells that express connexin43 (Cx43), resulting in "slow" ICW. Human osteoblastic cells transmit intercellular calcium signals by both of these mechanisms. In the current studies we have examined the mechanism of slow gap junction-dependent ICW in osteoblastic cells. In ROS rat osteoblastic cells, gap junction-dependent ICW were inhibited by removal of extracellular calcium, plasma membrane depolarization by high extracellular potassium, and the L-type voltage-operated calcium channel inhibitor, nifedipine. In contrast, all these treatments enhanced the spread of P2 receptor-mediated ICW in UMR rat osteoblastic cells. Using UMR cells transfected to express Cx43 (UMR/Cx43) we confirmed that nifedipine sensitivity of ICW required Cx43 expression. In human osteoblastic cells, gap junction-dependent ICW also required activation of L-type calcium channels and influx of extracellular calcium.

  14. The tight junction protein ZO-2 and Janus kinase 1 mediate intercellular communications in vascular smooth muscle cells

    SciTech Connect

    Tkachuk, Natalia; Tkachuk, Sergey; Patecki, Margret; Kusch, Angelika; Korenbaum, Elena; Haller, Hermann; Dumler, Inna

    2011-07-08

    Highlights: {yields} The tight junction protein ZO-2 associates with Jak1 in vascular smooth muscle cells via ZO-2 N-terminal fragment. {yields} Jak1 mediates ZO-2 tyrosine phosphorylation and ZO-2 localization to the sites of homotypic intercellular contacts. {yields} The urokinase receptor uPAR regulates ZO-2/Jak1 functional association. {yields} The ZO-2/Jak1/uPAR signaling complex is required for vascular smooth muscle cells functional network formation. -- Abstract: Recent evidence points to a multifunctional role of ZO-2, the tight junction protein of the MAGUK (membrane-associated guanylate kinase-like) family. Though ZO-2 has been found in cell types lacking tight junction structures, such as vascular smooth muscle cells (VSMC), little is known about ZO-2 function in these cells. We provide evidence that ZO-2 mediates specific homotypic cell-to-cell contacts between VSMC. Using mass spectrometry we found that ZO-2 is associated with the non-receptor tyrosine kinase Jak1. By generating specific ZO-2 constructs we further found that the N-terminal fragment of ZO-2 molecule is responsible for this interaction. Adenovirus-based expression of Jak1 inactive mutant demonstrated that Jak1 mediates ZO-2 tyrosine phosphorylation. By means of RNA silencing, expression of Jak1 mutant form and fluorescently labeled ZO-2 fusion protein we further specified that active Jak1, but not Jak1 inactive mutant, mediates ZO-2 localization to the sites of intercellular contacts. We identified the urokinase receptor uPAR as a pre-requisite for these cellular events. Functional requirement of the revealed signaling complex for VSMC network formation was confirmed in experiments using Matrigel and in contraction assay. Our findings imply involvement of the ZO-2 tight junction independent signaling complex containing Jak1 and uPAR in VSMC intercellular communications. This mechanism may contribute to vascular remodeling in occlusive cardiovascular diseases and in arteriogenesis.

  15. Intercellular calcium waves in the fire-diffuse-fire framework: Green's function for gap-junctional coupling.

    PubMed

    Harris, Jamie; Timofeeva, Yulia

    2010-11-01

    Calcium is a crucial component in a plethora of cellular processes involved in cell birth, life, and death. Intercellular calcium waves that can spread through multiple cells provide one form of cellular communication mechanism between various parts of cell tissues. Here we introduce a simple, yet biophysically realistic model for the propagation of intercellular calcium waves based on the fire-diffuse-fire type model for calcium dynamics. Calcium release sites are considered to be discretely distributed along individual linear cells that are connected by gap junctions and a solution of this model can be found in terms of the Green's function for this system. We develop the "sum-over-trips" formalism that takes into account the boundary conditions at gap junctions providing a generalization of the original sum-over-trips approach for constructing the response function for branched neural dendrites. We obtain the exact solution of the Green's function in the Laplace (frequency) domain for an infinite array of cells and show that this Green's function can be well approximated by its truncated version. This allows us to obtain an analytical traveling wave solution for an intercellular calcium wave and analyze the speed of solitary wave propagation as a function of physiologically important system parameters. Periodic and irregular traveling waves can be also sustained by the proposed model.

  16. Changing patterns of gap junctional intercellular communication and connexin distribution in mouse epidermis and hair follicles during embryonic development.

    PubMed

    Choudhry, R; Pitts, J D; Hodgins, M B

    1997-12-01

    In the mouse embryo between embryonic days 12 (E12) and 16, regular arrays of epidermal placodes on the mystacial pad develop into whisker follicles. This system was chosen for analysis of gap junctional intercellular communication during differentiation. The patterns of communication were studied by microinjection of the tracers Lucifer yellow-CH (LY-CH) and neurobiotin (NB), while immunofluorescent staining was used to study distribution of connexins 26 and 43. Extensive communication was seen between keratinocytes in developing hair pegs or, in later-stage hair follicles, in the germinative matrix. Coupling between adjacent hair pegs via interfollicular epidermis was not observed. Coupling also became restricted as follicular cells differentiated to form outer root sheath, inner root sheath, and hair shaft. Extensive gap junctional coupling is characteristic of keratinocytes that are rapidly proliferating (as in hair pegs and germinative matrix). Follicular keratinocytes commence differentiation shortly before restriction of gap junctional coupling becomes evident. Dermal mesenchymal cells undergoing different modes of differentiation also exhibit differences in gap junctional coupling, as evidenced by poor transfer of LY-CH between cells in dermal condensations of hair follicles compared with extensive transfer elsewhere in the dermis. LY-CH and NB were not transferred between epidermal or follicular epithelium and mesenchyme, arguing against a direct role for gap junctions permeable to known second messenger molecules or nucleotides in epithelial-mesenchymal interactions in this system. The distribution of connexins 26 and 43 in epidermis and hair follicles changed during differentiation but there was no correlation with changing patterns of dye transfer, indicating an unexpected degree of complexity in the relationship between gap junctional intercellular communication and connexin protein distribution during development.

  17. ESCRT-III assembly and cytokinetic abscission are induced by tension release in the intercellular bridge.

    PubMed

    Lafaurie-Janvore, Julie; Maiuri, Paolo; Wang, Irène; Pinot, Mathieu; Manneville, Jean-Baptiste; Betz, Timo; Balland, Martial; Piel, Matthieu

    2013-03-29

    The last step of cell division, cytokinesis, produces two daughter cells that remain connected by an intercellular bridge. This state often represents the longest stage of the division process. Severing the bridge (abscission) requires a well-described series of molecular events, but the trigger for abscission remains unknown. We found that pulling forces exerted by daughter cells on the intercellular bridge appear to regulate abscission. Counterintuitively, these forces prolonged connection, whereas a release of tension induced abscission. Tension release triggered the assembly of ESCRT-III (endosomal sorting complex required for transport-III), which was followed by membrane fission. This mechanism may allow daughter cells to remain connected until they have settled in their final locations, a process potentially important for tissue organization and morphogenesis.

  18. Estrogenic compounds inhibit gap junctional intercellular communication in mouse Leydig TM3 cells

    SciTech Connect

    Iwase, Yumiko . E-mail: Iwase.Yumiko@mg.m-pharma.co.jp; Fukata, Hideki . E-mail: fukata@faculty.chiba-u.jp; Mori, Chisato . E-mail: cmori@faculty.chiba-u.jp

    2006-05-01

    Some estrogenic compounds are reported to cause testicular disorders in humans and/or experimental animals by direct action on Leydig cells. In carcinogenesis and normal development, gap junctional intercellular communication (GJIC) plays an essential role in maintaining homeostasis. In this study, we examine the effects of diethylstilbestrol (DES, a synthetic estrogen), 17{beta}-estradiol (E{sub 2}, a natural estrogen), and genistein (GEN, a phytoestrogen) on GJIC between mouse Leydig TM3 cells using Lucifer yellow microinjection. The three compounds tested produced GJIC inhibition in the TM3 cells after 24 h. Gradually, 10 {mu}M DES began to inhibit GJIC for 24 h and this effect was observed until 72 h. On the other hand, both 20 {mu}M E{sub 2} and 25 {mu}M GEN rapidly inhibited GJIC in 6 h and 2 h, respectively. The effects continued until 24 h, but weakened by 72 h. Furthermore, a combined effect at {mu}M level between DES and E{sub 2} on GJIC inhibition was observed, but not between GEN and E{sub 2}. DES and E{sub 2} showed GJIC inhibition at low dose levels (nearly physiological estrogen levels) after 72 h, but GEN did not. DES-induced GJIC inhibition at 10 pM and 10 {mu}M was completely counteracted by ICI 182,780 (ICl), an estrogen receptor antagonist. On the other hand, the inhibitory effects on GJIC with E{sub 2} (10 pM and 20 {mu}M) and GEN (25 {mu}M) were partially blocked by ICI or calphostin C, a protein kinase C (PKC) inhibitor, and were completely blocked by the combination of ICI and calphostin C. These results demonstrate that DES inhibits GJIC between Leydig cells via the estrogen receptor (ER), and that E{sub 2} and GEN inhibit GJIC via ER and PKC. These estrogenic compounds may have different individual nongenotoxic mechanism including PKC pathway on testicular carcinogenesis or development.

  19. Noise magnetic fields abolish the gap junction intercellular communication suppression induced by 50 hz magnetic fields.

    PubMed

    Zeng, Qunli; Ke, Xueqin; Gao, Xiangwei; Fu, Yiti; Lu, Deqiang; Chiang, Huai; Xu, Zhengping

    2006-05-01

    Previously, we have reported that exposure to 50 Hz coherent sinusoidal magnetic fields (MF) for 24 h inhibits gap junction intercellular communication (GJIC) in mammalian cells at an intensity of 0.4 mT and enhances the inhibition effect of 12-O-tetradecanoylphorbol-13-acetate (TPA) at 0.2 mT. In the present study, we further explored the effects of incoherent noise MF on MF-induced GJIC inhibition. GJIC was determined by fluorescence recovery after photobleaching (FRAP) with a laser-scanning confocal microscope. The rate of fluorescence recovery (R) at 10 min after photobleaching was adopted as the functional index of GJIC. The R-value of NIH3T3 cells exposed to 50 Hz sinusoidal MF at 0.4 mT for 24 h was 30.85 +/- 14.70%, while the cells in sham exposure group had an R-value of 46.36 +/- 20.68%, demonstrating that the GJIC of NIH3T3 cells was significantly inhibited by MF exposure (P < .05). However, there were no significant differences in the R-values of the sham exposure, MF-plus-noise MF exposure (R: 49.58 +/- 19.38%), and noise MF exposure groups (R: 46.74 +/- 21.14%) (P > .05), indicating that the superposition of a noise MF alleviated the suppression of GJIC induced by the 50 Hz MF. In addition, although MF at an intensity of 0.2 mT synergistically enhanced TPA-induced GJIC inhibition (R: 24.90 +/- 13.50% vs. 35.82 +/- 17.18%, P < .05), further imposition of a noise MF abolished the synergistic effect of coherent MF (R: 32.51 +/- 18.37%). Overall, the present data clearly showed that although noise MF itself had no effect on GJIC of NIH3T3 cells, its superposition onto a coherent sinusoidal MF at the same intensity abolished MF-induced GJIC suppression. This is the first report showing that noise MF neutralizes 50 Hz MF-induced biological effect by using a signaling component as the test endpoint.

  20. Mouse Hepatitis Virus Infection Remodels Connexin43-Mediated Gap Junction Intercellular Communication In Vitro and In Vivo

    PubMed Central

    Basu, Rahul; Banerjee, Kaveri; Bose, Abhishek

    2015-01-01

    ABSTRACT Gap junctions (GJs) form intercellular channels which directly connect the cytoplasm between neighboring cells to facilitate the transfer of ions and small molecules. GJs play a major role in the pathogenesis of infection-associated inflammation. Mutations of gap junction proteins, connexins (Cxs), cause dysmyelination and leukoencephalopathy. In multiple sclerosis (MS) patients and its animal model experimental autoimmune encephalitis (EAE), Cx43 was shown to be modulated in the central nervous system (CNS). The mechanism behind Cx43 alteration and its role in MS remains unexplored. Mouse hepatitis virus (MHV) infection-induced demyelination is one of the best-studied experimental animal models for MS. Our studies demonstrated that MHV infection downregulated Cx43 expression at protein and mRNA levels in vitro in primary astrocytes obtained from neonatal mouse brains. After infection, a significant amount of Cx43 was retained in endoplasmic reticulum/endoplasmic reticulum Golgi intermediate complex (ER/ERGIC) and GJ plaque formation was impaired at the cell surface, as evidenced by a reduction of the Triton X-100 insoluble fraction of Cx43. Altered trafficking and impairment of GJ plaque formation may cause the loss of functional channel formation in MHV-infected primary astrocytes, as demonstrated by a reduced number of dye-coupled cells after a scrape-loading Lucifer yellow dye transfer assay. Upon MHV infection, a significant downregulation of Cx43 was observed in the virus-infected mouse brain. This study demonstrates that astrocytic Cx43 expression and function can be modulated due to virus stress and can be an appropriate model to understand the basis of cellular mechanisms involved in the alteration of gap junction intercellular communication (GJIC) in CNS neuroinflammation. IMPORTANCE We found that MHV infection leads to the downregulation of Cx43 in vivo in the CNS. In addition, results show that MHV infection impairs Cx43 expression in addition

  1. The B[a]P-increased intercellular communication via translocation of connexin-43 into gap junctions reduces apoptosis

    SciTech Connect

    Tekpli, X.; Rivedal, E.; Gorria, M.; Landvik, N.E.; Rissel, M.; Dimanche-Boitrel, M.-T.; Baffet, G.; Holme, J.A.; Lagadic-Gossmann, D.

    2010-01-15

    Gap junctions are channels in plasma membrane composed of proteins called connexins. These channels are organized in special domains between cells, and provide for direct gap junctional intercellular communication (GJIC), allowing diffusion of signalling molecules < 1 kD. GJIC regulates cell homeostasis and notably the balance between proliferation, cell cycle arrest, cell survival and apoptosis. Here, we have investigated benzo[a]pyrene (B[a]P) effects on GJIC and on the subcellular localization of the major protein of gap junction: connexin-43 (Cx43). Our results showed that B[a]P increased GJIC between mouse hepatoma Hepa1c1c7 cells via translocation of Cx43 from Golgi apparatus and lipid rafts into gap junction plaques. Interestingly, inhibition of GJIC by chlordane or small interference RNA directed against Cx43 enhanced B[a]P-induced apoptosis in Hepa1c1c7 cells. The increased apoptosis caused by inhibition of GJIC appeared to be mediated by ERK/MAPK pathway. It is suggested that B[a]P could induce transfer of cell survival signal or dilute cell death signal via regulation of ERK/MAPK through GJIC.

  2. Actin-interacting protein 1 controls assembly and permeability of intestinal epithelial apical junctions

    PubMed Central

    Baranwal, Somesh

    2015-01-01

    Adherens junctions (AJs) and tight junctions (TJs) are crucial regulators of the integrity and restitution of the intestinal epithelial barrier. The structure and function of epithelial junctions depend on their association with the cortical actin cytoskeleton that, in polarized epithelial cells, is represented by a prominent perijunctional actomyosin belt. The assembly and stability of the perijunctional cytoskeleton is controlled by constant turnover (disassembly and reassembly) of actin filaments. Actin-interacting protein (Aip) 1 is an emerging regulator of the actin cytoskeleton, playing a critical role in filament disassembly. In this study, we examined the roles of Aip1 in regulating the structure and remodeling of AJs and TJs in human intestinal epithelium. Aip1 was enriched at apical junctions in polarized human intestinal epithelial cells and normal mouse colonic mucosa. Knockdown of Aip1 by RNA interference increased the paracellular permeability of epithelial cell monolayers, decreased recruitment of AJ/TJ proteins to steady-state intercellular contacts, and attenuated junctional reassembly in a calcium-switch model. The observed defects of AJ/TJ structure and functions were accompanied by abnormal organization and dynamics of the perijunctional F-actin cytoskeleton. Moreover, loss of Aip1 impaired the apico-basal polarity of intestinal epithelial cell monolayers and inhibited formation of polarized epithelial cysts in 3-D Matrigel. Our findings demonstrate a previously unanticipated role of Aip1 in regulating the structure and remodeling of intestinal epithelial junctions and early steps of epithelial morphogenesis. PMID:25792565

  3. Cross regulation of intercellular gap junction communication and paracrine signaling pathways during organogenesis in Drosophila.

    PubMed

    Lechner, Hildegard; Josten, Frank; Fuss, Bernhard; Bauer, Reinhard; Hoch, Michael

    2007-10-01

    The spatial and temporal coordination of patterning and morphogenesis is often achieved by paracrine morphogen signals or by the direct coupling of cells via gap junctions. How paracrine signals and gap junction communication cooperate to control the coordinated behavior of cells and tissues is mostly unknown. We found that hedgehog signaling is required for the expression of wingless and of Delta/Notch target genes in a single row of boundary cells in the foregut-associated proventriculus organ of the Drosophila embryo. These cells coordinate the movement and folding of proventricular cells to generate a multilayered organ. hedgehog and wingless regulate gap junction communication by transcriptionally activating the innexin2 gene, which encodes a member of the innexin family of gap junction proteins. In innexin2 mutants, gap junction-mediated cell-to-cell communication is strongly reduced and the proventricular cell layers fail to fold and invaginate, similarly as in hedgehog or wingless mutants. We further found that innexin2 is required in a feedback loop for the transcriptional activation of the hedgehog and wingless morphogens and of Delta in the proventriculus primordium. We propose that the transcriptional cross regulation of paracrine and gap junction-mediated signaling is essential for organogenesis in Drosophila.

  4. Regulation of connexin 43-mediated gap junctional intercellular communication by Ca2+ in mouse epidermal cells is controlled by E- cadherin

    PubMed Central

    1991-01-01

    Gap junctional intercellular communication (GJIC) of cultured mouse epidermal cells is mediated by a gap junction protein, connexin 43, and is dependent on the calcium concentration in the medium, with higher GJIC in a high-calcium (1.2 mM) medium. In several mouse epidermal cell lines, we found a good correlation between the level of GJIC and that of immunohistochemical staining of E-cadherin, a calcium-dependent cell adhesion molecule, at cell-cell contact areas. The variant cell line P3/22 showed both low GJIC and E-cadherin protein expression in low- and high-Ca2+ media. P3/22 cells showed very low E-cadherin mRNA expression. To test directly whether E-cadherin is involved in the Ca(2+)-dependent regulation of GJIC, we transfected the E-cadherin expression vector into P3/22 cells and obtained several stable clones which expressed high levels of E-cadherin mRNA. All transfectants expressed E-cadherin molecules at cell-cell contact areas in a calcium- dependent manner. GJIC was also observed in these transfectants and was calcium dependent. These results suggest that Ca(2+)-dependent regulation of GJIC in mouse epidermal cells is directly controlled by a calcium-dependent cell adhesion molecule, E-cadherin. Furthermore, several lines of evidence suggest that GJIC control by E-cadherin involves posttranslational regulation (assembly and/or function) of the gap junction protein connexin 43. PMID:1650371

  5. Subsets of ATP-sensitive potassium channel (KATP) inhibitors increase gap junctional intercellular communication in metastatic cancer cell lines independent of SUR expression

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Gap junctional intercellular communication (GJIC) is a process whereby cells share molecules and nutrients with each other by physical contact through cell membrane pores. In tumor cells, GJIC is often altered, suggesting that this process may be important in the context of cancer. Certain ion chan...

  6. Intercellular communication in sensory ganglia by purinergic receptors and gap junctions: implications for chronic pain.

    PubMed

    Hanani, Menachem

    2012-12-03

    Peripheral injury can cause abnormal activity in sensory neurons, which is a major factor in chronic pain. Recent work has shown that injury induces major changes not only in sensory neurons but also in the main type of glial cells in sensory ganglia-satellite glial cells (SGCs), and that interactions between sensory neurons and SGCs contribute to neuronal activity in pain models. The main functional changes observed in SGCs after injury are an increased gap junction-mediated coupling among these cells, and augmented sensitivity to ATP. There is evidence that the augmented gap junctions contribute to neuronal hyperexcitability in pain models, but the mechanism underlying this effect is not known. The changes in SGCs described above have been found following a wide range of injuries (both axotomy and inflammation) in somatic, orofacial and visceral regions, and therefore appear to be a general feature in chronic pain. We have found that in cultures of sensory ganglia calcium signals can spread from an SGC to neighboring cells by calcium waves, which are mediated by gap junctions and ATP acting on purinergic P2 receptors. A model is proposed to explain how augmented gap junctions and greater sensitivity to ATP can combine to produce enhanced calcium waves, which can lead to neuronal excitation. Thus this simple scheme can account for several major changes in sensory ganglia that are common to a great variety of pain models.

  7. The interaction of Streptococcus pneumoniae with plasmin mediates transmigration across endothelial and epithelial monolayers by intercellular junction cleavage.

    PubMed

    Attali, Cécile; Durmort, Claire; Vernet, Thierry; Di Guilmi, Anne Marie

    2008-11-01

    for the plasminogen recruitment at the bacterial surface in facilitating adherence of pneumococci to endothelial and epithelial cells, while active plasmin degrades intercellular junctions. This process promotes migration of pneumococci through cell barriers by a pericellular route, a prerequisite for dissemination of S. pneumoniae in the host organism.

  8. Effects of estrogen and androgen on the ultrastructure of secretory granules and intercellular junctions in regressed canine prostate.

    PubMed

    Merk, F B; Leav, I; Kwan, P W; Ofner, P

    1980-06-01

    Epithelial cells in the prostate of the castrated or hypophysectomized dog were studied by thin-section and freeze-fracture electron microscopy to determine in vivo responses to estradiol-17 beta 17-cyclopentylpropionate (ECP) and testosterone cyclopentylpropionate (TCP). Particular attention was given to changes in specific organelles and intercellular junctions that might reflect hormone action. The few secretory granules that remain in the regressed epithelium (vestigial granules) serve as markers of prior androgen responsiveness. Pharmacologic doses of ECP caused regressed glandular cells to acquire a novel phenotype. Characteristic features of these estrogen-modified glandular (EMG) cells are newly formed secretory granules and tonofilament bundles that coexist with vestigial granules, thus demonstrating bipotentiality of response. Glandular cell-tight junctions appear unaltered by the endocrine manipulations. Although EMG cells have squamous cell features, tight junctions remain intact. Desmosomes in the canine prostate are dimorphic and are classified 70F and 100F according to the width of the filaments that converge on the dense plaques. In intact dogs, 100F desmosomes are associated with basal-reserve cells, whereas only the 70F variety is found between glandular cells. TCP treatment does not alter this distribution. Following ECP administration, both 70F and 100F desmosomes are present between EMG cells. The coexistence of newly formed secretory granules and tonofilaments of 100F desmosomes in the same EMG cell represents estrogen-induced bidirectional differentiation. Our findings indicate that androgens and estrogens are individually capable of controlling the expression of secretory granules and desmosomes. In intact animals, male and female sex hormones may act in concert to direct epithelial cell differentiation of the prostate.

  9. Lateral assembly of N-cadherin drives tissue integrity by stabilizing adherens junctions

    PubMed Central

    Garg, S.; Fischer, S. C.; Schuman, E. M.; Stelzer, E. H. K.

    2015-01-01

    Cadherin interactions ensure the correct registry and anchorage of cells during tissue formation. Along the plasma membrane, cadherins form inter-junctional lattices via cis- and trans-dimerization. While structural studies have provided models for cadherin interactions, the molecular nature of cadherin binding in vivo remains unexplored. We undertook a multi-disciplinary approach combining live cell imaging of three-dimensional cell assemblies (spheroids) with a computational model to study the dynamics of N-cadherin interactions. Using a loss-of-function strategy, we demonstrate that each N-cadherin interface plays a distinct role in spheroid formation. We found that cis-dimerization is not a prerequisite for trans-interactions, but rather modulates trans-interfaces to ensure tissue stability. Using a model of N-cadherin junction dynamics, we show that the absence of cis-interactions results in low junction stability and loss of tissue integrity. By quantifying the binding and unbinding dynamics of the N-cadherin binding interfaces, we determined that mutating either interface results in a 10-fold increase in the dissociation constant. These findings provide new quantitative information on the steps driving cadherin intercellular adhesion and demonstrate the role of cis-interactions in junction stability. PMID:25589573

  10. ASC Induces Apoptosis via Activation of Caspase-9 by Enhancing Gap Junction-Mediated Intercellular Communication.

    PubMed

    Kitazawa, Masato; Hida, Shigeaki; Fujii, Chifumi; Taniguchi, Shun'ichiro; Ito, Kensuke; Matsumura, Tomio; Okada, Nagisa; Sakaizawa, Takashi; Kobayashi, Akira; Takeoka, Michiko; Miyagawa, Shin-Ichi

    2017-01-01

    ASC (apoptosis-associated speck-like protein containing a CARD) is a key adaptor molecule of inflammasomes that mediates inflammatory and apoptotic signals. Aberrant methylation-induced silencing of ASC has been observed in a variety of cancer cells, thus implicating ASC in tumor suppression, although this role remains incompletely defined especially in the context of closely neighboring cell proliferation. As ASC has been confirmed to be silenced by abnormal methylation in HT1080 fibrosarcoma cells as well, this cell line was investigated to characterize the precise role and mechanism of ASC in tumor progression. The effects of ASC were examined using in vitro cell cultures based on comparisons between low and high cell density conditions as well as in a xenograft murine model. ASC overexpression was established by insertion of the ASC gene into pcDNA3 and pMX-IRES-GFP vectors, the latter being packed into a retrovirus and subjected to reproducible competitive assays using parental cells as an internal control, for evaluation of cell viability. p21 and p53 were silenced using shRNA. Cell viability was suppressed in ASC-expressing transfectants as compared with control cells at high cell density conditions in in vitro culture and colony formation assays and in in vivo ectopic tumor formation trials. This suppression was not detected in low cell density conditions. Furthermore, remarkable progression of apoptosis was observed in ASC-introduced cells at a high cell density, but not at a low one. ASC-dependent apoptosis was mediated not by p21, p53, or caspase-1, but rather by cleavage of caspase-9 as well as by suppression of the NF-κB-related X-linked inhibitor-of-apoptosis protein. Caspase-9 cleavage was observed to be dependent on gap junction formation. The remarkable effect of ASC on the induction of apoptosis through caspase-9 and gap junctions revealed in this study may lead to promising new approaches in anticancer therapy.

  11. Autophagy and gap junctional intercellular communication inhibition are involved in cadmium-induced apoptosis in rat liver cells

    SciTech Connect

    Zou, Hui; Zhuo, Liling; Han, Tao; Hu, Di; Yang, Xiaokang; Wang, Yi; Yuan, Yan; Gu, Jianhong; Bian, Jianchun; Liu, Xuezhong; Liu, Zongping

    2015-04-17

    Cadmium (Cd) is known to induce hepatotoxicity, yet the underlying mechanism of how this occurs is not fully understood. In this study, Cd-induced apoptosis was demonstrated in rat liver cells (BRL 3A) with apoptotic nuclear morphological changes and a decrease in cell index (CI) in a time- and concentration-dependent manner. The role of gap junctional intercellular communication (GJIC) and autophagy in Cd-induced apoptosis was investigated. Cd significantly induced GJIC inhibition as well as downregulation of connexin 43 (Cx43). The prototypical gap junction blocker carbenoxolone disodium (CBX) exacerbated the Cd-induced decrease in CI. Cd treatment was also found to cause autophagy, with an increase in mRNA expression of autophagy-related genes Atg-5, Atg-7, Beclin-1, and microtubule-associated protein light chain 3 (LC3) conversion from cytosolic LC3-I to membrane-bound LC3-II. The autophagic inducer rapamycin (RAP) prevented the Cd-induced CI decrease, while the autophagic inhibitor chloroquine (CQ) caused a further reduction in CI. In addition, CBX promoted Cd-induced autophagy, as well as changes in expression of Atg-5, Atg-7, Beclin-1 and LC3. CQ was found to block the Cd-induced decrease in Cx43 and GJIC inhibition, whereas RAP had opposite effect. These results demonstrate that autophagy plays a protective role during Cd-induced apoptosis in BRL 3A cells during 6 h of experiment, while autophagy exacerbates Cd-induced GJIC inhibition which has a negative effect on cellular fate. - Highlights: • GJIC and autophagy is crucial for biological processes. • Cd exposure causes GJIC inhibition and autophagy increase in BRL 3A cells. • Autophagy protects Cd induced BRL 3A cells apoptosis at an early stage. • Autophagy exacerbates Cd-induced GJIC inhibition. • GJIC plays an important role in autophagy induced cell death or survival.

  12. Influence of gap junction intercellular communication composed of connexin 43 on the antineoplastic effect of adriamycin in breast cancer cells

    PubMed Central

    Jiang, Guojun; Dong, Shuying; Yu, Meiling; Han, Xi; Zheng, Chao; Zhu, Xiaoguang; Tong, Xuhui

    2017-01-01

    Gap junctions (GJs) serve the principal role in the antineoplastic (cytotoxicity and induced apoptosis) effect of chemical drugs. The aim of the present study was to determine the effect of GJ intercellular communication (GJIC) composed of connexin 43 (Cx43) on adriamycin cytotoxicity in breast cancer cells. Four cell lines (Hs578T, MCF-7, MDA-MB-231 and SK-BR-3) with different degree of malignancy were used in the study. The results of western blotting and immunofluorescence revealed that, in Hs578T and MCF-7 cells, which have a low degree of malignancy, the expression levels of Cx43 and GJIC were higher than those in MDA-MB-231 and SK-BR-3 cells (which have a high degree of malignancy). In Hs578T and MCF-7 cells, where GJ could be formed, the function of GJ was modulated by a pharmacological potentiators [retinoid acid (RA)]/inhibitors [oleamide and 18-α-glycyrrhetinic acid (18-α-GA)] and small interfering RNA (siRNA). In high-density cells (where GJ was formed), enhancement of GJ function by RA increased the cytotoxicity of adriamycin, while inhibition of GJ function by oleamide/18-α-GA and siRNA decreased the cytotoxicity caused by adriamycin. Notably, the modulation of GJ did not affect the survival of cells treated with adriamycin when cells were in low density (no GJ was formed). The present study illustrated the association between GJIC and the antitumor effect of adriamycin in breast cancer cells. The cytotoxicity of adriamycin on breast cancer cells was increased when the function of gap junctions was enhanced. PMID:28356970

  13. ASC Induces Apoptosis via Activation of Caspase-9 by Enhancing Gap Junction-Mediated Intercellular Communication

    PubMed Central

    Hida, Shigeaki; Fujii, Chifumi; Taniguchi, Shun’ichiro; Ito, Kensuke; Matsumura, Tomio; Okada, Nagisa; Sakaizawa, Takashi; Kobayashi, Akira; Takeoka, Michiko; Miyagawa, Shin-ichi

    2017-01-01

    ASC (apoptosis-associated speck-like protein containing a CARD) is a key adaptor molecule of inflammasomes that mediates inflammatory and apoptotic signals. Aberrant methylation-induced silencing of ASC has been observed in a variety of cancer cells, thus implicating ASC in tumor suppression, although this role remains incompletely defined especially in the context of closely neighboring cell proliferation. As ASC has been confirmed to be silenced by abnormal methylation in HT1080 fibrosarcoma cells as well, this cell line was investigated to characterize the precise role and mechanism of ASC in tumor progression. The effects of ASC were examined using in vitro cell cultures based on comparisons between low and high cell density conditions as well as in a xenograft murine model. ASC overexpression was established by insertion of the ASC gene into pcDNA3 and pMX-IRES-GFP vectors, the latter being packed into a retrovirus and subjected to reproducible competitive assays using parental cells as an internal control, for evaluation of cell viability. p21 and p53 were silenced using shRNA. Cell viability was suppressed in ASC-expressing transfectants as compared with control cells at high cell density conditions in in vitro culture and colony formation assays and in in vivo ectopic tumor formation trials. This suppression was not detected in low cell density conditions. Furthermore, remarkable progression of apoptosis was observed in ASC-introduced cells at a high cell density, but not at a low one. ASC-dependent apoptosis was mediated not by p21, p53, or caspase-1, but rather by cleavage of caspase-9 as well as by suppression of the NF-κB-related X-linked inhibitor-of-apoptosis protein. Caspase-9 cleavage was observed to be dependent on gap junction formation. The remarkable effect of ASC on the induction of apoptosis through caspase-9 and gap junctions revealed in this study may lead to promising new approaches in anticancer therapy. PMID:28056049

  14. Effects of cathepsin K deficiency on intercellular junction proteins, luminal mucus layers, and extracellular matrix constituents in the mouse colon.

    PubMed

    Arampatzidou, Maria; Schütte, André; Hansson, Gunnar C; Saftig, Paul; Brix, Klaudia

    2012-12-01

    Cathepsin K has been shown to exhibit antimicrobial and anti-inflammatory activities in the mouse colon. To further elucidate its role, we used Ctsk-/- mice and demonstrated that the absence of cathepsin K was accompanied by elevated protein levels of related cysteine cathepsins (cathepsins B, L, and X) in the colon. In principle, such changes could result in altered subcellular localization; however, the trafficking of cysteine cathepsins was not affected in the colon of Ctsk-/- mice. However, cathepsin K deficiency affected the extracellular matrix constituents, as higher amounts of collagen IV and laminin were observed. Moreover, the localization pattern of the intercellular junction proteins E-cadherin and occludin was altered in the colon of Ctsk-/- mice, suggesting potential impairment of the barrier function. Thus, we used an ex vivo method for assessing the mucus layers and showed that the absence of cathepsin K had no influence on mucus organization and growth. The data of this study support the notion that cathepsin K contributes to intestinal homeostasis and tissue architecture, but the lack of cathepsin K activity is not expected to affect the mucus-depending barrier functions of the mouse colon. These results are important with regard to oral administration of cathepsin K inhibitors that are currently under investigation in clinical trials.

  15. Effect of perfluorooctane sulfonate on viability, maturation and gap junctional intercellular communication of porcine oocytes in vitro.

    PubMed

    Domínguez, A; Salazar, Z; Arenas, E; Betancourt, M; Ducolomb, Y; González-Márquez, H; Casas, E; Teteltitla, M; Bonilla, E

    2016-09-01

    Perfluorooctane sulfonate (PFOS) is a broadly used man-made surfactant whose long half-life has led to bioaccumulation. This perfluorinated compound is ubiquitous in human body fluids. PFOS concentrations as high as 26μM in plasma have been reported in occupationally exposed populations, and high levels of PFOS in human follicular fluid have been associated with subfertility. However, the effect of PFOS on the maturation of oocytes in mammals has not been reported to date. The aim of this study was to determine the effects of PFOS during oocyte maturation. Results indicate that PFOS inhibits oocyte viability (Lethal Concentration50=32μM) and maturation (inhibition of maturation50=22μM) at physiologically relevant concentrations. In order to evaluate the mechanisms of oocyte maturation inhibition by PFOS, gap junctional intercellular communication (GJIC) between oocytes and granulosa cells was assessed. GJIC between granulosa cells and the oocyte was significantly affected during the first 8h of maturation. However, the inhibitory effect of PFOS on GJIC was not due to an alteration on the expression of connexin genes Cx43, Cx45 and Cx60. These findings suggest that occupationally exposed populations could be at risk, and that PFOS might affect oocyte maturation by interfering the GJIC in the cumulus-oocyte complexes during the first hours of maturation.

  16. Effects of phenolics in Empire apples on hydrogen peroxide-induced inhibition of gap-junctional intercellular communication.

    PubMed

    Lee, Ki Won; Lee, Sang Jun; Kang, Nam Joo; Lee, Chang Yong; Lee, Hyong Joo

    2004-01-01

    The present study investigated antioxidant and antitumor-promoting activities of major phenolic phytochemicals of apples. The contents of each antioxidant in Empire apples was quantified and their contributions to total antioxidant activity of apples were determined using assay for inhibition of 12-O-tetradecanoylphorbol-13-acetate-induced superoxide radical generation in cell culture model and expressed in vitamin C equivalent antioxidant capacity (VCEAC). The estimated contribution of major phenolics and vitamin C to total anitoxidant capacity of 100 g fresh Empire apples is as follows: quercetin (60.05 VCEAC) > chlorogenic acid (12.32) > phloretin (7.41) > procyanidin B2 (7.22) > vitamin C (6.61) > epicatechin (5.10) in superoxide radical scavenging assay. Recent reports suggest that the mechanism of carcinogenic process of hydrogen peroxide (H2O2) may be associated with the inhibition of gap-junctional intercellular communication (GJIC), which is involved in tumor promotion process. Apple extracts showed the protective effects against the inhibition of GJIC by H2O2 in a dose-dependent manner. Quercetin exerted the strongest protective effects among major antioxidants in apples on H2O2-induced inhibition of GJIC, following epicatechin, procyanidin B2, and vitamin C, while chlorogenic acid and phloretin had no effects. Our results indicate that cancer chemopreventive activity of apples is associated with the combined antioxidant capacity and antitumor-promoting activities of diverse antioxidants.

  17. Supercritical CO(2)-extracted tomato Oleoresins enhance gap junction intercellular communications and recover from mercury chloride inhibition in keratinocytes.

    PubMed

    Leone, Antonella; Zefferino, Roberto; Longo, Cristiano; Leo, Lucia; Zacheo, Giuseppe

    2010-04-28

    A nutritionally relevant phytochemical such as lycopene, found in tomatoes and other fruits, has been proposed to have health-promoting effects by modulating hormonal and immune systems, metabolic pathways, and gap junction intercellular communication (GJIC). This work analyzes lycopene extracts, obtained from tomato and tomato added with grape seeds by using a safe and environmentally friendly extraction process, based on supercritical carbon dioxide technology (S-CO(2)). Analysis of the innovative S-CO(2)-extracted oleoresins showed peculiar chemical composition with high lycopene concentration and the presence of other carotenoids, lipids, and phenol compounds. The oleoresins showed a higher in vitro antioxidant activity compared with pure lycopene and beta-carotene and the remarkable ability to enhance the GJIC and to increase cx43 expression in keratinocytes. The oleoresins, (0.9 microM lycopene), were also able to overcome, completely, the GJIC inhibition induced by 10 nM HgCl(2), mercury(II) chloride, suggesting a possible action mechanism.

  18. Involvement of gap junctional intercellular communication in the bystander effect induced by broad-beam or microbeam heavy ions

    NASA Astrophysics Data System (ADS)

    Shao, Chunlin; Furusawa, Yoshiya; Kobayashi, Yasuhiko; Funayama, Tomoo

    2006-09-01

    Most of the reported bystander responses were studied by using low dose irradiation of γ-rays and light ions such as alpha-particles. In this study, primary human fibroblasts AG1522 in confluent cultures were irradiated with either broad-beam of 100 keV/μm 12C or microbeams of 380 keV/μm 20Ne and 1260 keV/μm 40Ar. When cells were irradiated with 12C ions, the induction of micronucleus (MN) had a low-dose sensitive effect, i.e. a lower dose of irradiation gave a higher yield of MN per cell-traversal. This phenomenon was further reinforced by using a microbeam to irradiate a fraction of cells within a population. Even when only a single cell was targeted with one particle of 40Ar or 20Ne, the MN yield was increased to 1.4-fold of the non-irradiated control. When the number of microbeam targeted cells increased, the MN yield per targeted-cell decreased drastically. In addition, the bystander MN induction did not vary significantly with the number and the linear energy transfer (LET) of microbeam particles. When the culture was treated with PMA, an inhibitor of gap junctional intercellular communication (GJIC), MN induction was decreased for both microbeam and broad-beam irradiations even at high-doses where all cells were hit. The present findings indicate that a GJIC-mediated signaling amplification mechanism was involved in the high-LET heavy ion irradiation induced bystander effect. Moreover, at high-doses of radiation, the bystander signals could perform a complex interaction with direct irradiation.

  19. Inhibition of gap junction intercellular communication by extremely low-frequency electromagnetic fields in osteoblast-like models is dependent on cell differentiation.

    PubMed

    Yamaguchi, Dean T; Huang, Jason; Ma, Defang; Wang, Paul K C

    2002-02-01

    Electromagnetic fields have been used to augment the healing of fractures because of its ability to increase new bone formation. The mechanism of how electromagnetic fields can promote new bone formation is unknown, although the interaction of electromagnetic fields with components of the plasma membrane of cells has been hypothesized to occur in bone cells. Gap junctions occur among bone forming cells, the osteoblasts, and have been hypothesized to play a role in new bone formation. Thus it was investigated whether extremely low-frequency (ELF) magnetic fields alter gap junction intercellular communication in the pre-osteoblastic model, MC3T3-E1, and the well-differentiated osteoblastic model, ROS 17/2.8. ELF magnetic field exposure systems were designed to be used for an inverted microscope stage and for a tissue culture incubator. Using these systems, it was found that magnetic fields over a frequency range from 30 to 120 Hz and field intensities up to 12.5 G dose dependently decreased gap junction intercellular communication in MC3T3-E1 cells during their proliferative phase of development. The total amount of connexin 43 protein and the distribution of connexin 43 gap junction protein between cytoplasmic and plasma membrane pools were unaltered by treatment with ELF magnetic fields. Cytosolic calcium ([Ca(2+)](i)) which can inhibit gap junction communication, was not altered by magnetic field exposure. Identical exposure conditions did not affect gap junction communication in the ROS 17/2.8 cell line and when MC3T3-E1 cells were more differentiated. Thus ELF magnetic fields may affect only less differentiated or pre-osteoblasts and not fully differentiated osteoblasts. Consequently, electromagnetic fields may aid in the repair of bone by effects exerted only on osteoprogenitor or pre-osteoblasts.

  20. Alix-mediated assembly of the actomyosin–tight junction polarity complex preserves epithelial polarity and epithelial barrier

    PubMed Central

    Campos, Yvan; Qiu, Xiaohui; Gomero, Elida; Wakefield, Randall; Horner, Linda; Brutkowski, Wojciech; Han, Young-Goo; Solecki, David; Frase, Sharon; Bongiovanni, Antonella; d'Azzo, Alessandra

    2016-01-01

    Maintenance of epithelial cell polarity and epithelial barrier relies on the spatial organization of the actin cytoskeleton and proper positioning/assembly of intercellular junctions. However, how these processes are regulated is poorly understood. Here we reveal a key role for the multifunctional protein Alix in both processes. In a knockout mouse model of Alix, we identified overt structural changes in the epithelium of the choroid plexus and in the ependyma, such as asymmetrical cell shape and size, misplacement and abnormal beating of cilia, blebbing of the microvilli. These defects culminate in excessive cell extrusion, enlargement of the lateral ventricles and hydrocephalus. Mechanistically, we find that by interacting with F-actin, the Par complex and ZO-1, Alix ensures the formation and maintenance of the apically restricted actomyosin–tight junction complex. We propose that in this capacity Alix plays a role in the establishment of apical–basal polarity and in the maintenance of the epithelial barrier. PMID:27336173

  1. Germ cell-somatic cell relationships: a comparative study of intercellular junctions during spermatogenesis in selected non-mammalian vertebrates.

    PubMed

    Sprando, R L; Russell, L D

    1987-09-01

    Specialized germ cell-somatic cell relationships were surveyed in the testis of species representative of four classes of non-mammalian vertebrates. Desmosome-like junctions were present in all classes studied. In the teleost fish studied (bluegill; Lepomis macrochirus), small, infrequent desmosomes, seen between the spherical cyst cells and spermatocytes, were characterized by poorly represented subsurface densities. In the bullfrog (Rana catesbeiana), similar desmosome-like junctions were found between cyst cell processes and spermatocytes. Reptilian (turtle; Pseudameys scripta) desmosome-like junctions between Sertoli cells and germ cells were heterogeneous and more numerous than those junctions found in fish and amphibians. In general, the reptilian desmosome-like junctions were extensive structures displaying 10 nm filaments associated with the Sertoli cell component of the junctions. Regions within the desmosome where the two plasma membranes converged suggested that gap junctions were a component of the desmosome-like junctions. "Desmosome-gap" junctions persisted in turtle spermatids for sometime after nuclear elongation had commenced. In birds (chicken; Gallus domesticus), "desmosome-gap" junctions, similar to those seen in turtles were described between both spermatocytes and Sertoli cells, and spermatids and Sertoli cells. These junctions were frequently lined by saccules of endoplasmic reticulum. The presence of gap junctions suggest the evolution of mechanisms for somatic cell-germ cell communication although more species should be examined to confirm this hypothesis.

  2. Self assembled silicon nanowire Schottky junction assisted by collagen

    NASA Astrophysics Data System (ADS)

    Stievenard, Didier; Sahli, Billel; Coffinier, Yannick; Boukherroub, Rabah; Melnyk, Oleg

    2008-03-01

    We present results on self assembled silicon nanowire Schottky junction assisted by collagen fibrous. The collagen is the principle protein of connective human tissues. It presents the double interest to be a low cost biological material with the possibility to be combed as the DNA molecule. First, the collagen was combed on OTS modified surface with gold electrodes. Second, silicon nanowires were grown on silicon substrate by CVD of silane gas (SiH4) at high temperature (500 C) using a vapor-liquid-solid (VLS) process and gold particles as catalysts. In order to increase electrostatic interaction between the collagen and the nanowires, these latters were chemically modified by mercaptopropylmethoxysilane (MPTS), then chemically oxidized. Therefore, the nanowires were transferred from their substrate into water and a drop of it deposited on the surface. Nanowires are only bound to collagen and in particular, in electrode gaps. The formation of spontaneous Schotkty junction is demonstrated by current-voltage characteristics.

  3. Resveratrol induces chemosensitization to 5-fluorouracil through up-regulation of intercellular junctions, Epithelial-to-mesenchymal transition and apoptosis in colorectal cancer.

    PubMed

    Buhrmann, Constanze; Shayan, Parviz; Kraehe, Patricia; Popper, Bastian; Goel, Ajay; Shakibaei, Mehdi

    2015-11-01

    5-Fluorouracil (5-FU), a common chemotherapeutic agent used for the treatment of colorectal cancer (CRC), by itself has inadequate response rates; highlighting the need for novel and improved treatment regimens for these patients. Resveratrol, a naturally-occurring polyphenol, has been linked with chemosensitizing potential and anticancer properties; however, the underlying mechanisms for these effects remain poorly understood. The effect of resveratrol in parental CRC cell lines (HCT116, SW480) and their corresponding isogenic 5-FU-chemoresistant derived clones (HCT116R, SW480R) was examined by MTT assays, intercellular junction formation and apoptosis by electron- and immunoelectron microscopy, nuclear factor-kappaB (NF-κB) and NF-κB regulated gene products by western blot analysis in a 3D-alginate microenvironment. Resveratrol blocked the proliferation of all four CRC cell lines and synergized the invasion inhibitory effects of 5-FU. Interestingly, resveratrol induced a transition from 5-FU-induced formation of microvilli to a planar cell surface, which was concomitant with up-regulation of desmosomes, gap- and tight junctions (claudin-2) and adhesion molecules (E-cadherin) expression in HCT116 and HCT116R cells. Further, resveratrol significantly attenuated drug resistance through inhibition of epithelial-mesenchymal transition (EMT) factors (decreased vimentin and slug, increased E-cadherin) and down-regulation of NF-κB activation and its translocation to the nucleus and abolished NF-κB-regulated gene end-products (MMP-9, caspase-3). Moreover, this suppression was mediated through inhibition of IκBα kinase and IκBα phosphorylation and degradation. Our results demonstrate that resveratrol can potentiate the anti-tumor effects of 5-FU on CRC cells by chemosensitizing them, inhibiting an EMT phenotype via up-regulation of intercellular junctions and by down-regulation of NF-κB pathway.

  4. Inhibition of gap junction intercellular communication is involved in silica nanoparticles-induced H9c2 cardiomyocytes apoptosis via the mitochondrial pathway

    PubMed Central

    Du, Zhong-jun; Cui, Guan-qun; Zhang, Juan; Liu, Xiao-mei; Zhang, Zhi-hu; Jia, Qiang; Ng, Jack C; Peng, Cheng; Bo, Cun-xiang; Shao, Hua

    2017-01-01

    Gap junction intercellular communication (GJIC) between cardiomyocytes is essential for synchronous heart contraction and relies on connexin-containing channels. Connexin 43 (Cx43) is a major component involved in GJIC in heart tissue, and its abnormal expression is closely associated with various cardiac diseases. Silica nanoparticles (SNPs) are known to induce cardiovascular toxicity. However, the mechanisms through which GJIC plays a role in cardiomyocytes apoptosis induced by SNPs remain unknown. The aim of the present study is to determine whether SNPs-decreased GJIC promotes apoptosis in rat cardiomyocytes cell line (H9c2 cells) via the mitochondrial pathway using CCK-8 Kit, scrape-loading dye transfer technique, Annexin V/PI double-staining assays, and Western blot analysis. The results showed that SNPs elicited cytotoxicity in H9c2 cells in a time- and concentration-dependent manner. SNPs also reduced GJIC in H9c2 cells in a concentration-dependent manner through downregulation of Cx43 and upregulation of P-Cx43. Inhibition of gap junctions by gap junction blocker carbenoxolone disodium resulted in decreased survival and increased apoptosis, whereas enhancement of the gap junctions by retinoic acid led to enhanced survival but decreased apoptosis. Furthermore, SNPs-induced apoptosis through the disrupted functional gap junction was correlated with abnormal expressions of the proteins involved in the mitochondrial pathway-related apoptosis such as Bcl-2/Bax, cytochrome C, Caspase-9, and Caspase-3. Taken together, our results provide the first evidence that SNPs-decreased GJIC promotes apoptosis in cardiomyocytes via the mitochondrial pathway. In addition, downregulation of GJIC by SNPs in cardiomyocytes is mediated through downregulation of Cx43 and upregulation of P-Cx43. These results suggest that in rat cardiomyocytes cell line, GJIC plays a protective role in SNPs-induced apoptosis and that GJIC may be one of the targets for SNPs-induced biological

  5. Tanshinone IIA increases the bystander effect of herpes simplex virus thymidine kinase/ganciclovir gene therapy via enhanced gap junctional intercellular communication.

    PubMed

    Xiao, Jianyong; Zhang, Guangxian; Qiu, Pengxiang; Liu, Xijuan; Wu, Yingya; Du, Biaoyan; Li, Jiefen; Zhou, Jing; Li, Jingjing; Tan, Yuhui

    2013-01-01

    The bystander effect is an intriguing phenomenon by which adjacent cells become sensitized to drug treatment during gene therapy with herpes simplex virus thymidine kinase/ganciclovir (HSV-tk/GCV). This effect is reported to be mediated by gap junctional intercellular communication (GJIC), and therefore, we postulated that upregulation of genes that facilitate GJIC may enhance the HSV-tk/GCV bystander effect. Previous findings have shown Tanshinone IIA (Tan IIA), a chemical substance derived from a Chinese medicine herb, promotes the upregulation of the connexins Cx26 and Cx43 in B16 cells. Because gap junctions are formed by connexins, we hypothesized that Tan IIA might increase GJIC. Our results show that Tan IIA increased GJIC in B16 melanoma cells, leading to more efficient GCV-induced bystander killing in cells stably expressing HSV-tk. Additionally, in vivo experiments demonstrated that tumors in mice with 10% HSV-tk positive B16 cells and 90% wild-type B16 cells became smaller following treatment with the combination of GCV and Tan IIA as compared to GCV or Tan IIA alone. These data demonstrate that Tan IIA can augment the bystander effect of HSV-tk/GCV system through increased gap junction coupling, which adds strength to the promising strategy that develops connexins inducer to potentiate the effects of suicide gene therapy.

  6. ROCK activity regulates functional tight junction assembly during blastocyst formation in porcine parthenogenetic embryos

    PubMed Central

    Kwon, Jeongwoo

    2016-01-01

    The Rho-associated coiled-coil-containing protein serine/threonine kinases 1 and 2 (ROCK1 and ROCK2) are Rho subfamily GTPase downstream effectors that regulate cell migration, intercellular adhesion, cell polarity, and cell proliferation by stimulating actin cytoskeleton reorganization. Inhibition of ROCK proteins affects specification of the trophectoderm (TE) and inner cell mass (ICM) lineages, compaction, and blastocyst cavitation. However, the molecules involved in blastocyst formation are not known. Here, we examined developmental competence and levels of adherens/tight junction (AJ/TJ) constituent proteins, such as CXADR, OCLN, TJP1, and CDH1, as well as expression of their respective mRNAs, after treating porcine parthenogenetic four-cell embryos with Y-27632, a specific inhibitor of ROCK, at concentrations of 0, 10, 20, 100 µM for 24 h. Following this treatment, the blastocyst development rates were 39.1, 20.7, 10.0, and 0% respectively. In embryos treated with 20 µM treatment, expression levels of CXADR, OCLN, TJP1, and CDH1 mRNA and protein molecules were significantly reduced (P < 0.05). FITC-dextran uptake assay revealed that the treatment caused an increase in TE TJ permeability. Interestingly, the majority of the four-cell and morula embryos treated with 20 µM Y-27643 for 24 h showed defective compaction and cavitation. Taken together, our results indicate that ROCK activity may differentially affect assembly of AJ/TJs as well as regulate expression of genes encoding junctional proteins. PMID:27077008

  7. Gap junctions in the ovary of Drosophila melanogaster: localization of innexins 1, 2, 3 and 4 and evidence for intercellular communication via innexin-2 containing channels

    PubMed Central

    Bohrmann, Johannes; Zimmermann, Jennifer

    2008-01-01

    Background In the Drosophila ovary, germ-line and soma cells are interconnected via gap junctions. The main gap-junction proteins in invertebrates are members of the innexin family. In order to reveal the role that innexins play in cell-cell communication during oogenesis, we investigated the localization of innexins 1, 2, 3 and 4 using immunohistochemistry, and analyzed follicle development following channel blockade. Results We found innexin 1 predominantly localized to the baso-lateral domain of follicle cells, whereas innexin 2 is positioned apico-laterally as well as apically between follicle cells and germ-line cells. Innexin 3 was observed laterally in follicle cells and also in nurse cells, and innexin 4 was detected in the oolemma up to stage 8 and in nurse-cell membranes up to stage 12. In order to test whether innexins form channels suitable for intercellular communication, we microinjected innexin antibodies in combination with a fluorescent tracer into the oocyte of stage-10 follicles. We found that dye-coupling between oocyte and follicle cells was largely reduced by innexin-2 antibodies directed against the intracellular C-terminus as well as against the intracellular loop. Analyzing in vitro, between stages 10 and 14, the developmental capacities of follicles following microinjections of innexin-2 antibodies revealed defects in follicle-cell differentiation, nurse-cell regression, oocyte growth and choriogenesis. Conclusion Our results suggest that all analyzed innexins are involved in the formation of gap junctions in the ovary. While innexins 2 and 3 are colocalized between soma cells, innexins 2 and 4 are colocalized between soma and germ-line cells. Innexin 2 is participating in cell-cell communication via hemichannels residing in the oolemma. It is obvious that gap-junctional communication between germ-line and soma cells is essential for several processes during oogenesis. PMID:19038051

  8. Evolution of Microbial Quorum Sensing to Human Global Quorum Sensing: An Insight into How Gap Junctional Intercellular Communication Might Be Linked to the Global Metabolic Disease Crisis

    PubMed Central

    Trosko, James E.

    2016-01-01

    The first anaerobic organism extracted energy for survival and reproduction from its source of nutrients, with the genetic means to ensure protection of its individual genome but also its species survival. While it had a means to communicate with its community via simple secreted molecules (“quorum sensing”), the eventual shift to an aerobic environment led to multi-cellular metazoan organisms, with evolutionary-selected genes to form extracellular matrices, stem cells, stem cell niches, and a family of gap junction or “connexin” genes. These germinal and somatic stem cells responded to extracellular signals that triggered intra-cellular signaling to regulate specific genes out of the total genome. These extra-cellular induced intra-cellular signals also modulated gap junctional intercellular communication (GJIC) in order to regulate the new cellular functions of symmetrical and asymmetrical cell division, cell differentiation, modes of cell death, and senescence. Within the hierarchical and cybernetic concepts, differentiated by neurons organized in the brain of the Homo sapiens, the conscious mind led to language, abstract ideas, technology, myth-making, scientific reasoning, and moral decision–making, i.e., the creation of culture. Over thousands of years, this has created the current collision between biological and cultural evolution, leading to the global “metabolic disease” crisis. PMID:27314399

  9. Evolution of Microbial Quorum Sensing to Human Global Quorum Sensing: An Insight into How Gap Junctional Intercellular Communication Might Be Linked to the Global Metabolic Disease Crisis.

    PubMed

    Trosko, James E

    2016-06-15

    The first anaerobic organism extracted energy for survival and reproduction from its source of nutrients, with the genetic means to ensure protection of its individual genome but also its species survival. While it had a means to communicate with its community via simple secreted molecules ("quorum sensing"), the eventual shift to an aerobic environment led to multi-cellular metazoan organisms, with evolutionary-selected genes to form extracellular matrices, stem cells, stem cell niches, and a family of gap junction or "connexin" genes. These germinal and somatic stem cells responded to extracellular signals that triggered intra-cellular signaling to regulate specific genes out of the total genome. These extra-cellular induced intra-cellular signals also modulated gap junctional intercellular communication (GJIC) in order to regulate the new cellular functions of symmetrical and asymmetrical cell division, cell differentiation, modes of cell death, and senescence. Within the hierarchical and cybernetic concepts, differentiated by neurons organized in the brain of the Homo sapiens, the conscious mind led to language, abstract ideas, technology, myth-making, scientific reasoning, and moral decision-making, i.e., the creation of culture. Over thousands of years, this has created the current collision between biological and cultural evolution, leading to the global "metabolic disease" crisis.

  10. Gap junctional intercellular communication as a biological "Rosetta stone" in understanding, in a systems biological manner, stem cell behavior, mechanisms of epigenetic toxicology, chemoprevention and chemotherapy.

    PubMed

    Trosko, James E

    2007-08-01

    In spite of the early speculation by Loewenstein that one of the critical distinguishing phenotypes of cancers from normal cells was the dysfunction of gap junctional intercellular communication (GJIC), this hypothesis has not captured the attention of most birth defects and cancer researchers. Moreover, even with later demonstrations that factors that influence normal development and carcinogenesis by modulating GJIC, such as chemical teratogens and tumor-promoting chemicals, inflammatory factors, hormones and growth factors, antisense connexin genes, knockout mouse models, human inherited mutated connexin genes, si-connexin RNA, chemopreventive and chemotherapeutic chemicals, it is rare that one sees any reference to these studies by the mainstream investigators in these fields. Based on the assumption that the evolutionarily conserved connexin genes found in metazoans are needed for normal development and the maintenance of health and T. Dobzhansky's statement "Nothing in biology makes sense except in the light of evolution," a short review of the roles of endogenous and exogenous modulators of GJIC will be made in the context of the multistage, multimechanism process of carcinogenesis, the stem cell theory of carcinogenesis, the discovery and characterization of normal adult stem "cancer stem" cells and the observation that two distinct classes of GJIC-deficient cancer cells are known. The implications of these observations to a "systems biological" view of the role of gap junctions and the nutritional prevention and treatment of several chronic diseases and cancer will be discussed.

  11. Analyzing phorbol ester effects on gap junctional communication: a dramatic inhibition of assembly

    PubMed Central

    1994-01-01

    The effect of 12-O-tetradeconylphorbol-13-acetate (TPA) on gap junction assembly between Novikoff hepatoma cells was examined. Cells were dissociated with EDTA to single cells and then reaggregated to form new junctions. When TPA (25 nM) was added to the cells at the onset of the 60-min reaggregation, dye transfer was detected at only 0.6% of the cell-cell interfaces compared to 72% for the untreated control and 74% for 4-alpha TPA, an inactive isomer of TPA. Freeze-fracture electron microscopy of reaggregated control cells showed interfaces containing an average of more than 600 aggregated intramembranous gap junction particles, while TPA-treated cells had no gap junctions. However, Lucifer yellow dye transfer between nondissociated cells via gap junctions was unaffected by 60 min of TPA treatment. Therefore, TPA dramatically inhibited gap junction assembly but did not alter channel gating nor enhance disassembly of preexisting gap junction structures. Short term TPA treatment (< 30 min) increased phosphorylation of the gap junction protein molecular weight of 43,000 (Cx43), but did not change the cellular level of Cx43. Cell surface biotinylation experiments suggested that TPA did not substantially reduce the plasma membrane concentration of Cx43. Therefore, the simple presence of Cx43 in the plasma membrane is not sufficient for gap junction assembly, and protein kinase C probably exerts an effect on assembly of gap junctions at the plasma membrane level. PMID:7806568

  12. The electrotonic location of low-resistance intercellular junctions between a pair of giant neurones in the snail Lymnaea.

    PubMed Central

    Benjamin, P R; Pilkington, J B

    1986-01-01

    The passive electrotonic properties of neurones VD1 and RPD2 in the brain of the snail Lymnaea can be represented by a soma-finite cable model with closed-circuit axon termination. There is a considerable individual variation in input resistance, membrane time constant, electrotonic length and axon-soma conductance ratio, but the average values for these parameters are similar in the two neurones. The cells are tightly coupled by an electrotonic synapse giving an average steady-state coupling coefficient of 0.68 and an average resistance measured between recording sites in the cell bodies of 20 M omega. Calculations using a model consisting of a symmetrical pair of cells with standard values for the electrotonic parameters show that in this system, for a soma-soma resistance of 20 M omega, the junction cannot be more than 0.16 length constants from the cell bodies. Reduction in coupling due to membrane current losses in such short proximal axon segments is insignificant. Intra-axonal recordings indicate that most of the coupling resistance is located at the junction between VD1 and RPD2, which must therefore be closer to the cell bodies than the limiting value of 0.16 length constants assuming an electrical equivalent model which includes the standard electrotonic parameters. If all the soma-soma resistance is located at the junction, then it could be physically a single array of gap-junction particles. Despite its low conductance (1/20 M omega = 50 nS) and possibly small physical dimensions, the electrotonic synapse is more than sufficient to ensure spike synchrony in the two cells. PMID:3958976

  13. Modulation of Intercellular Junctions by Cyclic-ADT Peptides as a Method to Reversibly Increase Blood-Brain Barrier Permeability

    PubMed Central

    Laksitorini, Marlyn D.; Kiptoo, Paul K.; On, Ngoc H.; Thliveris, James A.; Miller, Donald W.; Siahaan, Teruna J.

    2015-01-01

    It is challenging to deliver molecules to the brain for diagnosis and treatment of brain diseases. This is primarily due to the presence of the blood-brain barrier (BBB), which restricts the entry of many molecules into the brain. In this study, cyclic ADT peptides (ADTC1, ADTC5, and ADTC6) have been shown to modify the BBB to enhance the delivery of marker molecules (e.g., 14C-mannitol, Gd-DTPA) to the brain via the paracellular pathways of the BBB. The hypothesis is that these peptides modulate cadherin interactions in the adherens junctions of the vascular endothelial cells forming the BBB to increase paracellular drug permeation. In vitro studies indicated that ADTC5 had the best profile to inhibit adherens junction resealing in MDCK cell monolayers in a concentration-dependent manner (IC50 = 0.3 mM) with a maximal response at 0.4 mM. Under the current experimental conditions, ADTC5 improved the delivery of 14C-mannitol to the brain about twofold compared to the negative control in the in situ rat brain perfusion model. Furthermore, ADTC5 peptide increased in vivo delivery of Gd-DTPA to the brain of Balb/c mice when administered intravenously (i.v.). In conclusion, ADTC5 has the potential to improve delivery of diagnostic and therapeutic agents to the brain. PMID:25640479

  14. Modulation of intercellular junctions by cyclic-ADT peptides as a method to reversibly increase blood-brain barrier permeability.

    PubMed

    Laksitorini, Marlyn D; Kiptoo, Paul K; On, Ngoc H; Thliveris, James A; Miller, Donald W; Siahaan, Teruna J

    2015-03-01

    It is challenging to deliver molecules to the brain for diagnosis and treatment of brain diseases. This is primarily because of the presence of the blood-brain barrier (BBB), which restricts the entry of many molecules into the brain. In this study, cyclic-ADT peptides (ADTC1, ADTC5, and ADTC6) have been shown to modify the BBB to enhance the delivery of marker molecules [e.g., (14) C-mannitol, gadolinium-diethylenetriaminepentacetate (Gd-DTPA)] to the brain via the paracellular pathways of the BBB. The hypothesis is that these peptides modulate cadherin interactions in the adherens junctions of the vascular endothelial cells forming the BBB to increase paracellular drug permeation. In vitro studies indicated that ADTC5 had the best profile to inhibit adherens junction resealing in Madin-Darby canine kidney cell monolayers in a concentration-dependent manner (IC50 = 0.3 mM) with a maximal response at 0.4 mM. Under the current experimental conditions, ADTC5 improved the delivery of (14) C-mannitol to the brain about twofold compared with the negative control in the in situ rat brain perfusion model. Furthermore, ADTC5 peptide increased in vivo delivery of Gd-DTPA to the brain of Balb/c mice when administered intravenously. In conclusion, ADTC5 has the potential to improve delivery of diagnostic and therapeutic agents to the brain.

  15. The NLRP3 Inflammasome Is a Pathogen Sensor for Invasive Entamoeba histolytica via Activation of α5β1 Integrin at the Macrophage-Amebae Intercellular Junction

    PubMed Central

    Mortimer, Leanne; Moreau, France; Cornick, Steve; Chadee, Kris

    2015-01-01

    Entamoeba histolytica (Eh) is an extracellular protozoan parasite of humans that invades the colon to cause life-threatening intestinal and extra-intestinal amebiasis. Colonized Eh is asymptomatic, however, when trophozoites adhere to host cells there is a considerable inflammatory response that is critical in the pathogenesis of amebiasis. The host and/or parasite factors that trigger the inflammatory response to invading Eh are not well understood. We recently identified that Eh adherence to macrophages induces inflammasome activation and in the present study we sought to determine the molecular events upon contact that coordinates this response. Here we report that Eh contact-dependent activation of α5β1 integrin is critical for activation of the NLRP3 inflammasome. Eh-macrophage contact triggered recruitment of α5β1 integrin and NLRP3 into the intercellular junction, where α5β1 integrin underwent activation by an integrin-binding cysteine protease on the parasite surface, termed EhCP5. As a result of its activation, α5β1 integrin induced ATP release into the extracellular space through opening of pannexin-1 channels that signalled through P2X7 receptors to deliver a critical co-stimulatory signal that activated the NLRP3 inflammasome. Both the cysteine protease activity and integrin-binding domain of EhCP5 were required to trigger α5β1 integrin that led to ATP release and NLRP3 inflammasome activation. These findings reveal engagement of α5β1 integrin across the parasite-host junction is a key regulatory step that initiates robust inflammatory responses to Eh. We propose that α5β1 integrin distinguishes Eh direct contact and functions with NLRP3 as pathogenicity sensor for invasive Eh infection. PMID:25955828

  16. The NLRP3 Inflammasome Is a Pathogen Sensor for Invasive Entamoeba histolytica via Activation of α5β1 Integrin at the Macrophage-Amebae Intercellular Junction.

    PubMed

    Mortimer, Leanne; Moreau, France; Cornick, Steve; Chadee, Kris

    2015-05-01

    Entamoeba histolytica (Eh) is an extracellular protozoan parasite of humans that invades the colon to cause life-threatening intestinal and extra-intestinal amebiasis. Colonized Eh is asymptomatic, however, when trophozoites adhere to host cells there is a considerable inflammatory response that is critical in the pathogenesis of amebiasis. The host and/or parasite factors that trigger the inflammatory response to invading Eh are not well understood. We recently identified that Eh adherence to macrophages induces inflammasome activation and in the present study we sought to determine the molecular events upon contact that coordinates this response. Here we report that Eh contact-dependent activation of α5β1 integrin is critical for activation of the NLRP3 inflammasome. Eh-macrophage contact triggered recruitment of α5β1 integrin and NLRP3 into the intercellular junction, where α5β1 integrin underwent activation by an integrin-binding cysteine protease on the parasite surface, termed EhCP5. As a result of its activation, α5β1 integrin induced ATP release into the extracellular space through opening of pannexin-1 channels that signalled through P2X7 receptors to deliver a critical co-stimulatory signal that activated the NLRP3 inflammasome. Both the cysteine protease activity and integrin-binding domain of EhCP5 were required to trigger α5β1 integrin that led to ATP release and NLRP3 inflammasome activation. These findings reveal engagement of α5β1 integrin across the parasite-host junction is a key regulatory step that initiates robust inflammatory responses to Eh. We propose that α5β1 integrin distinguishes Eh direct contact and functions with NLRP3 as pathogenicity sensor for invasive Eh infection.

  17. Reversal of the TPA-induced inhibition of gap junctional intercellular communication by Chaga mushroom (Inonotus obliquus) extracts: effects on MAP kinases.

    PubMed

    Park, Jung-Ran; Park, Joon-Suk; Jo, Eun-Hye; Hwang, Jae-Woong; Kim, Sun-Jung; Ra, Jeong-Chan; Aruoma, Okezie I; Lee, Yong-Soon; Kang, Kyung-Sun

    2006-01-01

    Chaga mushroom (Inonotus obliquus) has continued to receive attention as a folk medicine with indications for the treatment of cancers and digestive diseases. The anticarcinogenic effect of Chaga mushroom extract was investigated using a model system of gap junctional intercellular communication (GJIC) in WB-F344 normal rat liver epithelial cells. The cells were pre-incubated with Chaga mushroom extracts (5, 10, 20 microg/ml) for 24 h and this was followed by co-treatment with Chaga mushroom extracts and TPA (12-O-tetradecanoylphorbol-13-acetate, 10 ng/ml) for 1 h. The inhibition of GJIC by TPA (12-O-tetradecanoylphorbol-13-acetate), promoter of cancer, was prevented with treatment of Chaga mushroom extracts. Similarly, the increased phosphorylated ERK1/2 and p38 protein kinases were markedly reduced in Chaga mushroom extracts-treated cells. There was no change in the JNK kinase protein level, suggesting that Chaga mushroom extracts could only block the activation of ERK1/2 and p38 MAP kinase. The Chaga mushroom extracts further prevented the inhibition of GJIC through the blocking of Cx43 phosphorylation. Indeed cell-to-cell communication through gap junctional channels is a critical factor in the life and death balance of cells because GJIC has an important function in maintaining tissue homeostasis through the regulation of cell growth, differentiation, apoptosis and adaptive functions of differentiated cells. Thus Chaga mushroom may act as a natural anticancer product by preventing the inhibition of GJIC through the inactivation of ERK1/2 and p38 MAP kinase.

  18. Surface-modified PLGA nanosphere with chitosan improved pulmonary delivery of calcitonin by mucoadhesion and opening of the intercellular tight junctions.

    PubMed

    Yamamoto, Hiromitsu; Kuno, Yoshio; Sugimoto, Shohei; Takeuchi, Hirofumi; Kawashima, Yoshiaki

    2005-02-02

    Surface-modified DL-lactide/glycolide copolymer (PLGA) nanospheres with chitosan (CS) were prepared by the emulsion solvent diffusion method for pulmonary delivery of peptide, i.e., elcatonin. The nanosphere suspension was successfully aerosolized with a nebulizer similar to the drug solution, whereas the microsphere suspensions could not be aerosolized. After pulmonary administration, CS-modified PLGA nanospheres were more slowly eliminated from the lungs than unmodified PLGA nanospheres. CS-modified PLGA nanospheres loaded with elcatonin reduced blood calcium levels to 80% of the initial calcium concentration and prolonged the pharmacological action to 24 h, which was a significantly longer duration of action than that by CS-unmodified nanospheres. These results were attributed to the retention of nanospheres adhered to the bronchial mucus and lung tissue and sustained drug release at the adherence site. In addition, CS and CS on the surface of the nanospheres enhanced the absorption of drug. The rank order of the absorption of the model drugs with CS solution was carboxyfluorescein>FITC-dextran-4 (FD-4; Mw. 4000)>FD-21 (Mw. 21,000)>FD70 (Mw. 70,000), which corresponded to the molecular weights ([Mw.] given in parentheses). The absorption-enhancing effect may have been caused by opening the intercellular tight junctions.

  19. Antiproliferative Action of Conjugated Linoleic Acid on Human MCF-7 Breast Cancer Cells Mediated by Enhancement of Gap Junctional Intercellular Communication through Inactivation of NF-κB

    PubMed Central

    Rakib, Md. Abdur; Lee, Won Sup; Kim, Gon Sup; Han, Jae Hee; Kim, Jeong Ok

    2013-01-01

    The major conjugated linoleic acid (CLA) isomers, c9,t11-CLA and t10,c12-CLA, have anticancer effects; however, the exact mechanisms underlying these effects are unknown. Evidence suggests that reversal of reduced gap junctional intercellular communication (GJIC) in cancer cells inhibits cell growth and induces cell death. Hence, we determined that CLA isomers enhance GJIC in human MCF-7 breast cancer cells and investigated the underlying molecular mechanisms. The CLA isomers significantly enhanced GJIC of MCF-7 cells at 40 μM concentration, whereas CLA inhibited cell growth and induced caspase-dependent apoptosis. CLA increased connexin43 (Cx43) expression both at the transcriptional and translational levels. CLA inhibited nuclear factor-κB (NF-κB) activity and enhanced reactive oxygen species (ROS) generation. No significant difference was observed in the efficacy of c9,t11-CLA and t10,c12-CLA. These results suggest that the anticancer effect of CLA is associated with upregulation of GJIC mediated by enhanced Cx43 expression through inactivation of NF-κB and generation of ROS in MCF-7 cells. PMID:24371460

  20. Inhibition of gap junctional intercellular communication and activation of mitogen-activated protein kinase by tumor-promoting organic peroxides and protection by resveratrol.

    PubMed

    Upham, Brad L; Guzvić, Miodrag; Scott, Jacob; Carbone, Joseph M; Blaha, Ludek; Coe, Chad; Li, Lan Lan; Rummel, Alisa M; Trosko, James E

    2007-01-01

    Dicumyl peroxide (di-CuOOH) and benzoyl peroxide (BzOOH) act as tumor promoters in SENCAR mice, whereas di-tert-butylhydroperoxide does not. Tumor promotion requires the removal of growth suppression by inhibition of gap junctional intercellular communication (GJIC) and the induction of mitogenic intracellular pathways. We showed that di-CuOOH and BzOOH both reversibly inhibited GJIC and transiently activated mitogen-activated protein kinase, specifically, the extracellular receptor kinase at noncytotoxic conditions in WB-F344 rat liver epithelial cells, whereas the non-tumor-promoting di-tert-butylhydroperoxide did not inhibit GJIC or activate extracellular receptor kinase. di-CuOOH but not BzOOH inhibited GJIC through a phosphatidylcholine-specific phospholipase C-dependent mechanism. N-acetylcysteine (NAC) was needed to prevent a cytotoxic, glutathione-depleting effect of BzOOH, whereas di-CuOOH was noncytotoxic and did not alter glutathione levels at all doses and times tested. Pretreatment of WB-F344 cells with resveratrol, a polyphenolic antioxidant present in red wine, prevented at physiological doses the inhibition of GJIC by di-CuOOH but not from BzOOH and was effective in significantly preventing extracellular receptor kinase activation by both peroxides. NAC did not prevent any of the peroxide effects on either GJIC or extracellular receptor kinase, suggesting a specific antioxidant effect of resveratrol.

  1. Neuropeptide Y, substance P, and human bone morphogenetic protein 2 stimulate human osteoblast osteogenic activity by enhancing gap junction intercellular communication

    PubMed Central

    Ma, W.H.; Liu, Y.J.; Wang, W.; Zhang, Y.Z.

    2015-01-01

    Bone homeostasis seems to be controlled by delicate and subtle “cross talk” between the nervous system and “osteo-neuromediators” that control bone remodeling. The purpose of this study was to evaluate the effect of interactions between neuropeptides and human bone morphogenetic protein 2 (hBMP2) on human osteoblasts. We also investigated the effects of neuropeptides and hBMP2 on gap junction intercellular communication (GJIC). Osteoblasts were treated with neuropeptide Y (NPY), substance P (SP), or hBMP2 at three concentrations. At various intervals after treatment, cell viability was measured by the MTT assay. In addition, cellular alkaline phosphatase (ALP) activity and osteocalcin were determined by colorimetric assay and radioimmunoassay, respectively. The effects of NPY, SP and hBMP on GJIC were determined by laser scanning confocal microscopy. The viability of cells treated with neuropeptides and hBMP2 increased significantly in a time-dependent manner, but was inversely associated with the concentration of the treatments. ALP activity and osteocalcin were both reduced in osteoblasts exposed to the combination of neuropeptides and hBMP2. The GJIC of osteoblasts was significantly increased by the neuropeptides and hBMP2. These results suggest that osteoblast activity is increased by neuropeptides and hBMP2 through increased GJIC. Identification of the GJIC-mediated signal transduction capable of modulating the cellular activities of bone cells represents a novel approach to studying the biology of skeletal innervation. PMID:25714881

  2. Red paprika (Capsicum annuum L.) and its main carotenoids, capsanthin and β-carotene, prevent hydrogen peroxide-induced inhibition of gap-junction intercellular communication.

    PubMed

    Kim, Ji-Sun; Lee, Woo-Moon; Rhee, Han Cheol; Kim, Suna

    2016-07-25

    This study was conducted to investigate the protective effect of red paprika extract (RPE) and its main carotenoids, namely, capsanthin (CST) and β-carotene (BCT), on the H2O2-induced inhibition of gap-junction intercellular communication (GJIC) in WB-F344 rat liver epithelial cells (WB cells). We found that pre-treatment with RPE, CST and BCT protected WB cells from H2O2-induced inhibition of GJIC. RPE, CST and BCT not only recovered connexin 43 (Cx43) mRNA expression but also prevented phosphorylation of Cx43 protein by H2O2 treatment. RPE attenuated the phosphorylation of ERK, p38 and JNK, whereas pre-treatment with CST and BCT only attenuated the phosphorylation of ERK and p38 and did not affect JNK in H2O2-treated WB cells. RPE, CST and BCT significantly suppressed the formation of reactive oxygen species (ROS) in H2O2-treated cells compared to untreated WB cells. These results suggest that dietary intake of red paprika might be helpful for lowering the risk of diseases caused by oxidative stress.

  3. Inhibition of gap junctional Intercellular communication in WB-F344 rat liver epithelial cells by triphenyltin chloride through MAPK and PI3-kinase pathways

    PubMed Central

    2010-01-01

    Background Organotin compounds (OTCs) have been widely used as stabilizers in the production of plastic, agricultural pesticides, antifoulant plaints and wood preservation. The toxicity of triphenyltin (TPT) compounds was known for their embryotoxic, neurotoxic, genotoxic and immunotoxic effects in mammals. The carcinogenicity of TPT was not well understood and few studies had discussed the effects of OTCs on gap junctional intercellular communication (GJIC) of cells. Method In the present study, the effects of triphenyltin chloride (TPTC) on GJIC in WB-F344 rat liver epithelial cells were evaluated, using the scrape-loading dye transfer technique. Results TPTC inhibited GJIC after a 30-min exposure in a concentration- and time-dependent manner. Pre-incubation of cells with the protein kinase C (PKC) inhibitor did not modify the response, but the specific MEK 1 inhibitor PD98059 and PI3K inhibitor LY294002 decreased substantially the inhibition of GJIC by TPTC. After WB-F344 cells were exposed to TPTC, phosphorylation of Cx43 increased as seen in Western blot analysis. Conclusions These results show that TPTC inhibits GJIC in WB-F344 rat liver epithelial cells by altering the Cx43 protein expression through both MAPK and PI3-kinase pathways. PMID:20591183

  4. ASSEMBLY AND CHARACTERIZATION OF 8-ARM AND 12-ARM DNA BRANCHED JUNCTIONS

    PubMed Central

    Wang, Xing

    2012-01-01

    Branched DNA molecules can be assembled into objects and networks directed by sticky-ended cohesion. The connectivity of these species is limited by the number of arms flanking the branch point. To date, the only branched junctions constructed contain six or fewer arms. We report the construction of DNA branched junctions that contain either 8 or 12 double helical arms surrounding a branch point. The design of the 8-arm junction expoits the limits of a previous approach to thwart branch migration, but the design of the 12-arm junction uses a new to principle achieve this end. The 8-arm junction is stable with 16 nucleotide pairs per arm, but the 12-arm junction has been stabilized by 24 nucleotide pairs per arm. Ferguson analysis of these junctions in combination with three, four, five, and six-arm junctions indicates a linear increase in friction constant as the number of arms increases; the four-arm junction migrates anomalously at 4°C., suggesting stacking of its domains. All strands in both the 8-arm and 12-arm junctions show similar responses to hydroxyl radical autofootprinting analysis, indicating that they lack any dominant stacking structures. The stability of the 12-arm junction demonstrates that the number of arms in a junction is not limited to the case of having adjacent identical base pairs flanking the junction. The ability to construct eight-arm and twelve-arm junctions increases the number of objects, graphs and networks that can be built from branched DNA components. In principle, the stick structure corresponding to cubic close packing is now a possible target for assembly by DNA nanotechnology. PMID:17564446

  5. Possible anti-tumour-promoting activity of components in Japanese soybean fermented food, Natto: effect on gap junctional intercellular communication.

    PubMed

    Takahashi, C; Kikuchi, N; Katou, N; Miki, T; Yanagida, F; Umeda, M

    1995-03-01

    In order to detect any protective agent against tumor formation, we examined the anti-tumor-promoting effect of a Japanese traditional soybean fermented food, Natto. Dye transfer was employed as an assay method. When fluorescent dye was microinjected into cultured BALB/3T3 cells, the dye was transformed into the neighboring cells through the gap junction. This dye transfer was blocked by the treatment with the tumor promoters 12-O-tetra-decanoylphorbol-13-acetate (TPA), a high concentration of NaCl and lithocholic acid (LCA). This reduction of the dye transfer by TPA treatment was not observed when the cells were pretreated with retinoic acid, an anti-tumor promoter. Thus, the recovery of the dye transfer in TPA-treated BALB/3T3 cells was proven to ge a good indicator for detecting some possible anti-tumor promoters. After extraction and fractionation of Natto, we obtained an active fraction (H1) which showed recovery of the dye transfer in TPA-treated cells. The fraction contained straight-chain saturated hydrocarbons. A comparison of the fraction and the authentic samples by GC analysis suggests that the H1 fraction contained straight-chain saturated hydrocarbons from around C30 to C32. Among these hydrocarbons, hentriacontane (C31) was found at the highest concentrations, and was shown to have the highest activity. Hentriacontane at a very low concentration of 0.65 ng/ml was shown to recover the dye transfer inhibited by the treatment with TPA as well as with NaCl and LCA.

  6. Collective cell guidance by cooperative intercellular forces

    NASA Astrophysics Data System (ADS)

    Tambe, Dhananjay T.; Corey Hardin, C.; Angelini, Thomas E.; Rajendran, Kavitha; Park, Chan Young; Serra-Picamal, Xavier; Zhou, Enhua H.; Zaman, Muhammad H.; Butler, James P.; Weitz, David A.; Fredberg, Jeffrey J.; Trepat, Xavier

    2011-06-01

    Cells comprising a tissue migrate as part of a collective. How collective processes are coordinated over large multi-cellular assemblies has remained unclear, however, because mechanical stresses exerted at cell-cell junctions have not been accessible experimentally. We report here maps of these stresses within and between cells comprising a monolayer. Within the cell sheet there arise unanticipated fluctuations of mechanical stress that are severe, emerge spontaneously, and ripple across the monolayer. Within that stress landscape, local cellular migrations follow local orientations of maximal principal stress. Migrations of both endothelial and epithelial monolayers conform to this behaviour, as do breast cancer cell lines before but not after the epithelial-mesenchymal transition. Collective migration in these diverse systems is seen to be governed by a simple but unifying physiological principle: neighbouring cells join forces to transmit appreciable normal stress across the cell-cell junction, but migrate along orientations of minimal intercellular shear stress.

  7. Ascorbic acid 6-palmitate suppresses gap-junctional intercellular communication through phosphorylation of connexin 43 via activation of the MEK-ERK pathway.

    PubMed

    Lee, Kyung Mi; Kwon, Jung Yeon; Lee, Ki Won; Lee, Hyong Joo

    2009-01-15

    Although the health benefits of dietary antioxidants have been extensively studied, their potential negative effects remain unclear. L-Ascorbic acid 6-palmitate (AAP), a synthetic derivative of ascorbic acid (AA), is widely used as an antioxidant and preservative in foods, vitamins, drugs, and cosmetics. Previously, we found that AA exerted an antitumor effect by protecting inhibition of gap-junctional intercellular communication (GJIC), which is closely associated with tumor progression. In this study, we examined whether AAP, an amphipathic derivative of AA, has chemopreventive effects using a GJIC model. AAP and AA exhibited dose-dependent free radical-scavenging activities and inhibited hydrogen peroxide (H(2)O(2))-induced intracellular reactive oxygen species (ROS) production in normal rat liver epithelial cells. Unexpectedly, however, AAP did not protect against the inhibition of GJIC induced by H(2)O(2); instead, it inhibited GJIC synergistically with H(2)O(2). AAP inhibited GJIC in a dose-dependent and reversible manner. This inhibitory effect was not due to the conjugated lipid structure of AAP, as treatment with palmitic acid alone failed to inhibit GJIC under the same conditions. The inhibition of GJIC by AAP was restored in the presence of mitogen-activated protein kinase/extracellular signal-regulated kinase (ERK) kinase (MEK) inhibitor U0126, but not in the presence of other signal inhibitors and antioxidant (PKC inhibitors, EGFR inhibitor, NADPH oxidase inhibitor, catalase, vitamin E, or AA), indicating the critical involvement of MEK signaling in the GJIC inhibitory activity of AAP. Phosphorylation of ERK and connexin 43 (Cx43) was observed following AAP treatment, and this was reversed by U0126. These results suggest that the AAP-induced inhibition of GJIC is mediated by the phosphorylation of Cx43 via activation of the MEK-ERK pathway. Taken together, our results indicate that AAP has a potent carcinogenic effect, and that the influence of dietary

  8. Inhibition of gap junctional intercellular communication in normal human breast epithelial cells after treatment with pesticides, PCBs, and PBBs, alone or in mixtures.

    PubMed Central

    Kang, K S; Wilson, M R; Hayashi, T; Chang, C C; Trosko, J E

    1996-01-01

    Chemical pollutants in the Great Lakes have found their way through the food chain into humans because of their environmental persistence and lipophilicity. Some epidemiological studies have claimed an association between metabolites of 2,2-bis(p-chlorophenyl)-1,1,1-trichloroethane (DDT), polychlorinated biphenyls (PCBs), and polybrominated biphenyls (PBBs) and breast cancer, but others have reported no such association. We examined various halogenated hydrocarbons for their capacity to inhibit gap junctional intercellular communication (GJIC) in normal human breast epithelial cells (HBEC) when given as single compounds or as mixtures. The scrape-loading/dye transfer and fluorescent redistribution after photobleaching techniques were used to measure GJIC; immunostaining and Western and Northern analyses were performed on connexin 43 (Cx43) gap junction protein and message to determine how halogenated hydrocarbons might affect GJIC. DDT, dieldrin, and toxaphene inhibited GJIC in a dose-responsive manner after 90 min treatments. Dieldrin suppressed GJIC within 30 min with no recovery after 24 hr. Inhibition of GJIC by DDT and toxaphene was partially restored after 12 hr and fully restored after 24 hr. Several PCB and PBB congeners inhibited GJIC in a dose-responsive and time-dependent manner, but GJIC was almost restored to control values 24 hr after exposure. The highest concentrations of the individual chemicals that did not inhibit GJIC was determined, and mixtures containing two of these chemicals were tested for their ability to inhibit GJIC. Significant inhibition of GJIC was observed when cells were treated with a mixture of DDT and 2,4,5-hexachlorobiphenyl (2,4,5-HCB), dieldrin and 2,4,5-HCB, or dieldrin and 2,4,5-hexabromobiphenyl (2,4,5-HBB). These results indicate that halogenated hydrocarbons, alone or in specific combinations, can alter GJIC at the post-translational level. These results are consistent with the hypothesis that DDT, dieldrin, toxaphene, 2

  9. Plakophilin 3 mediates Rap1-dependent desmosome assembly and adherens junction maturation

    PubMed Central

    Todorovic´, Viktor; Koetsier, Jennifer L.; Godsel, Lisa M.; Green, Kathleen J.

    2014-01-01

    The pathways driving desmosome and adherens junction assembly are temporally and spatially coordinated, but how they are functionally coupled is poorly understood. Here we show that the Armadillo protein plakophilin 3 (Pkp3) mediates both desmosome assembly and E-cadherin maturation through Rap1 GTPase, thus functioning in a manner distinct from the closely related plakophilin 2 (Pkp2). Whereas Pkp2 and Pkp3 share the ability to mediate the initial phase of desmoplakin (DP) accumulation at sites of cell–cell contact, they play distinct roles in later steps: Pkp3 is required for assembly of a cytoplasmic population of DP-enriched junction precursors, whereas Pkp2 is required for transfer of the precursors to the membrane. Moreover, Pkp3 forms a complex with Rap1 GTPase, promoting its activation and facilitating desmosome assembly. We show further that Pkp3 deficiency causes disruption of an E-cadherin/Rap1 complex required for adherens junction sealing. These findings reveal Pkp3 as a coordinator of desmosome and adherens junction assembly and maturation through its functional association with Rap1. PMID:25208567

  10. Lack of tumor-promoting effects of flavonoids: studies on rat liver preneoplastic foci and on in vivo and in vitro gap junctional intercellular communication.

    PubMed

    Chaumontet, C; Suschetet, M; Honikman-Leban, E; Krutovskikh, V A; Berges, R; Le Bon, A M; Heberden, C; Shahin, M M; Yamasaki, H; Martel, P

    1996-01-01

    Possible tumor-promoting activity of four flavonoids, quercetin (QC), tangeretin (TG), flavone (FO), and flavanone (FN), was examined in a rat liver short-term carcinogenesis assay as well as with in vivo and in vitro assays of inhibition of gap junctional intercellular communication (GJIC). Rat hepatocarcinogenesis was induced by aflatoxin B1 treatment followed by a selection phase (2-acetylaminofluorene treatment and partial hepatectomy), then treatment with or without test chemicals (in vivo studies of antipromotion were not performed). Using glutathione S-transferase placental form (GST-P)-positive foci, we compared the effects of flavonoids (at 1,000 ppm in the diet) with the effects of phenobarbital (PB) on the occurrence of liver preneoplastic lesions. In addition, we studied the effects of flavonoids on GJIC in the livers derived from these experiments and in two types of cultured cells. No significant difference in the number and area of GST-P-positive foci was found after one or three months of treatment between any flavonoid group and control group. In the positive control group, PB markedly increased the numbers and areas of preneoplastic lesions at three months. Whereas PB also decreased by 60% the average size of lucifer yellow dye spread in slices of liver parenchyma free of preneoplastic lesions among the different flavonoids, only TG decreased the dye transfer in vivo: by 30% at one month and 50% at three months. With the dye transfer assay applied to a rat liver epithelial cell line (REL) and the Chinese hamster V79 metabolic cooperation assay, none of the tested flavonoids (< or = 25 microM) inhibited GJIC. Conversely, protective properties were seen for some of the compounds in antipromotion in vitro studies, because TG and FN enhanced the dye transfer in REL cells and FO, TG, and QC partly prevented the inhibition of metabolic cooperation by 12-O-tetradecanoylphorbol-13-acetate. Thus, taken together, our results suggest that QC, FO, and FN do

  11. Reciprocal influence of connexins and apical junction proteins on their expressions and functions

    PubMed Central

    Derangeon, Mickaël; Spray, David C.; Bourmeyster, Nicolas; Sarrouilhe, Denis; Hervé, Jean-Claude

    2009-01-01

    Membranes of adjacent cells form intercellular junctional complexes to mechanically anchor neighbour cells (anchoring junctions), to seal the paracellular space and to prevent diffusion of integral proteins within the plasma membrane (tight junctions) and to allow cell-to-cell diffusion of small ions and molecules (gap junctions). These different types of specialised plasma membrane microdomains, sharing common adaptor molecules, particularly zonula occludens proteins, frequently present intermingled relationships where the different proteins co-assemble into macromolecular complexes and their expressions are co-ordinately regulated. Proteins forming gap junction channels (connexins, particularly) and proteins fulfilling cell attachment or forming tight junction strands mutually influence expression and functions of one another. PMID:19046940

  12. Biochemical analysis of connexin43 intracellular transport, phosphorylation, and assembly into gap junctional plaques

    PubMed Central

    1991-01-01

    We previously demonstrated that the gap junction protein connexin43 is translated as a 42-kD protein (connexin43-NP) that is efficiently phosphorylated to a 46,000-Mr species (connexin43-P2) in gap junctional communication-competent, but not in communication-deficient, cells. In this study, we used a combination of metabolic radiolabeling and immunoprecipitation to investigate the assembly of connexin43 into gap junctions and the relationship of this event to phosphorylation of connexin43. Examination of the detergent solubility of connexin43 in communication-competent NRK cells revealed that processing of connexin43 to the P2 form was accompanied by acquisition of resistance to solubilization in 1% Triton X-100. Immunohistochemical localization of connexin43 in Triton-extracted NRK cells demonstrated that connexin43-P2 (Triton-insoluble) was concentrated in gap junctional plaques, whereas connexin43-NP (Triton-soluble) was predominantly intracellular. Using either a 20 degrees C intracellular transport block or cell-surface protein biotinylation, we determined that connexin43 was transported to the plasma membrane in the Triton-soluble connexin43-NP form. Cell-surface biotinylated connexin43-NP was processed to Triton-insoluble connexin43-P2 at 37 degrees C. Connexin43- NP was also transported to the plasma membrane in communication defective, gap junction-deficient S180 and L929 cells but was not processed to Triton-insoluble connexin43-P2. Taken together, these results demonstrate that gap junction assembly is regulated after arrival of connexin43 at the plasma membrane and is temporally associated with acquisition of insolubility in Triton X-100 and phosphorylation to the connexin43-P2 form. PMID:1659577

  13. Charge transport in vertically aligned, self-assembled peptide nanotube junctions

    NASA Astrophysics Data System (ADS)

    Mizrahi, Mordechay; Zakrassov, Alexander; Lerner-Yardeni, Jenny; Ashkenasy, Nurit

    2012-01-01

    The self-assembly propensity of peptides has been extensively utilized in recent years for the formation of supramolecular nanostructures. In particular, the self-assembly of peptides into fibrils and nanotubes makes them promising building blocks for electronic and electro-optic applications. However, the mechanisms of charge transfer in these wire-like structures, especially in ambient conditions, are not yet fully understood. We describe here a layer-by-layer deposition methodology of short self-assembled cyclic peptide nanotubes, which results in vertically oriented nanotubes on gold substrates. Using this novel deposition methodology, we have fabricated molecular junctions with a conductive atomic force microscopy tip as a second electrode. Studies of the junctions' current-voltage characteristics as a function of the nanotube length revealed an efficient charge transfer in these supramolecular structures, with a low current attenuation constant of 0.1 Å-1, which indicate that electron transfer is dominated by hopping. Moreover, the threshold voltage to field-emission dominated transport was found to increase with peptide length in a manner that depends on the nature of the contact with the electrodes. The flexibility in the design of the peptide monomers and the ability to control their sequential order over the nanotube by means of the layer-by-layer assembly process, which is demonstrated in this work, can be used to engineer the electronic properties of self-assembled peptide nanotubes toward device applications.

  14. Charge transport in vertically aligned, self-assembled peptide nanotube junctions.

    PubMed

    Mizrahi, Mordechay; Zakrassov, Alexander; Lerner-Yardeni, Jenny; Ashkenasy, Nurit

    2012-01-21

    The self-assembly propensity of peptides has been extensively utilized in recent years for the formation of supramolecular nanostructures. In particular, the self-assembly of peptides into fibrils and nanotubes makes them promising building blocks for electronic and electro-optic applications. However, the mechanisms of charge transfer in these wire-like structures, especially in ambient conditions, are not yet fully understood. We describe here a layer-by-layer deposition methodology of short self-assembled cyclic peptide nanotubes, which results in vertically oriented nanotubes on gold substrates. Using this novel deposition methodology, we have fabricated molecular junctions with a conductive atomic force microscopy tip as a second electrode. Studies of the junctions' current-voltage characteristics as a function of the nanotube length revealed an efficient charge transfer in these supramolecular structures, with a low current attenuation constant of 0.1 Å(-1), which indicate that electron transfer is dominated by hopping. Moreover, the threshold voltage to field-emission dominated transport was found to increase with peptide length in a manner that depends on the nature of the contact with the electrodes. The flexibility in the design of the peptide monomers and the ability to control their sequential order over the nanotube by means of the layer-by-layer assembly process, which is demonstrated in this work, can be used to engineer the electronic properties of self-assembled peptide nanotubes toward device applications.

  15. HIV-1 gp120 Glycoprotein Interacting with Dendritic Cell-specific Intercellular Adhesion Molecule 3-grabbing Non-integrin (DC-SIGN) Down-Regulates Tight Junction Proteins to Disrupt the Blood Retinal Barrier and Increase Its Permeability.

    PubMed

    Qian, Yi-Wen; Li, Chuan; Jiang, Ai-Ping; Ge, Shengfang; Gu, Ping; Fan, Xianqun; Li, Tai-Sheng; Jin, Xia; Wang, Jian-Hua; Wang, Zhi-Liang

    2016-10-28

    Approximately 70% of HIV-1 infected patients acquire ocular opportunistic infections and manifest eye disorders during the course of their illness. The mechanisms by which pathogens invade the ocular site, however, are unclear. Under normal circumstances, vascular endothelium and retinal pigment epithelium (RPE), which possess a well developed tight junction complex, form the blood-retinal barrier (BRB) to prevent pathogen invasion. We hypothesize that disruption of the BRB allows pathogen entry into ocular sites. The hypothesis was tested using in vitro models. We discovered that human RPE cells could bind to either HIV-1 gp120 glycoproteins or HIV-1 viral particles. Furthermore, the binding was mediated by dendritic cell-specific intercellular adhesion molecule 3-grabbing non-integrin (DC-SIGN) expressed on RPE cells. Upon gp120 binding to DC-SIGN, cellular NF-κB signaling was triggered, leading to the induction of matrix metalloproteinases, which subsequently degraded tight junction proteins and disrupted the BRB integrity. DC-SIGN knockdown or prior blocking with a specific antibody abolished gp120-induced matrix metalloproteinase expression and reduced the degradation of tight junction proteins. This study elucidates a novel mechanism by which HIV, type 1 invades ocular tissues and provides additional insights into the translocation or invasion process of ocular complication-associated pathogens.

  16. Cooperation of bcl-2 and myc in the neoplastic transformation of normal rat liver epithelial cells is related to the down-regulation of gap junction-mediated intercellular communication.

    PubMed

    DeoCampo, N D; Wilson, M R; Trosko, J E

    2000-08-01

    The objectives of this study were to isolate several rat liver epithelial cell clones containing the human bcl-2 and myc/bcl-2 genes in order to study their potential cooperative effect on neoplastic transformation and gap junction-mediated intercellular communication (GJIC) and to test the hypothesis that the loss of GJIC leads to tumorigenesis. Using anchorage-independent growth as a surrogate marker for neoplastic transformation, we transfected both normal rat liver epithelial cells, WB-F344, and a WB-F344 cell line overexpressing v-myc with human bcl-2 cDNA. Those cell lines that only expressed v-myc or human bcl-2 were unable to form colonies in soft agar. However, those cell lines that overexpressed both v-myc and human bcl-2 showed varying ability to form colonies in soft agar, which did not correlate with their human bcl-2 expression level. In order to test if there was a correlation between cell line growth in soft agar and the ability to communicate through gap junctions, we performed scrape load dye transfer and fluorescence recovery after photobleaching assays. Our results show that v-myc and human bcl-2 can cooperate in the transformation of normal cells, but the degree to which the cells are transformed is dependent on the cells' ability to communicate through gap junctions.

  17. Gap junctions.

    PubMed

    Goodenough, Daniel A; Paul, David L

    2009-07-01

    Gap junctions are aggregates of intercellular channels that permit direct cell-cell transfer of ions and small molecules. Initially described as low-resistance ion pathways joining excitable cells (nerve and muscle), gap junctions are found joining virtually all cells in solid tissues. Their long evolutionary history has permitted adaptation of gap-junctional intercellular communication to a variety of functions, with multiple regulatory mechanisms. Gap-junctional channels are composed of hexamers of medium-sized families of integral proteins: connexins in chordates and innexins in precordates. The functions of gap junctions have been explored by studying mutations in flies, worms, and humans, and targeted gene disruption in mice. These studies have revealed a wide diversity of function in tissue and organ biology.

  18. Gap Junctions

    PubMed Central

    Goodenough, Daniel A.; Paul, David L.

    2009-01-01

    Gap junctions are aggregates of intercellular channels that permit direct cell–cell transfer of ions and small molecules. Initially described as low-resistance ion pathways joining excitable cells (nerve and muscle), gap junctions are found joining virtually all cells in solid tissues. Their long evolutionary history has permitted adaptation of gap-junctional intercellular communication to a variety of functions, with multiple regulatory mechanisms. Gap-junctional channels are composed of hexamers of medium-sized families of integral proteins: connexins in chordates and innexins in precordates. The functions of gap junctions have been explored by studying mutations in flies, worms, and humans, and targeted gene disruption in mice. These studies have revealed a wide diversity of function in tissue and organ biology. PMID:20066080

  19. Phosphorylation on Ser-279 and Ser-282 of connexin43 regulates endocytosis and gap junction assembly in pancreatic cancer cells

    PubMed Central

    Johnson, Kristen E.; Mitra, Shalini; Katoch, Parul; Kelsey, Linda S.; Johnson, Keith R.; Mehta, Parmender P.

    2013-01-01

    The molecular mechanisms regulating the assembly of connexins (Cxs) into gap junctions are poorly understood. Using human pancreatic tumor cell lines BxPC3 and Capan-1, which express Cx26 and Cx43, we show that, upon arrival at the cell surface, the assembly of Cx43 is impaired. Connexin43 fails to assemble, because it is internalized by clathrin-mediated endocytosis. Assembly is restored upon expressing a sorting-motif mutant of Cx43, which does not interact with the AP2 complex, and by expressing mutants that cannot be phosphorylated on Ser-279 and Ser-282. The mutants restore assembly by preventing clathrin-mediated endocytosis of Cx43. Our results also document that the sorting-motif mutant is assembled into gap junctions in cells in which the expression of endogenous Cx43 has been knocked down. Remarkably, Cx43 mutants that cannot be phosphorylated on Ser-279 or Ser-282 are assembled into gap junctions only when connexons are composed of Cx43 forms that can be phosphorylated on these serines and forms in which phosphorylation on these serines is abolished. Based on the subcellular fate of Cx43 in single and contacting cells, our results document that the endocytic itinerary of Cx43 is altered upon cell–cell contact, which causes Cx43 to traffic by EEA1-negative endosomes en route to lysosomes. Our results further show that gap-junctional plaques formed of a sorting motif–deficient mutant of Cx43, which is unable to be internalized by the clathrin-mediated pathway, are predominantly endocytosed in the form of annular junctions. Thus the differential phosphorylation of Cx43 on Ser-279 and Ser-282 is fine-tuned to control Cx43’s endocytosis and assembly into gap junctions. PMID:23363606

  20. ADAM10 Is Involved in Cell Junction Assembly in Early Porcine Embryo Development

    PubMed Central

    Kwon, Jeongwoo; Jeong, Sung-min; Choi, Inchul; Kim, Nam-Hyung

    2016-01-01

    ADAM10 (A Disintegrin and Metalloprotease domain-containing protein 10) is a cell surface protein with a unique structure possessing both potential adhesion and protease domains. However, the role of ADAM10 in preimplantation stage embryos is not clear. In this study, we examined the expression patterns and functional roles of ADAM10 in porcine parthenotes during preimplantation development. The transcription level of ADAM10 dramatically increased from the morula stage onward. Immunostaining revealed that ADAM10 was present in both the nucleus and cytoplasm in early cleavage stage embryos, and localized to the apical region of the outer cells in morula and blastocyst embryos. Knockdown (KD) of ADAM10 using double strand RNA did not alter preimplantation embryo development until morula stage, but resulted in significantly reduced development to blastocyst stage. Moreover, the KD blastocyst showed a decrease in gene expression of adherens and tight junction (AJ/TJ), and an increase in trophectoderm TJ permeability by disrupting TJ assembly. Treatment with an ADAM10 specific chemical inhibitor, GI254023X, at the morula stage also inhibited blastocyst development and led to disruption of TJ assembly. An in situ proximity ligation assay demonstrated direct interaction of ADAM10 with coxsackie virus and adenovirus receptor (CXADR), supporting the involvement of ADAM10 in TJ assembly. In conclusion, our findings strongly suggest that ADADM10 is important for blastocyst formation rather than compaction, particularly for TJ assembly and stabilization in preimplantation porcine parthenogenetic development. PMID:27043020

  1. ADAM10 Is Involved in Cell Junction Assembly in Early Porcine Embryo Development.

    PubMed

    Kwon, Jeongwoo; Jeong, Sung-min; Choi, Inchul; Kim, Nam-Hyung

    2016-01-01

    ADAM10 (A Disintegrin and Metalloprotease domain-containing protein 10) is a cell surface protein with a unique structure possessing both potential adhesion and protease domains. However, the role of ADAM10 in preimplantation stage embryos is not clear. In this study, we examined the expression patterns and functional roles of ADAM10 in porcine parthenotes during preimplantation development. The transcription level of ADAM10 dramatically increased from the morula stage onward. Immunostaining revealed that ADAM10 was present in both the nucleus and cytoplasm in early cleavage stage embryos, and localized to the apical region of the outer cells in morula and blastocyst embryos. Knockdown (KD) of ADAM10 using double strand RNA did not alter preimplantation embryo development until morula stage, but resulted in significantly reduced development to blastocyst stage. Moreover, the KD blastocyst showed a decrease in gene expression of adherens and tight junction (AJ/TJ), and an increase in trophectoderm TJ permeability by disrupting TJ assembly. Treatment with an ADAM10 specific chemical inhibitor, GI254023X, at the morula stage also inhibited blastocyst development and led to disruption of TJ assembly. An in situ proximity ligation assay demonstrated direct interaction of ADAM10 with coxsackie virus and adenovirus receptor (CXADR), supporting the involvement of ADAM10 in TJ assembly. In conclusion, our findings strongly suggest that ADADM10 is important for blastocyst formation rather than compaction, particularly for TJ assembly and stabilization in preimplantation porcine parthenogenetic development.

  2. Evidence for intercellular communication in mosquito renal tubules: a putative role of gap junctions in coordinating and regulating the rapid diuretic effects of neuropeptides

    PubMed Central

    Piermarini, Peter M.; Calkins, Travis L.

    2014-01-01

    Adult female mosquitoes require a blood meal from a vertebrate host to successfully reproduce. During a single blood feeding, a female may ingest more than the equivalent of her own body mass, resulting in an acute stress to osmotic and ionic homeostasis. In response to this stress, the renal (Malpighian) tubules mediate a rapid diuresis that commences as soon as blood is ingested. The diuresis is regulated by neuropeptides (e.g., kinins, calcitonin-like peptide) that act on receptors in the Malpighian tubule epithelium. Interestingly, the expression of these receptors is discontinuous throughout the epithelium, which raises the question as to how Malpighian tubules mount such a rapid and synchronized response to neuropeptide stimulation. Here we propose a hypothesis that gap junctions functionally couple the epithelial cells of Malpighian tubules, resulting in a coordinated physiological response to the binding of neuropeptides. We review recent, relevant literature on the electrophysiology, physiology, and molecular biology of mosquito Malpighian tubules that indicate the presence of gap junctions in the epithelium. We also provide new physiological and immunochemical data that are consistent with the proposed hypothesis. PMID:24316302

  3. Junctional actin assembly is mediated by Formin-like 2 downstream of Rac1

    PubMed Central

    Grikscheit, Katharina; Frank, Tanja; Wang, Ying

    2015-01-01

    Epithelial integrity is vitally important, and its deregulation causes early stage cancer. De novo formation of an adherens junction (AJ) between single epithelial cells requires coordinated, spatial actin dynamics, but the mechanisms steering nascent actin polymerization for cell–cell adhesion initiation are not well understood. Here we investigated real-time actin assembly during daughter cell–cell adhesion formation in human breast epithelial cells in 3D environments. We identify formin-like 2 (FMNL2) as being specifically required for actin assembly and turnover at newly formed cell–cell contacts as well as for human epithelial lumen formation. FMNL2 associates with components of the AJ complex involving Rac1 activity and the FMNL2 C terminus. Optogenetic control of Rac1 in living cells rapidly drove FMNL2 to epithelial cell–cell contact zones. Furthermore, Rac1-induced actin assembly and subsequent AJ formation critically depends on FMNL2. These data uncover FMNL2 as a driver for human epithelial AJ formation downstream of Rac1. PMID:25963818

  4. Gap Junctions

    PubMed Central

    Nielsen, Morten Schak; Axelsen, Lene Nygaard; Sorgen, Paul L.; Verma, Vandana; Delmar, Mario; Holstein-Rathlou, Niels-Henrik

    2013-01-01

    Gap junctions are essential to the function of multicellular animals, which require a high degree of coordination between cells. In vertebrates, gap junctions comprise connexins and currently 21 connexins are known in humans. The functions of gap junctions are highly diverse and include exchange of metabolites and electrical signals between cells, as well as functions, which are apparently unrelated to intercellular communication. Given the diversity of gap junction physiology, regulation of gap junction activity is complex. The structure of the various connexins is known to some extent; and structural rearrangements and intramolecular interactions are important for regulation of channel function. Intercellular coupling is further regulated by the number and activity of channels present in gap junctional plaques. The number of connexins in cell-cell channels is regulated by controlling transcription, translation, trafficking, and degradation; and all of these processes are under strict control. Once in the membrane, channel activity is determined by the conductive properties of the connexin involved, which can be regulated by voltage and chemical gating, as well as a large number of posttranslational modifications. The aim of the present article is to review our current knowledge on the structure, regulation, function, and pharmacology of gap junctions. This will be supported by examples of how different connexins and their regulation act in concert to achieve appropriate physiological control, and how disturbances of connexin function can lead to disease. © 2012 American Physiological Society. Compr Physiol 2:1981-2035, 2012. PMID:23723031

  5. Benzo[a]pyrene-7,8-diol-9,10-epoxide inhibits gap junction intercellular communication via phosphorylation of tumor progression locus 2 in WB-F344 rat liver epithelial cells.

    PubMed

    Lee, Bo Kyung; Chung, Min-Yu; Lee, Ki Won

    2015-05-01

    Benzo[a]pyrene-7,8-diol-9,10-epoxide (B[a]PDE), a major metabolite of benzo[a]pyrene, has been reported to function as a human carcinogen. However, the molecular mechanism of how B[a]PDE regulates signaling pathways during tumor promotion remains unclear. In this study, we investigated the effects of B[a]PDE on the regulation of gap junction intercellular communication (GJIC), one of the major carcinogenic processes, and its main regulatory signaling pathways using WB-F344 rat liver epithelial (WB-F344 RLE) cells. Treatment of benzo[a]pyrene or B[a]PDE resulted in GJIC inhibition, and B[a]PDE was more active at lower concentrations than benzo[a]pyrene in the suppression of GJIC. This suggests that B[a]PDE is a stronger GJIC inhibitor. B[a]PDE at 1 µM reversibly inhibited GJIC in WB-F344 RLE cells, which was attributable to hyperphosphorylation of connexin43 (Cx43) via phosphorylation of mitogen-activated protein kinase/extracellular signal-regulated kinase (MEK) and extracellular signal-regulated kinase (ERK). We found that B[a]PDE induced phosphorylation of tumor progression locus 2 (Tpl2), a direct upstream regulator of MEK. Tpl2 inhibitor recovered B[a]PDE-induced GJIC inhibition and attenuated B[a]PDE-induced MEK/ERK phosphorylation in WB-F344 RLE cells. Collectively, our results suggest that B[a]PDE suppresses GJIC by activating Tpl2 and subsequently the MEK/ERK pathway and Cx43 phosphorylation in WB-F344 RLE cells. These results outline the potential importance of Tpl2 as a novel therapeutic target for B[a]PDE-induced GJIC inhibition during cancer promotion.

  6. Creating Sub-50 Nm Nanofluidic Junctions in PDMS Microfluidic Chip via Self-Assembly Process of Colloidal Particles.

    PubMed

    Wei, Xi; Syed, Abeer; Mao, Pan; Han, Jongyoon; Song, Yong-Ak

    2016-03-13

    Polydimethylsiloxane (PDMS) is the prevailing building material to make microfluidic devices due to its ease of molding and bonding as well as its transparency. Due to the softness of the PDMS material, however, it is challenging to use PDMS for building nanochannels. The channels tend to collapse easily during plasma bonding. In this paper, we present an evaporation-driven self-assembly method of silica colloidal nanoparticles to create nanofluidic junctions with sub-50 nm pores between two microchannels. The pore size as well as the surface charge of the nanofluidic junction is tunable simply by changing the colloidal silica bead size and surface functionalization outside of the assembled microfluidic device in a vial before the self-assembly process. Using the self-assembly of nanoparticles with a bead size of 300 nm, 500 nm, and 900 nm, it was possible to fabricate a porous membrane with a pore size of ~45 nm, ~75 nm and ~135 nm, respectively. Under electrical potential, this nanoporous membrane initiated ion concentration polarization (ICP) acting as a cation-selective membrane to concentrate DNA by ~1,700 times within 15 min. This non-lithographic nanofabrication process opens up a new opportunity to build a tunable nanofluidic junction for the study of nanoscale transport processes of ions and molecules inside a PDMS microfluidic chip.

  7. Role of redox centre in charge transport investigated by novel self-assembled conjugated polymer molecular junctions

    PubMed Central

    Wang, Zongrui; Dong, Huanli; Li, Tao; Hviid, Rune; Zou, Ye; Wei, Zhongming; Fu, Xiaolong; Wang, Erjing; Zhen, Yonggang; Nørgaard, Kasper; Laursen, Bo W.; Hu, Wenping

    2015-01-01

    Molecular electronics describes a field that seeks to implement electronic components made of molecular building blocks. To date, few studies have used conjugated polymers in molecular junctions despite the fact that they potentially transport charge more efficiently than the extensively investigated small-molecular systems. Here we report a novel type of molecular tunnelling junction exploring the use of conjugated polymers, which are self-assembled into ultrathin films in a distinguishable ‘planar' manner from the traditional vertically oriented small-molecule monolayers. Electrical measurements on the junctions reveal molecular-specific characteristics of the polymeric molecules in comparison with less conjugated small molecules. More significantly, we decorate redox-active functionality into polymeric backbones, demonstrating a key role of redox centre in the modulation of charge transport behaviour via energy level engineering and external stimuli, and implying the potential of employing tailor-made polymeric components as alternatives to small molecules for future molecular-scale electronics. PMID:26085081

  8. Bottom head to shell junction assembly for a boiling water nuclear reactor

    DOEpatents

    Fife, A.B.; Ballas, G.J.

    1998-02-24

    A bottom head to shell junction assembly which, in one embodiment, includes an annular forging having an integrally formed pump deck and shroud support is described. In the one embodiment, the annular forging also includes a top, cylindrical shaped end configured to be welded to one end of the pressure vessel cylindrical shell and a bottom, conical shaped end configured to be welded to the disk shaped bottom head. Reactor internal pump nozzles also are integrally formed in the annular forging. The nozzles do not include any internal or external projections. Stubs are formed in each nozzle opening to facilitate welding a pump housing to the forging. Also, an upper portion of each nozzle opening is configured to receive a portion of a diffuser coupled to a pump shaft which extends through the nozzle opening. Diffuser openings are formed in the integral pump deck to provide additional support for the pump impellers. The diffuser opening is sized so that a pump impeller can extend at least partially therethrough. The pump impeller is connected to the pump shaft which extends through the nozzle opening. 5 figs.

  9. Bottom head to shell junction assembly for a boiling water nuclear reactor

    DOEpatents

    Fife, Alex Blair; Ballas, Gary J.

    1998-01-01

    A bottom head to shell junction assembly which, in one embodiment, includes an annular forging having an integrally formed pump deck and shroud support is described. In the one embodiment, the annular forging also includes a top, cylindrical shaped end configured to be welded to one end of the pressure vessel cylindrical shell and a bottom, conical shaped end configured to be welded to the disk shaped bottom head. Reactor internal pump nozzles also are integrally formed in the annular forging. The nozzles do not include any internal or external projections. Stubs are formed in each nozzle opening to facilitate welding a pump housing to the forging. Also, an upper portion of each nozzle opening is configured to receive a portion of a diffuser coupled to a pump shaft which extends through the nozzle opening. Diffuser openings are formed in the integral pump deck to provide additional support for the pump impellers. The diffuser opening is sized so that a pump impeller can extend at least partially therethrough. The pump impeller is connected to the pump shaft which extends through the nozzle opening.

  10. Butyrate enhances the intestinal barrier by facilitating tight junction assembly via activation of AMP-activated protein kinase in Caco-2 cell monolayers.

    PubMed

    Peng, Luying; Li, Zhong-Rong; Green, Robert S; Holzman, Ian R; Lin, Jing

    2009-09-01

    Butyrate, one of the SCFA, promotes the development of the intestinal barrier. However, the molecular mechanisms underlying the butyrate regulation of the intestinal barrier are unknown. To test the hypothesis that the effect of butyrate on the intestinal barrier is mediated by the regulation of the assembly of tight junctions involving the activation of the AMP-activated protein kinase (AMPK), we determined the effect of butyrate on the intestinal barrier by measuring the transepithelial electrical resistance (TER) and inulin permeability in a Caco-2 cell monolayer model. We further used a calcium switch assay to study the assembly of epithelial tight junctions and determined the effect of butyrate on the assembly of epithelial tight junctions and AMPK activity. We demonstrated that the butyrate treatment increased AMPK activity and accelerated the assembly of tight junctions as shown by the reorganization of tight junction proteins, as well as the development of TER. AMPK activity was also upregulated by butyrate during calcium switch-induced tight junction assembly. Compound C, a specific AMPK inhibitor, inhibited the butyrate-induced activation of AMPK. The facilitating effect of butyrate on the increases in TER in standard culture media, as well as after calcium switch, was abolished by compound C. We conclude that butyrate enhances the intestinal barrier by regulating the assembly of tight junctions. This dynamic process is mediated by the activation of AMPK. These results suggest an intriguing link between SCFA and the intracellular energy sensor for the development of the intestinal barrier.

  11. Claudin-16 and claudin-19 interaction is required for their assembly into tight junctions and for renal reabsorption of magnesium.

    PubMed

    Hou, Jianghui; Renigunta, Aparna; Gomes, Antonio S; Hou, Mingli; Paul, David L; Waldegger, Siegfried; Goodenough, Daniel A

    2009-09-08

    Claudins are tight junction integral membrane proteins that are key regulators of the paracellular pathway. Defects in claudin-16 (CLDN16) and CLDN19 function result in the inherited human renal disorder familial hypomagnesemia with hypercalciuria and nephrocalcinosis (FHHNC). Previous studies showed that siRNA knockdown of CLDN16 in mice results in a mouse model for FHHNC. Here, we show that CLDN19-siRNA mice also developed the FHHNC symptoms of chronic renal wasting of magnesium and calcium together with defective renal salt handling. siRNA knockdown of CLDN19 caused a loss of CLDN16 from tight junctions in the thick ascending limb (TAL) without a decrease in CLDN16 expression level, whereas siRNA knockdown of CLDN16 produced a similar effect on CLDN19. In both mouse lines, CLDN10, CLDN18, occludin, and ZO-1, normal constituents of TAL tight junctions, remained correctly localized. CLDN16- and CLDN19-depleted tight junctions had normal barrier function but defective ion selectivity. These data, together with yeast two-hybrid binding studies, indicate that a heteromeric CLDN16 and CLDN19 interaction was required for assembling them into the tight junction structure and generating cation-selective paracellular channels.

  12. A relay mechanism between EB1 and APC facilitate STIM1 puncta assembly at endoplasmic reticulum-plasma membrane junctions.

    PubMed

    Asanov, Alexander; Sherry, Ryan; Sampieri, Alicia; Vaca, Luis

    2013-09-01

    The assembly of STIM1 protein puncta near endoplasmic reticulum-plasma membrane (ER-PM) junctions is required for optimal activation of store-operated channels (SOC). The mechanisms controlling the translocation of STIM1 puncta to ER-PM junctions remain largely unknown. In the present study, we have explored the role of the microtubule binding protein adenomatous polyposis coli (APC), on STIM1 puncta and store-operated calcium entry (SOCE). APC-depleted cells showed reduced STIM1 puncta near ER-PM junctions, instead puncta is found at the ER surrounding the cell nucleus. Reduced STIM1 puncta near ER-PM junctions in APC-depleted cells correlates with a strong inhibition of SOCE and diminished Orai whole-cell currents. Immunoprecipitation and confocal microscopy co-localization studies indicate that, upon depletion of the ER, STIM1 dissociates from EB1 and associates to APC. Deletion analysis identified an APC-binding domain in the carboxyl terminus of STIM1 (STIM1 650-685). These results together position APC as an important element in facilitating the translocation of STIM1 puncta near ER-PM junctions, which in turn is required for efficient SOCE and Orai activation upon depletion of the ER.

  13. A complex of ZO-1 and the BAR-domain protein TOCA-1 regulates actin assembly at the tight junction

    PubMed Central

    Van Itallie, Christina M.; Tietgens, Amber Jean; Krystofiak, Evan; Kachar, Bechara; Anderson, James M.

    2015-01-01

    Assembly and sealing of the tight junction barrier are critically dependent on the perijunctional actin cytoskeleton, yet little is known about physical and functional links between barrier-forming proteins and actin. Here we identify a novel functional complex of the junction scaffolding protein ZO-1 and the F-BAR–domain protein TOCA-1. Using MDCK epithelial cells, we show that an alternative splice of TOCA-1 adds a PDZ-binding motif, which binds ZO-1, targeting TOCA-1 to barrier contacts. This isoform of TOCA-1 recruits the actin nucleation–promoting factor N-WASP to tight junctions. CRISPR-Cas9–mediated knockout of TOCA-1 results in increased paracellular flux and delayed recovery in a calcium switch assay. Knockout of TOCA-1 does not alter FRAP kinetics of GFP ZO-1 or occludin, but longer term (12 h) time-lapse microscopy reveals strikingly decreased tight junction membrane contact dynamics in knockout cells compared with controls. Reexpression of TOCA-1 with, but not without, the PDZ-binding motif rescues both altered flux and membrane contact dynamics. Ultrastructural analysis shows actin accumulation at the adherens junction in TOCA-1–knockout cells but unaltered freeze-fracture fibril morphology. Identification of the ZO-1/TOCA-1 complex provides novel insights into the underappreciated dependence of the barrier on the dynamic nature of cell-to-cell contacts and perijunctional actin. PMID:26063734

  14. The Interaction of JRAB/MICAL-L2 with Rab8 and Rab13 Coordinates the Assembly of Tight Junctions and Adherens Junctions

    PubMed Central

    Yamamura, Rie; Nishimura, Noriyuki; Nakatsuji, Hiroyoshi; Arase, Seiji

    2008-01-01

    The assembly of tight junctions (TJs) and adherens junctions (AJs) is regulated by the transport of integral TJ and AJ proteins to and/or from the plasma membrane (PM) and it is tightly coordinated in epithelial cells. We previously reported that Rab13 and a junctional Rab13-binding protein (JRAB)/molecule interacting with CasL-like 2 (MICAL-L2) mediated the endocytic recycling of an integral TJ protein occludin and the formation of functional TJs. Here, we investigated the role of Rab13 and JRAB/MICAL-L2 in the transport of other integral TJ and AJ proteins claudin-1 and E-cadherin to the PM by using a Ca2+-switch model. Although knockdown of Rab13 specifically suppressed claudin-1 and occludin but not E-cadherin transport, knockdown of JRAB/MICAL-L2 and expression of its Rab13-binding domain (JRAB/MICAL-L2-C) inhibited claudin-1, occludin, and E-cadherin transport. We then identified Rab8 as another JRAB/MICAL-L2-C-binding protein. Knockdown of Rab8 inhibited the Rab13-independent transport of E-cadherin to the PM. Rab8 and Rab13 competed with each other for the binding to JRAB/MICAL-L2 and functionally associated with JRAB/MICAL-L2 at the perinuclear recycling/storage compartments and PM, respectively. These results suggest that the interaction of JRAB/MICAL-L2 with Rab8 and Rab13 coordinates the assembly of AJs and TJs. PMID:18094055

  15. Effect of trunnion roughness and length on the modular taper junction strength under typical intraoperative assembly forces.

    PubMed

    Jauch-Matt, S Y; Miles, A W; Gill, H S

    2017-01-01

    Modular hip implants are at risk of fretting-induced postoperative complications most likely initiated by micromotion between adjacent implant components. A stable fixation between ball head and stem-neck taper is critical to avoid excessive interface motions. Therefore, the aim of this study was to identify the effect of trunnion roughness and length on the modular taper strength under typical intraoperative assembly forces. Custom-made Titanium trunnions (standard/mini taper, smooth/grooved surface finish) were assembled with modular Cobalt-chromium heads by impaction with peak forces ranging from 2kN to 6kN. After each assembly process these were disassembled with a materials testing machine to detect the pull-off force as a measure for the taper strength. As expected, the pull-off forces increased with rising peak assembly force (p < 0.001). For low and moderate assembly forces, smooth standard tapers offered higher pull-off forces compared to grooved tapers (p < 0.038). In the case of an assembly force of 2kN, mini tapers showed a higher taper strength than standard ones (p=0.037). The results of this study showed that smooth tapers provided a higher strength for taper junctions. This higher taper strength may reduce the risk of fretting-related complications especially in the most common range of intraoperative assembly forces.

  16. The Extracellular Architecture of Adherens Junctions Revealed by Crystal Structures of Type I Cadherins

    SciTech Connect

    O Harrison; X Jin; S Hong; F Bahna; G Ahlsen; J Brasch; Y Wu; J Vendome; K Felsovalyi; et al.

    2011-12-31

    Adherens junctions, which play a central role in intercellular adhesion, comprise clusters of type I classical cadherins that bind via extracellular domains extended from opposing cell surfaces. We show that a molecular layer seen in crystal structures of E- and N-cadherin ectodomains reported here and in a previous C-cadherin structure corresponds to the extracellular architecture of adherens junctions. In all three ectodomain crystals, cadherins dimerize through a trans adhesive interface and are connected by a second, cis, interface. Assemblies formed by E-cadherin ectodomains coated on liposomes also appear to adopt this structure. Fluorescent imaging of junctions formed from wild-type and mutant E-cadherins in cultured cells confirm conclusions derived from structural evidence. Mutations that interfere with the trans interface ablate adhesion, whereas cis interface mutations disrupt stable junction formation. Our observations are consistent with a model for junction assembly involving strong trans and weak cis interactions localized in the ectodomain.

  17. Synaptopodin couples epithelial contractility to α-actinin-4–dependent junction maturation

    PubMed Central

    Kannan, Nivetha

    2015-01-01

    The epithelial junction experiences mechanical force exerted by endogenous actomyosin activities and from interactions with neighboring cells. We hypothesize that tension generated at cell–cell adhesive contacts contributes to the maturation and assembly of the junctional complex. To test our hypothesis, we used a hydraulic apparatus that can apply mechanical force to intercellular junction in a confluent monolayer of cells. We found that mechanical force induces α-actinin-4 and actin accumulation at the cell junction in a time- and tension-dependent manner during junction development. Intercellular tension also induces α-actinin-4–dependent recruitment of vinculin to the cell junction. In addition, we have identified a tension-sensitive upstream regulator of α-actinin-4 as synaptopodin. Synaptopodin forms a complex containing α-actinin-4 and β-catenin and interacts with myosin II, indicating that it can physically link adhesion molecules to the cellular contractile apparatus. Synaptopodin depletion prevents junctional accumulation of α-actinin-4, vinculin, and actin. Knockdown of synaptopodin and α-actinin-4 decreases the strength of cell–cell adhesion, reduces the monolayer permeability barrier, and compromises cellular contractility. Our findings underscore the complexity of junction development and implicate a control process via tension-induced sequential incorporation of junctional components. PMID:26504173

  18. Electrical conductivity of single molecular junctions assembled from Co- and Co3C-encapsulating carbon nanocapsules.

    PubMed

    Matsuura, Daisuke; Kizuka, Tokushi

    2014-03-01

    Single molecular junctions (SMJs) were assembled from cobalt (Co)- and Co carbide (Co3C)-encapsulating carbon nanocapsules (CNCs) and two gold electrodes inside a high-resolution transmission electron microscope equipped with a specimen-piezomanipulation system. The structure and electrical transport properties of the SMJs were investigated in situ. The current density depended on the perimeter of the contact area between CNCs and the electrodes, showing that the current flowed not through the encapsulated region but rather along the graphene layers of CNCs. It was demonstrated that the properties of graphene can be applied to nanodevices using CNCs irrespective of the encapsulating materials.

  19. Connexin26 regulates assembly and maintenance of cochlear gap junction macromolecular complex for normal hearing

    NASA Astrophysics Data System (ADS)

    Kamiya, Kazusaku; Fukunaga, Ichiro; Hatakeyama, Kaori; Ikeda, Katsuhisa

    2015-12-01

    Hereditary deafness affects about 1 in 2000 children and GJB2 gene mutation is most frequent cause for this disease in the world. GJB2 encodes connexin26 (Cx26), a component in cochlear gap junction. Recently, we found macromolecular change of gap junction plaques with two different types of Cx26 mutation as major classification of clinical case, one is a model of dominant negative type, Cx26R75W+ and the other is conditional gene deficient mouse, Cx26f/fP0Cre as a model for insufficiency of gap junction protein [6]. Gap junction composed mainly of Cx26 and Cx30 in wild type mice formed large planar gap junction plaques (GJP). In contrast, Cx26R75W+ and Cx26f/fP0Cre showed fragmented small round GJPs around the cell border. In Cx26f/fP0Cre, some of the cells with Cx26 expression due to their cellular mosaicism showed normal large GJP with Cx26 and Cx30 only at the cell junction site between two Cx26 positive cells. These indicate that bilateral Cx26 expressions from both adjacent cells are essential for the formation of the cochlear linear GJP, and it is not compensated by other cochlear Connexins such as Connexin30. In the present study, we demonstrated a new molecular pathology in most common hereditary deafness with different types of Connexin26 mutations, and this machinery can be a new target for drag design of hereditary deafness.

  20. Systems and assemblies for transferring high power laser energy through a rotating junction

    DOEpatents

    Norton, Ryan J.; McKay, Ryan P.; Fraze, Jason D.; Rinzler, Charles C.; Grubb, Daryl L.; Faircloth, Brian O.; Zediker, Mark S.

    2016-01-26

    There are provided high power laser devices and systems for transmitting a high power laser beam across a rotating assembly, including optical slip rings and optical rotational coupling assemblies. These devices can transmit the laser beam through the rotation zone in free space or within a fiber.

  1. Assembly of the cochlear gap junction macromolecular complex requires connexin 26.

    PubMed

    Kamiya, Kazusaku; Yum, Sabrina W; Kurebayashi, Nagomi; Muraki, Miho; Ogawa, Kana; Karasawa, Keiko; Miwa, Asuka; Guo, Xueshui; Gotoh, Satoru; Sugitani, Yoshinobu; Yamanaka, Hitomi; Ito-Kawashima, Shioko; Iizuka, Takashi; Sakurai, Takashi; Noda, Tetsuo; Minowa, Osamu; Ikeda, Katsuhisa

    2014-04-01

    Hereditary deafness affects approximately 1 in 2,000 children. Mutations in the gene encoding the cochlear gap junction protein connexin 26 (CX26) cause prelingual, nonsyndromic deafness and are responsible for as many as 50% of hereditary deafness cases in certain populations. Connexin-associated deafness is thought to be the result of defective development of auditory sensory epithelium due to connexion dysfunction. Surprisingly, CX26 deficiency is not compensated for by the closely related connexin CX30, which is abundantly expressed in the same cochlear cells. Here, using two mouse models of CX26-associated deafness, we demonstrate that disruption of the CX26-dependent gap junction plaque (GJP) is the earliest observable change during embryonic development of mice with connexin-associated deafness. Loss of CX26 resulted in a drastic reduction in the GJP area and protein level and was associated with excessive endocytosis with increased expression of caveolin 1 and caveolin 2. Furthermore, expression of deafness-associated CX26 and CX30 in cell culture resulted in visible disruption of GJPs and loss of function. Our results demonstrate that deafness-associated mutations in CX26 induce the macromolecular degradation of large gap junction complexes accompanied by an increase in caveolar structures.

  2. Occludin oligomeric assemblies at tight junctions of the blood-brain barrier are altered by hypoxia and reoxygenation stress.

    PubMed

    McCaffrey, Gwen; Willis, Colin L; Staatz, William D; Nametz, Nicole; Quigley, Carolyn A; Hom, Sharon; Lochhead, Jeffrey J; Davis, Thomas P

    2009-07-01

    Hypoxic (low oxygen) and reperfusion (post-hypoxic reoxygenation) phases of stroke promote an increase in microvascular permeability at tight junctions (TJs) of the blood-brain barrier (BBB) that may lead to cerebral edema. To investigate the effect of hypoxia (Hx) and reoxygenation on oligomeric assemblies of the transmembrane TJ protein occludin, rats were subjected to either normoxia (Nx, 21% O(2), 60 min), Hx (6% O(2), 60 min), or hypoxia/reoxygenation (H/R, 6% O(2), 60 min followed by 21% O(2), 10 min). After treatment, cerebral microvessels were isolated, fractionated by detergent-free density gradient centrifugation, and occludin oligomeric assemblies associated with plasma membrane lipid rafts were solubilized by perfluoro-octanoic acid (PFO) exclusively as high molecular weight protein complexes. Analysis by non-reducing and reducing sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis/western blot of PFO-solubilized occludin revealed that occludin oligomeric assemblies co-localizing with 'TJ-associated' raft domains contained a high molecular weight 'structural core' that was resistant to disassembly by either SDS or a hydrophilic reducing agent ex vivo, and by Hx and H/R conditions in vivo. However, exposure of PFO-solubilized occludin oligomeric assemblies to SDS ex vivo revealed the non-covalent association of a significant amount of dimeric and monomeric occludin isoforms to the disulfide-bonded inner core, and dispersal of these non-covalently attached occludin subunits to lipid rafts of higher density in vivo was differentially promoted by Hx and H/R. Our data suggest a model of isoform interaction within occludin oligomeric assemblies at the BBB that enables occludin to simultaneously perform a structural role in inhibiting paracellular diffusion, and a signaling role involving interactions of dimeric and monomeric occludin isoforms with a variety of regulatory molecules within different plasma membrane lipid raft domains.

  3. Self-assembled molecular p/n junctions for applications in dye-sensitized solar energy conversion.

    PubMed

    Farnum, Byron H; Wee, Kyung-Ryang; Meyer, Thomas J

    2016-09-01

    The achievement of long-lived photoinduced redox separation lifetimes has long been a central goal of molecular-based solar energy conversion strategies. The longer the redox-separation lifetime, the more time available for useful work to be extracted from the absorbed photon energy. Here we describe a novel strategy for dye-sensitized solar energy applications in which redox-separated lifetimes on the order of milliseconds to seconds can be achieved based on a simple toolkit of molecular components. Specifically, molecular chromophores (C), electron acceptors (A) and electron donors (D) were self-assembled on the surfaces of mesoporous, transparent conducting indium tin oxide nanoparticle (nanoITO) electrodes to prepare both photoanode (nanoITO|-A-C-D) and photocathode (nanoITO|-D-C-A) assemblies. Nanosecond transient-absorption and steady-state photolysis measurements show that the electrodes function microscopically as molecular analogues of semiconductor p/n junctions. These results point to a new chemical strategy for dye-sensitized solar energy conversion based on molecular excited states and electron acceptors/donors on the surfaces of transparent conducting oxide nanoparticle electrodes.

  4. Intracellular Transport, Assembly, and Degradation of Wild-Type and Disease-linked Mutant Gap Junction Proteins

    PubMed Central

    VanSlyke, Judy K.; Deschenes, Suzanne M.; Musil, Linda S.

    2000-01-01

    More than 130 different mutations in the gap junction integral plasma membrane protein connexin32 (Cx32) have been linked to the human peripheral neuropathy X-linked Charcot–Marie–Tooth disease (CMTX). How these various mutants are processed by the cell and the mechanism(s) by which they cause CMTX are unknown. To address these issues, we have studied the intracellular transport, assembly, and degradation of three CMTX-linked Cx32 mutants stably expressed in PC12 cells. Each mutant had a distinct fate: E208K Cx32 appeared to be retained in the endoplasmic reticulum (ER), whereas both the E186K and R142W mutants were transported to perinuclear compartments from which they trafficked either to lysosomes (R142W Cx32) or back to the ER (E186K Cx32). Despite these differences, each mutant was soluble in nonionic detergent but unable to assemble into homomeric connexons. Degradation of both mutant and wild-type connexins was rapid (t1/2 < 3 h) and took place at least in part in the ER by a process sensitive to proteasome inhibitors. The mutants studied are therefore unlikely to cause disease by accumulating in degradation-resistant aggregates but instead are efficiently cleared from the cell by quality control processes that prevent abnormal connexin molecules from traversing the secretory pathway. PMID:10848620

  5. Sample Preconcentration Utilizing Nanofractures Generated by Junction Gap Breakdown Assisted by Self-Assembled Monolayer of Gold Nanoparticles

    PubMed Central

    Jen, Chun-Ping; Amstislavskaya, Tamara G.; Chen, Kuan-Fu; Chen, Yu-Hung

    2015-01-01

    The preconcentration of proteins with low concentrations can be used to increase the sensitivity and accuracy of detection. A nonlinear electrokinetic flow is induced in a nanofluidic channel due to the overlap of electrical double layers, resulting in the fast accumulation of proteins, referred to as the exclusion-enrichment effect. The proposed chip for protein preconcentration was fabricated using simple standard soft lithography with a polydimethylsiloxane replica. This study extends our previous paper, in which gold nanoparticles were manually deposited onto the surface of a protein preconcentrator. In the present work, nanofractures were formed by utilizing the self-assembly of gold-nanoparticle-assisted electric breakdown. This reliable method for nanofracture formation, involving self-assembled monolayers of nanoparticles at the junction gap between microchannels, also decreases the required electric breakdown voltage. The experimental results reveal that a high concentration factor of 1.5×104 for a protein sample with an extremely low concentration of 1 nM was achieved in 30 min by using the proposed chip, which is faster than our previously proposed chip at the same conditions. Moreover, an immunoassay of bovine serum albumin (BSA) and anti-BSA was carried out to demonstrate the applicability of the proposed chip. PMID:25970592

  6. Self-assembled molecular p/n junctions for applications in dye-sensitized solar energy conversion

    NASA Astrophysics Data System (ADS)

    Farnum, Byron H.; Wee, Kyung-Ryang; Meyer, Thomas J.

    2016-09-01

    The achievement of long-lived photoinduced redox separation lifetimes has long been a central goal of molecular-based solar energy conversion strategies. The longer the redox-separation lifetime, the more time available for useful work to be extracted from the absorbed photon energy. Here we describe a novel strategy for dye-sensitized solar energy applications in which redox-separated lifetimes on the order of milliseconds to seconds can be achieved based on a simple toolkit of molecular components. Specifically, molecular chromophores (C), electron acceptors (A) and electron donors (D) were self-assembled on the surfaces of mesoporous, transparent conducting indium tin oxide nanoparticle (nanoITO) electrodes to prepare both photoanode (nanoITO|-A-C-D) and photocathode (nanoITO|-D-C-A) assemblies. Nanosecond transient-absorption and steady-state photolysis measurements show that the electrodes function microscopically as molecular analogues of semiconductor p/n junctions. These results point to a new chemical strategy for dye-sensitized solar energy conversion based on molecular excited states and electron acceptors/donors on the surfaces of transparent conducting oxide nanoparticle electrodes.

  7. Optical modulation of nano-gap tunnelling junctions comprising self-assembled monolayers of hemicyanine dyes

    PubMed Central

    Pourhossein, Parisa; Vijayaraghavan, Ratheesh K.; Meskers, Stefan C. J.; Chiechi, Ryan C.

    2016-01-01

    Light-driven conductance switching in molecular tunnelling junctions that relies on photoisomerization is constrained by the limitations of kinetic traps and either by the sterics of rearranging atoms in a densely packed monolayer or the small absorbance of individual molecules. Here we demonstrate light-driven conductance gating; devices comprising monolayers of hemicyanine dyes trapped between two metallic nanowires exhibit higher conductance under irradiation than in the dark. The modulation of the tunnelling current occurs faster than the timescale of the measurement (∼1 min). We propose a mechanism in which a fraction of molecules enters an excited state that brings the conjugated portion of the monolayer into resonance with the electrodes. This mechanism is supported by calculations showing the delocalization of molecular orbitals near the Fermi energy in the excited and cationic states, but not the ground state and a reasonable change in conductance with respect to the effective barrier width. PMID:27272394

  8. A Single-Level Tunnel Model to Account for Electrical Transport through Single Molecule- and Self-Assembled Monolayer-based Junctions

    PubMed Central

    Garrigues, Alvar R.; Yuan, Li; Wang, Lejia; Mucciolo, Eduardo R.; Thompon, Damien; del Barco, Enrique; Nijhuis, Christian A.

    2016-01-01

    We present a theoretical analysis aimed at understanding electrical conduction in molecular tunnel junctions. We focus on discussing the validity of coherent versus incoherent theoretical formulations for single-level tunneling to explain experimental results obtained under a wide range of experimental conditions, including measurements in individual molecules connecting the leads of electromigrated single-electron transistors and junctions of self-assembled monolayers (SAM) of molecules sandwiched between two macroscopic contacts. We show that the restriction of transport through a single level in solid state junctions (no solvent) makes coherent and incoherent tunneling formalisms indistinguishable when only one level participates in transport. Similar to Marcus relaxation processes in wet electrochemistry, the thermal broadening of the Fermi distribution describing the electronic occupation energies in the electrodes accounts for the exponential dependence of the tunneling current on temperature. We demonstrate that a single-level tunnel model satisfactorily explains experimental results obtained in three different molecular junctions (both single-molecule and SAM-based) formed by ferrocene-based molecules. Among other things, we use the model to map the electrostatic potential profile in EGaIn-based SAM junctions in which the ferrocene unit is placed at different positions within the molecule, and we find that electrical screening gives rise to a strongly non-linear profile across the junction. PMID:27216489

  9. Printing-based assembly of quadruple-junction four-terminal microscale solar cells and their use in high-efficiency modules.

    PubMed

    Sheng, Xing; Bower, Christopher A; Bonafede, Salvatore; Wilson, John W; Fisher, Brent; Meitl, Matthew; Yuen, Homan; Wang, Shuodao; Shen, Ling; Banks, Anthony R; Corcoran, Christopher J; Nuzzo, Ralph G; Burroughs, Scott; Rogers, John A

    2014-06-01

    Expenses associated with shipping, installation, land, regulatory compliance and on-going maintenance and operations of utility-scale photovoltaics can be significantly reduced by increasing the power conversion efficiency of solar modules through improved materials, device designs and strategies for light management. Single-junction cells have performance constraints defined by their Shockley-Queisser limits. Multi-junction cells can achieve higher efficiencies, but epitaxial and current matching requirements between the single junctions in the devices hinder progress. Mechanical stacking of independent multi-junction cells circumvents these disadvantages. Here we present a fabrication approach for the realization of mechanically assembled multi-junction cells using materials and techniques compatible with large-scale manufacturing. The strategy involves printing-based stacking of microscale solar cells, sol-gel processes for interlayers with advanced optical, electrical and thermal properties, together with unusual packaging techniques, electrical matching networks, and compact ultrahigh-concentration optics. We demonstrate quadruple-junction, four-terminal solar cells with measured efficiencies of 43.9% at concentrations exceeding 1,000 suns, and modules with efficiencies of 36.5%.

  10. Printing-based assembly of quadruple-junction four-terminal microscale solar cells and their use in high-efficiency modules

    NASA Astrophysics Data System (ADS)

    Sheng, Xing; Bower, Christopher A.; Bonafede, Salvatore; Wilson, John W.; Fisher, Brent; Meitl, Matthew; Yuen, Homan; Wang, Shuodao; Shen, Ling; Banks, Anthony R.; Corcoran, Christopher J.; Nuzzo, Ralph G.; Burroughs, Scott; Rogers, John A.

    2014-06-01

    Expenses associated with shipping, installation, land, regulatory compliance and on-going maintenance and operations of utility-scale photovoltaics can be significantly reduced by increasing the power conversion efficiency of solar modules through improved materials, device designs and strategies for light management. Single-junction cells have performance constraints defined by their Shockley-Queisser limits. Multi-junction cells can achieve higher efficiencies, but epitaxial and current matching requirements between the single junctions in the devices hinder progress. Mechanical stacking of independent multi-junction cells circumvents these disadvantages. Here we present a fabrication approach for the realization of mechanically assembled multi-junction cells using materials and techniques compatible with large-scale manufacturing. The strategy involves printing-based stacking of microscale solar cells, sol-gel processes for interlayers with advanced optical, electrical and thermal properties, together with unusual packaging techniques, electrical matching networks, and compact ultrahigh-concentration optics. We demonstrate quadruple-junction, four-terminal solar cells with measured efficiencies of 43.9% at concentrations exceeding 1,000 suns, and modules with efficiencies of 36.5%.

  11. Mesenchymal-epithelial transitions: spontaneous and cumulative syntheses of epithelial marker molecules and their assemblies to novel cell junctions connecting human hematopoietic tumor cells to carcinomatoid tissue structures.

    PubMed

    Franke, Werner W; Rickelt, Steffen

    2011-12-01

    Using biochemical as well as light- and electron-microscopic immunolocalization methods, in cultures of unicellular human blood tumor cells, we have studied the phenomenon of spontaneous and cumulative syntheses of certain epithelial proteins and glycoproteins and their assemblies to two major kinds of novel cell-cell junctions, adhering junctions (AJs) and junctions based on the epithelial cell adhesion molecule (EpCAM). More than two decades, we have selected and characterized clonal sublines of multipotential hematopoietic K562 cells, which are enriched in newly formed AJs based on cis-clusters of desmoglein Dsg2, in some sublines accompanied by desmocollin Dsc2. Both desmosomal cadherins can be anchored in a submembranous plaque containing plakoglobin and plakophilins Pkp2 and Pkp3, with or without other armadillo proteins and desmoplakin. Also, these cells are often connected by an additional, extended junction system, in which the transmembrane epithelial glycoprotein EpCAM is associated with a cytoplasmic plaque rich in several actin-binding proteins such as afadin, α-actinin, ezrin and vinculin. Both kinds of junctions contribute to connections of K562 cells into epithelioid monolayers or even three-dimensional, tissue-like structures, thus markedly changing the cell biological nature and behavior of the resulting tumor subforms (mesenchymal-epithelial transitions). We discuss molecular mechanisms involved in the formation and function of these junctions, also with respect to tumor spread and metastasis, as well as diagnostic and therapeutic consequences.

  12. Desmosome assembly and dynamics.

    PubMed

    Nekrasova, Oxana; Green, Kathleen J

    2013-11-01

    Desmosomes are intercellular junctions that anchor intermediate filaments (IFs) to the plasma membrane, forming a supracellular scaffold that provides mechanical resilience to tissues. This anchoring function is accomplished by specialized members of the cadherin family and associated cytoskeletal linking proteins, which together form a highly organized membrane core flanked by mirror-image cytoplasmic plaques. Due to the biochemical insolubility of desmosomes, the mechanisms that govern assembly of these components into a functional organelle remained elusive. Recently developed molecular reporters and live cell imaging approaches have provided powerful new tools to monitor this finely tuned process in real time. Here we discuss studies that are beginning to decipher the machinery and regulation governing desmosome assembly and homeostasis in situ and how these mechanisms are affected during disease pathogenesis.

  13. Fluid shear, intercellular stress, and endothelial cell alignment.

    PubMed

    Steward, Robert; Tambe, Dhananjay; Hardin, C Corey; Krishnan, Ramaswamy; Fredberg, Jeffrey J

    2015-04-15

    Endothelial cell alignment along the direction of laminar fluid flow is widely understood to be a defining morphological feature of vascular homeostasis. While the role of associated signaling and structural events have been well studied, associated intercellular stresses under laminar fluid shear have remained ill-defined and the role of these stresses in the alignment process has remained obscure. To fill this gap, we report here the tractions as well as the complete in-plane intercellular stress fields measured within the human umbilical vein endothelial cell (HUVEC) monolayer subjected to a steady laminar fluid shear of 1 Pa. Tractions, intercellular stresses, as well as their time course, heterogeneity, and anisotropy, were measured using monolayer traction microscopy and monolayer stress microscopy. Prior to application of laminar fluid flow, intercellular stresses were largely tensile but fluctuated dramatically in space and in time (317 ± 122 Pa). Within 12 h of the onset of laminar fluid flow, the intercellular stresses decreased substantially but continued to fluctuate dramatically (142 ± 84 Pa). Moreover, tractions and intercellular stresses aligned strongly and promptly (within 1 h) along the direction of fluid flow, whereas the endothelial cell body aligned less strongly and substantially more slowly (12 h). Taken together, these results reveal that steady laminar fluid flow induces prompt reduction in magnitude and alignment of tractions and intercellular stress tensor components followed by the retarded elongation and alignment of the endothelial cell body. Appreciably smaller intercellular stresses supported by cell-cell junctions logically favor smaller incidence of gap formation and thus improved barrier integrity.

  14. Structures and electrical properties of single nanoparticle junctions assembled using LaC2-encapsulating carbon nanocapsules

    PubMed Central

    Tezura, Manabu; Kizuka, Tokushi

    2016-01-01

    As the miniaturization of integrated circuits advances, electronics using single molecules and nanosize particles are being studied increasingly. Single nanoparticle junctions (SNPJs) consist of two electrodes sandwiching a single nanoparticle. Nanocarbons with nanospaces in their center, such as fullerenes, carbon nanotubes, and carbon nanocapsules (CNCs), are expected to be elements of advanced SNPJs. In this study, SNPJs were assembled using lanthanum dicarbide (LaC2)-encapsulating CNCs and two gold (Au) electrodes by a nanotip operation inside a high-resolution transmission electron microscope. The atomic configuration and electrical resistance of the SNPJs were investigated in situ. The results implied that the electrical resistance of the SNPJ depended on the interface structures of the contacts between the CNC and Au electrodes, i.e., the contact electrical resistance, and the greatest portion of the current through the SNPJ flowed along the outermost carbon layer of the CNC. Thus, the resistance of the SNPJs using the CNCs was demonstrated and the electrical conduction mechanism of one of the CNC was discussed in this study. PMID:27412856

  15. Thiol-terminated monolayers on oxide-free Si: assembly of semiconductor-alkyl-S-metal junctions.

    PubMed

    Böcking, Till; Salomon, Adi; Cahen, David; Gooding, J Justin

    2007-03-13

    Self-assembled monolayers formed by thermal hydrosilylation of a trifluoroacetyl-protected alkenylthiol on Si-H surfaces, followed by removal of the protecting groups, yield essentially oxide-free monolayers suitable for the formation of Si-C11H22-S-Hg and Si-C11H22-S-Au junctions in which the alkyl chains are chemically bound to the silicon surface (via Si-C bonds) and the metal electrode (via Hg-S or Au-S bonds). Two barriers to charge transport are present in the system: at low bias the current is temperature activated and hence limited by thermionic emission over the Schottky barrier in the silicon, whereas as at high bias transport is limited by tunneling through the organic monolayer. The thiol-terminated monolayer on oxide-free silicon provides a well-characterized system allowing a careful study of the importance of the interfacial bond to the metal electrode for current transport through saturated molecules.

  16. Structures and electrical properties of single nanoparticle junctions assembled using LaC2-encapsulating carbon nanocapsules

    NASA Astrophysics Data System (ADS)

    Tezura, Manabu; Kizuka, Tokushi

    2016-07-01

    As the miniaturization of integrated circuits advances, electronics using single molecules and nanosize particles are being studied increasingly. Single nanoparticle junctions (SNPJs) consist of two electrodes sandwiching a single nanoparticle. Nanocarbons with nanospaces in their center, such as fullerenes, carbon nanotubes, and carbon nanocapsules (CNCs), are expected to be elements of advanced SNPJs. In this study, SNPJs were assembled using lanthanum dicarbide (LaC2)-encapsulating CNCs and two gold (Au) electrodes by a nanotip operation inside a high-resolution transmission electron microscope. The atomic configuration and electrical resistance of the SNPJs were investigated in situ. The results implied that the electrical resistance of the SNPJ depended on the interface structures of the contacts between the CNC and Au electrodes, i.e., the contact electrical resistance, and the greatest portion of the current through the SNPJ flowed along the outermost carbon layer of the CNC. Thus, the resistance of the SNPJs using the CNCs was demonstrated and the electrical conduction mechanism of one of the CNC was discussed in this study.

  17. Gap junctions and tissue business: problems and strategies for developing specific functional reagents.

    PubMed

    Goodenough, D A; Musil, L S

    1993-01-01

    The complex and overlapping tissue distribution of different members of the gap junctional connexin protein family is reviewed. Intermixing of different connexins in the building of intercellular channels and translational and posttranslational regulation of gap junctional channels add additional challenges to the interpretation of the possible functions played by gap junction-mediated intercellular communication in tissue business.

  18. Intercellular Ca2+ Waves: Mechanisms and Function

    PubMed Central

    Sanderson, Michael J.

    2012-01-01

    Intercellular calcium (Ca2+) waves (ICWs) represent the propagation of increases in intracellular Ca2+ through a syncytium of cells and appear to be a fundamental mechanism for coordinating multicellular responses. ICWs occur in a wide diversity of cells and have been extensively studied in vitro. More recent studies focus on ICWs in vivo. ICWs are triggered by a variety of stimuli and involve the release of Ca2+ from internal stores. The propagation of ICWs predominately involves cell communication with internal messengers moving via gap junctions or extracellular messengers mediating paracrine signaling. ICWs appear to be important in both normal physiology as well as pathophysiological processes in a variety of organs and tissues including brain, liver, retina, cochlea, and vascular tissue. We review here the mechanisms of initiation and propagation of ICWs, the key intra- and extracellular messengers (inositol 1,4,5-trisphosphate and ATP) mediating ICWs, and the proposed physiological functions of ICWs. PMID:22811430

  19. Structure of components of an intercellular channel complex in sporulating Bacillus subtilis

    PubMed Central

    Levdikov, Vladimir M.; Blagova, Elena V.; McFeat, Amanda; Fogg, Mark J.; Wilson, Keith S.; Wilkinson, Anthony J.

    2012-01-01

    Following asymmetric cell division during spore formation in Bacillus subtilis, a forespore expressed membrane protein SpoIIQ, interacts across an intercellular space with a mother cell-expressed membrane protein, SpoIIIAH. Their interaction can serve as a molecular “ratchet” contributing to the migration of the mother cell membrane around that of the forespore in a phagocytosis-like process termed engulfment. Upon completion of engulfment, SpoIIQ and SpoIIIAH are integral components of a recently proposed intercellular channel allowing passage from the mother cell into the forespore of factors required for late gene expression in this compartment. Here we show that the extracellular domains of SpoIIQ and SpoIIIAH form a heterodimeric complex in solution. The crystal structure of this complex reveals that SpoIIQ has a LytM-like zinc-metalloprotease fold but with an incomplete zinc coordination sphere and no metal. SpoIIIAH has an α-helical subdomain and a protruding β-sheet subdomain, which mediates interactions with SpoIIQ. SpoIIIAH has sequence and structural homology to EscJ, a type III secretion system protein that forms a 24-fold symmetric ring. Superposition of the structures of SpoIIIAH and EscJ reveals that the SpoIIIAH protomer overlaps with two adjacent protomers of EscJ, allowing us to generate a dodecameric SpoIIIAH ring by using structural homology. Following this superposition, the SpoIIQ chains also form a closed dodecameric ring abutting the SpoIIIAH ring, producing an assembly surrounding a 60 Å channel. The dimensions and organization of the proposed complex suggest it is a plausible model for the extracellular component of a gap junction-like intercellular channel. PMID:22431604

  20. Hepatocyte growth factor/scatter factor effects on epithelia. Regulation of intercellular junctions in transformed and nontransformed cell lines, basolateral polarization of c-met receptor in transformed and natural intestinal epithelia, and induction of rapid wound repair in a transformed model epithelium.

    PubMed Central

    Nusrat, A; Parkos, C A; Bacarra, A E; Godowski, P J; Delp-Archer, C; Rosen, E M; Madara, J L

    1994-01-01

    Intestinal epithelial cells rest on a fibroblast sheath. Thus, factors produced by these fibroblasts may influence epithelial function in a paracrine fashion. We examined modulation of intestinal epithelial function by one such fibroblast product, scatter factor/hepatocyte growth factor (HGF/SF). This effect was studied in vitro by using model T84 intestinal epithelial cells. When applied to confluent T84 monolayers, HGF/SF attenuates transepithelial resistance to passive ion flow in a dose-dependent manner (maximum fall at 300 ng/ml, 28% control monolayer resistance, P < 0.001, ED50 of 1.2 nM), t1/2 of 20 h. This functional effect of HGF/SF and distribution of its receptor, c-met, are polarized to the basolateral membranes of T84 intestinal epithelial cells. HGF/SF effects on resistance are not attributable to altered transcellular resistance (opening of Cl- and/or basolateral K+ channels), cytotoxicity, or enhanced cell proliferation; they therefore represent specific regulation of paracellular tight junction resistance. Analysis with biochemically purified rodent HGF/SF and Madin-Darby canine kidney cells reveals that effects on paracellular tight junctions also occur in other nontransformed epithelia. Binding of HGF/SF to its receptor in T84 intestinal epithelial cells is accompanied by tyrosine phosphorylation of the receptor. Because loosening of intercellular junctions between cells could facilitate separation, spreading, and migration of epithelial cells during physiologic processes such as wound resealing, we determined the effects of HGF/SF on intestinal epithelial wound resealing using our previously published in vitro model (Nusrat, A., C. Delp, and J. L. Madara. 1992. J. Clin. Invest. 89:1501-1511). HGF/SF markedly enhanced wound closure (> 450% increase in rate, P < 0.001) by influencing the migratory and spreading response in not only cells adjoining the wound but also cells many positions removed from the wound. We thus speculate that HGF/SF may

  1. Intercellular communication and human prostate carcinogenesis.

    PubMed

    Carruba, Giuseppe; Stefano, Rosalba; Cocciadiferro, Letizia; Saladino, Francesca; Di Cristina, Antonietta; Tokar, Erik; Quader, Salmann T A; Webber, Mukta M; Castagnetta, Luigi

    2002-06-01

    Gap-junction-mediated intercellular communication (GJIC) is required for completion of embryonic development, tissue homeostasis, and regulation of cell proliferation and death. Although, as emphasized in several reports, defects or disruption of GJIC may be important in carcinogenesis, the potential role of GJIC in the onset and progression of human prostate cancer remains ill-defined. The gap junction channel-forming connexins (Cx) comprise a multigene family of highly conserved proteins that are differentially expressed in a tissue- and development-specific manner; changes in connexin expression are also commonly seen during cellular differentiation. However, when multiple connexins are concurrently expressed, gap junction channels may consist of more than one connexin species. This is important, because only certain pairings give rise to functional channels. In our studies, we investigated GJIC in a panel of both nontumorigenic (RWPE-1) and malignant (RWPE-2, LNCaP, DU-145) human prostate epithelial cells, compared to a normal rat liver epithelial F344 (WB-1) cell line, as it was found to be junctionally proficient. In addition, expression and regulation of Cx43 and Cx32 were also inspected using western blot analysis. The ability of hormones, antihormones, and the antihypertensive drug forskolin to restore GJIC in nontumorigenic and malignant human prostate epithelial cells was examined by the scrape-loading/dye transfer (SL/DT) or fluorescence recovery after photobleaching (FRAP) methods using an Ultima laser cytometer. Results from both assays showed that neither nontumorigenic nor malignant prostate cells have functional GJIC. However, both estrone (E1) and forskolin (FK) induced a significant increase (4.4- and 2.8-fold, respectively) in cell-cell communication only in the RWPE-1 cells. Interestingly, the use of Matrigel, a solubilized basement membrane, as substrate for cell attachment and growth resulted in the rescue of GJIC activity in RWPE-1 cells, as

  2. On the ovarian bursa of the golden hamster. II. Intercellular connections in the bursal epithelium and passage of ferritin from the cavity into lymphatics.

    PubMed Central

    Nakatani, T; Shinohara, H; Matsuda, T

    1986-01-01

    Cell abutments in the ovarian bursal epithelium of the golden hamster included tight junctions, desmosome-like junctions, gap junctions and zonulae adhaerentes. The plasma membranes of adjacent epithelial cells were often closely apposed forming a zonula adherens at the apex of the intercellular space, but these did not fuse along the entire length of the intercellular space. Gap junctions and other intercellular junctions did not allow diffusion of ferritin into the junctional areas, but failed to prevent ferritin from filling the intercellular space basal to the junction. This suggested that these junctions were not continuous around the entire circumference of the cell. In general, intercellular connections in the bursal epithelium may be labile and this lability, as well as periodic distention of the bursa, may give rise to separation of epithelial cells. Ferritin injected into the bursal cavity reached lymphatics via three routes: (1) via stomata; (2) via gaps and pores; (3) via intercellular spaces. Transcellular transport did not make a major contribution to permeation of ferritin across the bursal epithelium, and ferritin particles did not enter fenestrated blood vessels within 30 minutes after injection. The variety of routes may facilitate rapid and complete drainage of fluid and cellular components from the bursal cavity. Images Fig. 1 Fig. 2 Fig. 3 Fig. 4 Fig. 5 Fig. 6 Fig. 7 Fig. 8 Fig. 9 Fig. 10 Fig. 11 Figs. 12-13 Fig. 14 Fig. 15 Fig. 16 Figs. 17-18 PMID:3693080

  3. [Gap junctions and cancer: implications and perspectives].

    PubMed

    Mesnil, Marc

    2004-02-01

    Gap junctions are made of intercellular channels which permit the diffusion from cytoplasm to cytoplasm of small hydrophilic molecules (<1,200 Da) such as ions, sugars, amino acids, nucleotides, second messengers (calcium, inositol triphosphate, etc.). Since their discovery in the early sixties, several groups have described the loss of their function in cancer cells. The accumulation of such data led to the hypothesis that gap junctions are involved in the carcinogenesis process. This assumption has been confirmed by data establishing that gap junctional intercellular communication is inhibited by most of the tumor promoters and that the restoration of such a communication, by transfection of cDNAs encoding gap junction proteins (connexins), inhibits the aberrant growth rates of tumorigenic cells. Despite these important informations, several fundamental questions remain still open. First, we do not know how gap junctions mediate such a tumor suppressor effect and whether it may depend either on the cell type or on the connexin type. Moreover, most of the data concerning a possible involvement of gap junctions in carcinogenesis have been obtained from in vitro and animal models. The very few results which have been currently collected from human tumors are not sufficient to have a clear idea concerning the real involvement of gap junctions in sporadic human cancers. These points as well as other unresolved questions about the role of gap junctional intercellular communication in carcinogenesis are mentioned. To bring some answers, some prospects are proposed with the objective to use gap junctions for increasing the effect of anticancer therapies.

  4. Multifaceted Roles of Tunneling Nanotubes in Intercellular Communication

    PubMed Central

    Marzo, Ludovica; Gousset, Karine; Zurzolo, Chiara

    2012-01-01

    Cell-to-cell communication and exchange of materials are vital processes in multicellular organisms during cell development, cell repair, and cell survival. In neuronal and immunological cells, intercellular transmission between neighboring cells occurs via different complex junctions or synapses. Recently, long distance intercellular connections in mammalian cells called tunneling nanotubes (TNTs) have been described. These structures have been found in numerous cell types and shown to transfer signals and cytosolic materials between distant cells, suggesting that they might play a prominent role in intercellular trafficking. However, these cellular connections are very heterogeneous in both structure and function, giving rise to more questions than answers as to their nature and role as intercellular conduits. To better understand and characterize the functions of TNTs, we have highlighted here the latest discoveries regarding the formation, structure, and role of TNTs in cell-to-cell spreading of various signals and materials. We first gathered information regarding their formation with an emphasis on the triggering mechanisms observed, such as stress and potentially important proteins and/or signaling pathways. We then describe the various types of transfer mechanisms, in relation to signals and cargoes that have been shown recently to take advantage of these structures for intercellular transfer. Because a number of pathogens were shown to use these membrane bridges to spread between cells we also draw attention to specific studies that point toward a role for TNTs in pathogen spreading. In particular we discuss the possible role that TNTs might play in prion spreading, and speculate on their role in neurological diseases in general. PMID:22514537

  5. Gene correction of integrin beta4-dependent pyloric atresia-junctional epidermolysis bullosa keratinocytes establishes a role for beta4 tyrosines 1422 and 1440 in hemidesmosome assembly.

    PubMed

    Dellambra, E; Prislei, S; Salvati, A L; Madeddu, M L; Golisano, O; Siviero, E; Bondanza, S; Cicuzza, S; Orecchia, A; Giancotti, F G; Zambruno, G; De Luca, M

    2001-11-02

    The cytoplasmic domain of beta4 integrin contains two pairs of fibronectin-like repeats separated by a connecting segment. The connecting segment harbors a putative tyrosine activation motif in which tyrosines 1422 and 1440 are phosphorylated in response to alpha6beta4 binding to laminin-5. Primary beta4-null keratinocytes, obtained from a newborn suffering from lethal junctional epidermolysis bullosa, were stably transduced with retroviruses carrying a full-length beta4 cDNA or a beta4 cDNA with phenylalanine substitutions at Tyr-1422 and Tyr-1440. Hemidesmosome assembly was evaluated on organotypic skin cultures. beta4-corrected keratinocytes were indistinguishable from normal cells in terms of alpha6beta4 expression, the localization of hemidesmosome components, and hemidesmosome structure and density, suggesting full genetic and functional correction of beta4-null keratinocytes. In cultures generated from beta4(Y1422F/Y1440F) keratinocytes, beta4 mutants as well as alpha6 integrin, HD1/plectin, and BP180 were not concentrated at the dermal-epidermal junction. Furthermore, the number of hemidesmosomes was strikingly reduced as compared with beta4-corrected keratinocytes. The rare hemidesmosomes detected in beta4(Y1422F/Y1440F) cells were devoid of sub-basal dense plates and of inner cytoplasmic plaques with keratin filament insertion. Collectively, our data demonstrate that the beta4 tyrosine activation motif is not required for the localization of alpha6beta4 at the keratinocyte plasma membrane but is essential for optimal assembly of bona fide hemidesmosomes.

  6. Integration and Modulation of Intercellular Signaling Underlying Blood Flow Control

    PubMed Central

    Segal, Steven S.

    2015-01-01

    Vascular resistance networks control tissue blood flow in concert with regulating arterial perfusion pressure. In response to increased metabolic demand, vasodilation arising in arteriolar networks ascends to encompass proximal feed arteries. By reducing resistance upstream, ascending vasodilation (AVD) increases blood flow into the microcirculation. Once initiated [e.g., through local activation of K+ channels in endothelial cells (ECs)], hyperpolarization is conducted through gap junctions along the endothelium. Via EC projections through the internal elastic lamina, hyperpolarization spreads into the surrounding smooth muscle cells (SMCs) through myoendothelial gap junctions (MEGJs) to promote their relaxation. Intercellular signaling through electrical signal transmission (i.e., cell-to-cell conduction) can thereby coordinate vasodilation along and among the branches of microvascular resistance networks. Perivascular sympathetic nerve fibers course through the adventitia and release norepinephrine to stimulate SMCs via α-adrenoreceptors to produce contraction. In turn, SMCs can signal ECs through MEGJs to activate K+ channels and attenuate sympathetic vasoconstriction. Activation of K+ channels along the endothelium will dissipate electrical signal transmission and inhibit AVD, thereby restricting blood flow into the microcirculation while maintaining peripheral resistance and perfusion pressure. This review explores the origins and nature of intercellular signaling governing blood flow control in skeletal muscle with respect to the interplay between AVD and sympathetic innervation. Whereas these interactions are integral to physical daily activity and athletic performance, resolving the interplay between respective signaling events provides insight into how selective interventions can improve tissue perfusion and oxygen delivery during vascular disease. PMID:26368324

  7. Gap junctional communication modulates gene transcription by altering the recruitment of Sp1 and Sp3 to connexin-response elements in osteoblast promoters

    NASA Technical Reports Server (NTRS)

    Stains, Joseph P.; Lecanda, Fernando; Screen, Joanne; Towler, Dwight A.; Civitelli, Roberto

    2003-01-01

    Loss-of-function mutations of gap junction proteins, connexins, represent a mechanism of disease in a variety of tissues. We have shown that recessive (gene deletion) or dominant (connexin45 overexpression) disruption of connexin43 function results in osteoblast dysfunction and abnormal expression of osteoblast genes, including down-regulation of osteocalcin transcription. To elucidate the molecular mechanisms of gap junction-sensitive transcriptional regulation, we systematically analyzed the rat osteocalcin promoter for sensitivity to gap junctional intercellular communication. We identified an Sp1/Sp3 containing complex that assembles on a minimal element in the -70 to -57 region of the osteocalcin promoter in a gap junction-dependent manner. This CT-rich connexin-response element is necessary and sufficient to confer gap junction sensitivity to the osteocalcin proximal promoter. Repression of osteocalcin transcription occurs as a result of displacement of the stimulatory Sp1 by the inhibitory Sp3 on the promoter when gap junctional communication is perturbed. Modulation of Sp1/Sp3 recruitment also occurs on the collagen Ialpha1 promoter and translates into gap junction-sensitive transcriptional control of collagen Ialpha1 gene expression. Thus, regulation of Sp1/Sp3 recruitment to the promoter may represent a potential general mechanism for transcriptional control of target genes by signals passing through gap junctions.

  8. Modular Nuclease-Responsive DNA Three-Way Junction-Based Dynamic Assembly of a DNA Device and Its Sensing Application.

    PubMed

    Zhu, Jing; Wang, Lei; Xu, Xiaowen; Wei, Haiping; Jiang, Wei

    2016-04-05

    Here, we explored a modular strategy for rational design of nuclease-responsive three-way junctions (TWJs) and fabricated a dynamic DNA device in a "plug-and-play" fashion. First, inactivated TWJs were designed, which contained three functional domains: the inaccessible toehold and branch migration domains, the specific sites of nucleases, and the auxiliary complementary sequence. The actions of different nucleases on their specific sites in TWJs caused the close proximity of the same toehold and branch migration domains, resulting in the activation of the TWJs and the formation of a universal trigger for the subsequent dynamic assembly. Second, two hairpins (H1 and H2) were introduced, which could coexist in a metastable state, initially to act as the components for the dynamic assembly. Once the trigger initiated the opening of H1 via TWJs-driven strand displacement, the cascade hybridization of hairpins immediately switched on, resulting in the formation of the concatemers of H1/H2 complex appending numerous integrated G-quadruplexes, which were used to obtain label-free signal readout. The inherent modularity of this design allowed us to fabricate a flexible DNA dynamic device and detect multiple nucleases through altering the recognition pattern slightly. Taking uracil-DNA glycosylase and CpG methyltransferase M.SssI as models, we successfully realized the butt joint between the uracil-DNA glycosylase and M.SssI recognition events and the dynamic assembly process. Furthermore, we achieved ultrasensitive assay of nuclease activity and the inhibitor screening. The DNA device proposed here will offer an adaptive and flexible tool for clinical diagnosis and anticancer drug discovery.

  9. Distribution of the feline calicivirus receptor junctional adhesion molecule a in feline tissues.

    PubMed

    Pesavento, P A; Stokol, T; Liu, H; van der List, D A; Gaffney, P M; Parker, J S

    2011-03-01

    Junctional adhesion molecule A (JAM-A) is an immunoglobulin superfamily protein that plays an important role in the assembly and maintenance of tight junctions and the establishment of epithelial cell polarity. The feline JAM-A (fJAM-A) is a functional receptor for feline calicivirus (FCV). Among natural diseases associated with FCV infection, isolates that cause oral vesicular disease are detected in epithelial cells; however, isolates that cause systemic disease are detected in multiple cell types. The distribution of an FCV receptor or receptors in feline tissues is relevant to viral pathogenesis in that it should reflect the wide latitude of clinical sequelae associated with FCV infection. The authors examined the expression of feline JAM-A in the cat by using confocal immunofluorescence localization on normal tissues, with special regard to tissue targets of naturally occurring FCV. As described in the human and the mouse, fJAM-A was widely distributed in feline tissues, where it localized at cell-cell junctions of epithelial and endothelial cells. fJAM-A was highly expressed on feline platelets, with lower levels of expression on feline peripheral blood leukocytes. Additionally, FCV infection of a feline epithelial cell monolayer causes redistribution of fJAM-A to the cytosol of infected cells. It is reasonable to propose that the spectrum of lesions caused by FCV reflects disruption of intercellular junctions that rely on fJAM-A function and tight junctional integrity.

  10. Septal Junctions in Filamentous Heterocyst-Forming Cyanobacteria.

    PubMed

    Flores, Enrique; Herrero, Antonia; Forchhammer, Karl; Maldener, Iris

    2016-02-01

    In the filaments of heterocyst-forming cyanobacteria, septal junctions that traverse the septal peptidoglycan join adjacent cells, allowing intercellular communication. Perforations in the septal peptidoglycan have been observed, and proteins involved in the formation of such perforations and putative protein components of the septal junctions have been identified, but their relationships are debated.

  11. Cleavage of transmembrane junction proteins and their role in regulating epithelial homeostasis

    PubMed Central

    Nava, Porfirio; Kamekura, Ryuta; Nusrat, Asma

    2013-01-01

    Epithelial tissues form a selective barrier that separates the external environment from the internal tissue milieu. Single epithelial cells are densely packed and associate via distinct intercellular junctions. Intercellular junction proteins not only control barrier properties of the epithelium but also play an important role in regulating epithelial homeostasis that encompasses cell proliferation, migration, differentiation and regulated shedding. Recent studies have revealed that several proteases target epithelial junction proteins during physiological maturation as well as in pathologic states such as inflammation and cancer. This review discusses mechanisms and biological consequences of transmembrane junction protein cleavage. The influence of junction protein cleavage products on pathogenesis of inflammation and cancer is discussed. PMID:24665393

  12. Ischemic preconditioning protects against gap junctional uncoupling in cardiac myofibroblasts.

    PubMed

    Sundset, Rune; Cooper, Marie; Mikalsen, Svein-Ole; Ytrehus, Kirsti

    2004-01-01

    Ischemic preconditioning increases the heart's tolerance to a subsequent longer ischemic period. The purpose of this study was to investigate the role of gap junction communication in simulated preconditioning in cultured neonatal rat cardiac myofibroblasts. Gap junctional intercellular communication was assessed by Lucifer yellow dye transfer. Preconditioning preserved intercellular coupling after prolonged ischemia. An initial reduction in coupling in response to the preconditioning stimulus was also observed. This may protect neighboring cells from damaging substances produced during subsequent regional ischemia in vivo, and may preserve gap junctional communication required for enhanced functional recovery during subsequent reperfusion.

  13. Gap Junction in the Teleost Fish Lineage: Duplicated Connexins May Contribute to Skin Pattern Formation and Body Shape Determination

    PubMed Central

    Watanabe, Masakatsu

    2017-01-01

    Gap junctions are intercellular channels that allow passage of ions and small molecules between adjacent cells. Gap junctions in vertebrates are composed of connexons, which are an assembly of six proteins, connexins. Docking of two connexons on the opposite cell surfaces forms a gap junction between the cytoplasm of two neighboring cells. Connexins compose a family of structurally related four-pass transmembrane proteins. In mammals, there are ~20 connexins, each of which contributes to unique permeability of gap junctions, and mutations of some connexin-encoding genes are associated with human diseases. Zebrafish has been predicted to contain 39 connexin-encoding genes; the high number can be attributed to gene duplication during fish evolution, which resulted in diversified functions of gap junctions in teleosts. The determination of body shapes and skin patterns in animal species is an intriguing question. Mathematical models suggest principle mechanisms explaining the diversification of animal morphology. Recent studies have revealed the involvement of gap junctions in fish morphological diversity, including skin pattern formation and body shape determination. This review focuses on connexins in teleosts, which are integrated in the mathematical models explaining morphological diversity of animal skin patterns and body shapes. PMID:28271062

  14. Structure and function of gap junction proteins: role of gap junction proteins in embryonic heart development.

    PubMed

    Ahir, Bhavesh K; Pratten, Margaret K

    2014-01-01

    Intercellular (cell-to-cell) communication is a crucial and complex mechanism during embryonic heart development. In the cardiovascular system, the beating of the heart is a dynamic and key regulatory process, which is functionally regulated by the coordinated spread of electrical activity through heart muscle cells. Heart tissues are composed of individual cells, each bearing specialized cell surface membrane structures called gap junctions that permit the intercellular exchange of ions and low molecular weight molecules. Gap junction channels are essential in normal heart function and they assist in the mediated spread of electrical impulses that stimulate synchronized contraction (via an electrical syncytium) of cardiac tissues. This present review describes the current knowledge of gap junction biology. In the first part, we summarise some relevant biochemical and physiological properties of gap junction proteins, including their structure and function. In the second part, we review the current evidence demonstrating the role of gap junction proteins in embryonic development with particular reference to those involved in embryonic heart development. Genetics and transgenic animal studies of gap junction protein function in embryonic heart development are considered and the alteration/disruption of gap junction intercellular communication which may lead to abnormal heart development is also discussed.

  15. Axonal Ensheathment and Intercellular Barrier Formation in Drosophila

    PubMed Central

    Blauth, Kevin; Banerjee, Swati; Bhat, Manzoor A.

    2014-01-01

    Glial cells are critical players in every major aspect of nervous system development, function, and disease. Other than their traditional supportive role, glial cells perform a variety of important functions such as myelination, synapse formation and plasticity, and establishment of blood–brain and blood–nerve barriers in the nervous system. Recent studies highlight the striking functional similarities between Drosophila and vertebrate glia. In both systems, glial cells play an essential role in neural ensheathment thereby isolating the nervous system and help to create a local ionic microenvironment for conduction of nerve impulses. Here, we review the anatomical aspects and the molecular players that underlie ensheathment during different stages of nervous system development in Drosophila and how these processes lead to the organization of neuroglial junctions. We also discuss some key aspects of the invertebrate axonal ensheathment and junctional organization with that of vertebrate myelination and axon–glial interactions. Finally, we highlight the importance of intercellular junctions in barrier formation in various cellular contexts in Drosophila. We speculate that unraveling the genetic and molecular mechanisms of ensheathment across species might provide key insights into human myelin-related disorders and help in designing therapeutic interventions. PMID:20801419

  16. Self-Assembly Assisted Fabrication of Dextran-Based Nanohydrogels with Reduction-Cleavable Junctions for Applications as Efficient Drug Delivery Systems

    PubMed Central

    Wang, Hao; Dai, Tingting; Zhou, Shuyan; Huang, Xiaoxiao; Li, Songying; Sun, Kang; Zhou, Guangdong; Dou, Hongjing

    2017-01-01

    In order to overcome the key challenge in improving both fabrication efficiency and their drug delivery capability of anti-cancer drug delivery systems (ACDDS), here polyacrylic acid (PAA) grafted dextran (Dex) nanohydrogels (NGs) with covalent crosslinked structure bearing redox sensitive disulfide crosslinking junctions (Dex-SS-PAA) were synthesized efficiently through a one-step self-assembly assisted methodology (SAA). The Dex-SS-PAA were subsequently conjugated with doxorubicin through an acid-labile hydrazone bond (Dex-SS-PAA-DOX). The in vitro drug release behavior, anti-cancer effects in vivo, and biosafety of the as-prepared acid- and redox-dual responsive biodegradable NGs were systematically investigated. The results revealed that the Dex-SS-PAA-DOX exhibited pH- and redox-controlled drug release, greatly reduced the toxicity of free DOX, while exhibiting a strong ability to inhibit the growth of MDA-MB-231 tumors. Our study demonstrated that the Dex-SS-PAA-DOX NGs are very promising candidates as ACDDS for anti-cancer therapeutics. PMID:28071743

  17. Self-Assembly Assisted Fabrication of Dextran-Based Nanohydrogels with Reduction-Cleavable Junctions for Applications as Efficient Drug Delivery Systems

    NASA Astrophysics Data System (ADS)

    Wang, Hao; Dai, Tingting; Zhou, Shuyan; Huang, Xiaoxiao; Li, Songying; Sun, Kang; Zhou, Guangdong; Dou, Hongjing

    2017-01-01

    In order to overcome the key challenge in improving both fabrication efficiency and their drug delivery capability of anti-cancer drug delivery systems (ACDDS), here polyacrylic acid (PAA) grafted dextran (Dex) nanohydrogels (NGs) with covalent crosslinked structure bearing redox sensitive disulfide crosslinking junctions (Dex-SS-PAA) were synthesized efficiently through a one-step self-assembly assisted methodology (SAA). The Dex-SS-PAA were subsequently conjugated with doxorubicin through an acid-labile hydrazone bond (Dex-SS-PAA-DOX). The in vitro drug release behavior, anti-cancer effects in vivo, and biosafety of the as-prepared acid- and redox-dual responsive biodegradable NGs were systematically investigated. The results revealed that the Dex-SS-PAA-DOX exhibited pH- and redox-controlled drug release, greatly reduced the toxicity of free DOX, while exhibiting a strong ability to inhibit the growth of MDA-MB-231 tumors. Our study demonstrated that the Dex-SS-PAA-DOX NGs are very promising candidates as ACDDS for anti-cancer therapeutics.

  18. Dilated intercellular spaces as a marker of GERD.

    PubMed

    Orlando, Lori A; Orlando, Roy C

    2009-06-01

    Gastroesophageal reflux disease (GERD) is typically heralded by the substernal burning pain of heartburn. On endoscopic examination, about one third of GERD subjects with heartburn have erosive disease, and the remainder have nonerosive reflux disease (NERD). Unlike patients with erosive disease, those with NERD (approximately 50%) often do not respond to therapy with proton pump inhibitors (PPIs), raising the question of whether they have NERD and, if they do, whether the cause of their symptoms is similar to those who respond to PPIs. Recently, biopsies established that subjects with heartburn and PPI-responsive NERD, like those with erosive esophagitis, have lesions within the esophageal epithelium known as dilated intercellular space (DIS). In this article, we discuss the physicochemical basis for DIS in acid-injured esophageal epithelium and its significance in GERD. Although DIS is not pathognomic of GERD, it is a marker of a break in the epithelial (junctional) barrier reflecting an increase in paracellular permeability.

  19. Intercellular communication within the rat anterior pituitary gland: X. Immunohistocytochemistry of S-100 and connexin 43 of folliculo-stellate cells in the rat anterior pituitary gland.

    PubMed

    Shirasawa, Nobuyuki; Mabuchi, Yoshio; Sakuma, Eisuke; Horiuchi, Osamu; Yashiro, Takashi; Kikuchi, Motoshi; Hashimoto, Yasuo; Tsuruo, Yoshihiro; Herbert, Damon C; Soji, Tsuyoshi

    2004-05-01

    Since Rinehart and Farquhar reported the presence of agranulated cells in the anterior pituitary gland in 1953, the functions of the folliculo-stellate cell remain to be clarified. Intercellular junctions have been described in the monkey, rat, and teleost anterior pituitary glands, indicating the existence of cell-to-cell communication within the organ. We pointed to their possible role in the rapid dissemination of information through a complex interconnecting system of follicles involving gap junctions. The gap junctional/folliculo-stellate cellular network was essential in the maturation and regulation of the pituitary gland system such as the hypothalamic-pituitary-gonadal axis. It has been was shown that a network participated in the conduction of electrophysiological information over a long distance using the ion Ca(++), which propagates to other folliculo-stellate cells by signaling through gap junctions. Sixty-day-old male rats were used in this study for light microscopic immunohistochemistry of S-100 protein, type I collagen, and connexin 43, and for electron microscopy to observe the morphological relationships between the cellular networks of folliculo-stellate cells and granulated pituitary cells. Clusters of anti-S-100 protein-positive cells were clearly observed in a region of the hypophysis tentatively named the transition zone. Anti-S-100 protein-positive cells and their cytoplasmic processes were also present in the anterior lobe and assembled together to form follicular lumina. Type I collagen was clearly shown outlining the incomplete lobular or ductule-like structure making cell cords in the anterior pituitary gland. Numerous microvilli were present within the follicular lumen while around the lumina, junctional specializations including gap junctions were positive for the connexin 43 protein. A nonuniform distribution of the connexin 43-positive sites were observed. Small or dot-shaped positive sites were noted where two clusters of cells

  20. Opto-electrical studies of self-assembled monolayer diodes and bulk hetero-junction organic photovoltaic devices

    NASA Astrophysics Data System (ADS)

    Ndobe, Alexandre

    The present dissertation is the result of our studies of the optical and electrical properties of self-assembled monolayer (SAM) diodes and bulk heterojunction organic photovoltaic(BOPV) devices. In our studies of SAM diodes, we fabricated solid-state mixtures of two different kinds of molecules; 1,4 benzene-dimethane-thiol (MeBDT) and 1-pentanethiol (PT). By varying the concentration r of MeBDT with respect to PT, we can go from a regime of isolated molecular wires (10--8 < r 10-3). For r = 0, we found that a potential barrier dominated the transport properties of the device. In the isolated molecules regime, the conductance of MeBDT dominates the transport. In this regime, because of the linearity of the conductance with respect to r, we were able to obtain a "single molecule resistance" at V = 0.1 V of RM = 6x 10--9 . In the aggregated molecules regime, an ohmic response in the current-voltage (I-V) characteristics was observed for bias voltages ≤ 0.5V with the appearance of a new band in the differential conductance around V = 0 along with a new double band in the optical gap at 2.4eV resulting in yellow/red photoluminescence emission. Opto-electrical studies of BOPV devices reveal that there are very few similarities between these types of solar cells and conventional solar cells. From simulations and experiemental measurements of the I-V characteristics, we found that while the open voltage circuit (Voc) is important for engineers, it carries no intrinsic information of the device. It cannot exceed the built-in potential of the device (Vbuilt--in ). The later origin was found to be dependent on electrode work function difference for a non-Ohmic contact configuration and on the active layer's blend in an Ohmic contact configuration. In a bid to improve BOPV device performance, we added to the blend spin 1/2 radical molecules. At concentration ( ≤2%), an increase in device performance was observed. The principal cause for this increase was the increase in

  1. Intercellular Communication in the Adaptive Immune System

    NASA Astrophysics Data System (ADS)

    Chakraborty, Arup

    2004-03-01

    Higher organisms, like humans, have an adaptive immune system that can respond to pathogens that have not been encountered before. T lymphocytes (T cells) are the orchestrators of the adaptive immune response. They interact with cells, called antigen presenting cells (APC), that display molecular signatures of pathogens. Recently, video microscopy experiments have revealed that when T cells detect antigen on APC surfaces, a spatially patterned supramolecular assembly of different types of molecules forms in the junction between cell membranes. This recognition motif is implicated in information transfer between APC and T cells, and so, is labeled the immunological synapse. The observation of synapse formation sparked two broad questions: How does the synapse form? Why does the synapse form? I will describe progress made in answering these fundamental questions in biology by synergistic use of statistical mechanical theory/computation, chemical engineering principles, and genetic and biochemical experiments. The talk will also touch upon mechanisms that may underlie the extreme sensitivity with which T cells discriminate between self and non-self.

  2. Nanotube junctions

    DOEpatents

    Crespi, Vincent Henry; Cohen, Marvin Lou; Louie, Steven Gwon Sheng; Zettl, Alexander Karlwalter

    2003-01-01

    The present invention comprises a new nanoscale metal-semiconductor, semiconductor-semiconductor, or metal-metal junction, designed by introducing topological or chemical defects in the atomic structure of the nanotube. Nanotubes comprising adjacent sections having differing electrical properties are described. These nanotubes can be constructed from combinations of carbon, boron, nitrogen and other elements. The nanotube can be designed having different indices on either side of a junction point in a continuous tube so that the electrical properties on either side of the junction vary in a useful fashion. For example, the inventive nanotube may be electrically conducting on one side of a junction and semiconducting on the other side. An example of a semiconductor-metal junction is a Schottky barrier. Alternatively, the nanotube may exhibit different semiconductor properties on either side of the junction. Nanotubes containing heterojunctions, Schottky barriers, and metal-metal junctions are useful for microcircuitry.

  3. Nanotube junctions

    DOEpatents

    Crespi, Vincent Henry; Cohen, Marvin Lou; Louie, Steven Gwon; Zettl, Alexander Karlwalte

    2004-12-28

    The present invention comprises a new nanoscale metal-semiconductor, semiconductor-semiconductor, or metal-metal junction, designed by introducing topological or chemical defects in the atomic structure of the nanotube. Nanotubes comprising adjacent sections having differing electrical properties are described. These nanotubes can be constructed from combinations of carbon, boron, nitrogen and other elements. The nanotube can be designed having different indices on either side of a junction point in a continuous tube so that the electrical properties on either side of the junction vary in a useful fashion. For example, the inventive nanotube may be electrically conducting on one side of a junction and semiconducting on the other side. An example of a semiconductor-metal junction is a Schottky barrier. Alternatively, the nanotube may exhibit different semiconductor properties on either side of the junction. Nanotubes containing heterojunctions, Schottky barriers, and metal-metal junctions are useful for microcircuitry.

  4. Lens ion homeostasis relies on the assembly and/or stability of large connexin 46 gap junction plaques on the broad sides of differentiating fiber cells

    PubMed Central

    Cheng, Catherine; Nowak, Roberta B.; Gao, Junyuan; Sun, Xiurong; Biswas, Sondip K.; Lo, Woo-Kuen; Mathias, Richard T.

    2015-01-01

    The eye lens consists of layers of tightly packed fiber cells, forming a transparent and avascular organ that is important for focusing light onto the retina. A microcirculation system, facilitated by a network of gap junction channels composed of connexins 46 and 50 (Cx46 and Cx50), is hypothesized to maintain and nourish lens fiber cells. We measured lens impedance in mice lacking tropomodulin 1 (Tmod1, an actin pointed-end capping protein), CP49 (a lens-specific intermediate filament protein), or both Tmod1 and CP49. We were surprised to find that simultaneous loss of Tmod1 and CP49, which disrupts cytoskeletal networks in lens fiber cells, results in increased gap junction coupling resistance, hydrostatic pressure, and sodium concentration. Protein levels of Cx46 and Cx50 in Tmod1−/−;CP49−/− double-knockout (DKO) lenses were unchanged, and electron microscopy revealed normal gap junctions. However, immunostaining and quantitative analysis of three-dimensional confocal images showed that Cx46 gap junction plaques are smaller and more dispersed in DKO differentiating fiber cells. The localization and sizes of Cx50 gap junction plaques in DKO fibers were unaffected, suggesting that Cx46 and Cx50 form homomeric channels. We also demonstrate that gap junction plaques rest in lacunae of the membrane-associated actin-spectrin network, suggesting that disruption of the actin-spectrin network in DKO fibers may interfere with gap junction plaque accretion into micrometer-sized domains or alter the stability of large plaques. This is the first work to reveal that normal gap junction plaque localization and size are associated with normal lens coupling conductance. PMID:25740157

  5. [Intracellular calcium channels, hormone receptors and intercellular calcium waves].

    PubMed

    Tordjmann, T; Tran, D; Berthon, B; Jacquemin, E; Guillon, G; Combettes, L; Claret, M

    1998-01-01

    from permeabilized PP and PV hepatocytes. In permeabilized PP and PV hepatocytes, internal Ca2+ stores displayed the same loading-kinetics, the responses to InsP3 were similar, and the sizes of InsP3-sensitive compartment were not different. In a further study, we investigated by video microscopy in fura2-loaded multicellular systems of rat hepatocytes, the mechanisms controlling intercellular propagation of the Ca2+ wave and coordination of Ca2+ signals induced by the different hormones. Using focal microperfusion which allows local perfusion of any cell of the multiplet, rapid agonist removal during the Ca2+ response and microinjection, we found that second messengers and [Ca2+]i rises in one hepatocyte cannot trigger Ca2+ responses in connected adjacent cells, suggesting that diffusion across gap junctions, while required for coordination, is not sufficient by itself for the propagation of the intercellular Ca2+ wave. In addition, focal microperfusion and intermediate cell disruption experiments revealed very fine functional differences (hormonal delay, frequency of [Ca2+]i oscillations) between hormone-induced Ca2+ signals, even between two adjacent connected hepatocytes. Recent unpublished results performed in suspensions of PP and PV rat hepatocytes supported the view of a major role played by vasopressin receptors (V1a) in genesis and orientation of the Ca2+ wave. Vasopressin binding sites, V1a mRNAs detected by RNAse Protection Assay, and vasopressin-induced InsP3 production, were more abundant in PV than in PP cells. A gradient of hormone receptors could orientate the propagation of the Ca2+ wave in multicellular systems and in liver cell plate. These results suggest that the intercellular Ca2+ wave in multicellular systems of rat hepatocytes is propagated through mechanisms involving at least three factors. (ABSTRACT TRUNCATED)

  6. Relationship between intercellular communication and adriamycin resistance in non-small cell lung cancer.

    PubMed

    Bradley, C; Freshney, R I; Pitts, J

    1994-01-01

    The adriamycin chemosensitivity and extent of gap junctional intercellular communication were assessed in a panel of seven human non-small cell lung cancer (NSCLC) cell lines. Communication was assessed by autoradiographic detection of transfer of 3H uridine nucleotides between coupled cells. The strength of coupling varied widely between the cell lines and they could be separated into 3 groups: those which exhibited strong coupling, L-DAN and A549; those which exhibited weak coupling, SK-MES-1, Calu-3 and NCI-H125; and an intermediate group, WIL and NCI-H23. Adriamycin chemosensitivity was assessed by both clonogenic and MTT assays. The range of IC50 values as measured by either assay was extremely narrow, with no important differences between the lines. Thus, despite the wide spectrum of intercellular communication observed in these lines, this did not correlate with their adriamycin resistance.

  7. 1,4-naphthoquinones: from oxidative damage to cellular and inter-cellular signaling.

    PubMed

    Klotz, Lars-Oliver; Hou, Xiaoqing; Jacob, Claus

    2014-09-17

    Naphthoquinones may cause oxidative stress in exposed cells and, therefore, affect redox signaling. Here, contributions of redox cycling and alkylating properties of quinones (both natural and synthetic, such as plumbagin, juglone, lawsone, menadione, methoxy-naphthoquinones, and others) to cellular and inter-cellular signaling processes are discussed: (i) naphthoquinone-induced Nrf2-dependent modulation of gene expression and its potentially beneficial outcome; (ii) the modulation of receptor tyrosine kinases, such as the epidermal growth factor receptor by naphthoquinones, resulting in altered gap junctional intercellular communication. Generation of reactive oxygen species and modulation of redox signaling are properties of naphthoquinones that render them interesting leads for the development of novel compounds of potential use in various therapeutic settings.

  8. Evidence for an organized lattice in the intercellular space of vestibular sensory cat epithelia.

    PubMed

    Favre, D; Sans, A

    1984-01-01

    The intracellular space between sensory hair cells and nerve endings and between supporting cells of the vestibular epithelia in the cat contains a fine network of microstructures arranged obliquely between the cell membranes of adjacent cells. In situ immersion-perfusion fixation, electron microscopy complemented by lanthanum negative-contrast and freeze-fracture methods suggest that these intercellular structures are tubular with an external diameter between 3 and 6 nm and an internal diameter of 1.5 nm. They could serve as intercytoplasmic tunnels. We discuss the evidence concerning the analogy between the present microchannels and the passageways regularly arranged in the intercellular space of gap-junctions which would permit the transport of low molecular weight substances from one cell to other.

  9. Josephson junction

    DOEpatents

    Wendt, J.R.; Plut, T.A.; Martens, J.S.

    1995-05-02

    A novel method for fabricating nanometer geometry electronic devices is described. Such Josephson junctions can be accurately and reproducibly manufactured employing photolithographic and direct write electron beam lithography techniques in combination with aqueous etchants. In particular, a method is described for manufacturing planar Josephson junctions from high temperature superconducting material. 10 figs.

  10. Josephson junction

    DOEpatents

    Wendt, Joel R.; Plut, Thomas A.; Martens, Jon S.

    1995-01-01

    A novel method for fabricating nanometer geometry electronic devices is described. Such Josephson junctions can be accurately and reproducibly manufactured employing photolithographic and direct write electron beam lithography techniques in combination with aqueous etchants. In particular, a method is described for manufacturing planar Josephson junctions from high temperature superconducting material.

  11. Current trends in salivary gland tight junctions

    PubMed Central

    Baker, Olga J.

    2016-01-01

    ABSTRACT Tight junctions form a continuous intercellular barrier between epithelial cells that is required to separate tissue spaces and regulate selective movement of solutes across the epithelium. They are composed of strands containing integral membrane proteins (e.g., claudins, occludin and tricellulin, junctional adhesion molecules and the coxsackie adenovirus receptor). These proteins are anchored to the cytoskeleton via scaffolding proteins such as ZO-1 and ZO-2. In salivary glands, tight junctions are involved in polarized saliva secretion and barrier maintenance between the extracellular environment and the glandular lumen. This review seeks to provide an overview of what is currently known, as well as the major questions and future research directions, regarding tight junction expression, organization and function within salivary glands. PMID:27583188

  12. Molecular Diffusion through Cyanobacterial Septal Junctions.

    PubMed

    Nieves-Morión, Mercedes; Mullineaux, Conrad W; Flores, Enrique

    2017-01-03

    Heterocyst-forming cyanobacteria grow as filaments in which intercellular molecular exchange takes place. During the differentiation of N2-fixing heterocysts, regulators are transferred between cells. In the diazotrophic filament, vegetative cells that fix CO2 through oxygenic photosynthesis provide the heterocysts with reduced carbon and heterocysts provide the vegetative cells with fixed nitrogen. Intercellular molecular transfer has been traced with fluorescent markers, including calcein, 5-carboxyfluorescein, and the sucrose analogue esculin, which are observed to move down their concentration gradient. In this work, we used fluorescence recovery after photobleaching (FRAP) assays in the model heterocyst-forming cyanobacterium Anabaena sp. strain PCC 7120 to measure the temperature dependence of intercellular transfer of fluorescent markers. We find that the transfer rate constants are directly proportional to the absolute temperature. This indicates that the "septal junctions" (formerly known as "microplasmodesmata") linking the cells in the filament allow molecular exchange by simple diffusion, without any activated intermediate state. This constitutes a novel mechanism for molecular transfer across the bacterial cytoplasmic membrane, in addition to previously characterized mechanisms for active transport and facilitated diffusion. Cyanobacterial septal junctions are functionally analogous to the gap junctions of metazoans.

  13. Patterning of Wound-Induced Intercellular Ca2+ Flashes in a Developing Epithelium

    PubMed Central

    Narciso, Cody; Wu, Qinfeng; Brodskiy, Pavel; Garston, George; Baker, Ruth; Fletcher, Alexander; Zartman, Jeremiah

    2015-01-01

    Differential mechanical force distributions are increasingly recognized to provide important feedback into the control of an organ’s final size and shape. As a second messenger that integrates and relays mechanical information to the cell, calcium ions (Ca2+) are a prime candidate for providing important information on both the overall mechanical state of the tissue and resulting behavior at the individual-cell level during development. Still, how the spatiotemporal properties of Ca2+ transients reflect the underlying mechanical characteristics of tissues is still poorly understood. Here we use an established model system of an epithelial tissue, the Drosophila wing imaginal disc, to investigate how tissue properties impact the propagation of Ca2+ transients induced by laser ablation. The resulting intercellular Ca2+ flash is found to be mediated by inositol 1,4,5-trisphosphate (IP3) and depends on gap junction communication. Further, we find that intercellular Ca2+ transients show spatially nonuniform characteristics across the proximal-distal (PD) axis of the larval wing imaginal disc, which exhibit a gradient in cell size and anisotropy. A computational model of Ca2+ transients is employed to identify the principle factors explaining the spatiotemporal patterning dynamics of intercellular Ca2+ flashes. The relative Ca2+ flash anisotropy is principally explained by local cell shape anisotropy. Further, Ca2+ velocities are relatively uniform throughout the wing disc, irrespective of cell size or anisotropy. This can be explained by the opposing effects of cell diameter and cell elongation on intercellular Ca2+ propagation. Thus, intercellular Ca2+ transients follow lines of mechanical tension at velocities that are largely independent of tissue heterogeneity and reflect the mechanical state of the underlying tissue. PMID:26331891

  14. Patterning of wound-induced intercellular Ca2+ flashes in a developing epithelium

    NASA Astrophysics Data System (ADS)

    Narciso, Cody; Wu, Qinfeng; Brodskiy, Pavel; Garston, George; Baker, Ruth; Fletcher, Alexander; Zartman, Jeremiah

    2015-10-01

    Differential mechanical force distributions are increasingly recognized to provide important feedback into the control of an organ’s final size and shape. As a second messenger that integrates and relays mechanical information to the cell, calcium ions (Ca2+) are a prime candidate for providing important information on both the overall mechanical state of the tissue and resulting behavior at the individual-cell level during development. Still, how the spatiotemporal properties of Ca2+ transients reflect the underlying mechanical characteristics of tissues is still poorly understood. Here we use an established model system of an epithelial tissue, the Drosophila wing imaginal disc, to investigate how tissue properties impact the propagation of Ca2+ transients induced by laser ablation. The resulting intercellular Ca2+ flash is found to be mediated by inositol 1,4,5-trisphosphate and depends on gap junction communication. Further, we find that intercellular Ca2+ transients show spatially non-uniform characteristics across the proximal-distal axis of the larval wing imaginal disc, which exhibit a gradient in cell size and anisotropy. A computational model of Ca2+ transients is employed to identify the principle factors explaining the spatiotemporal patterning dynamics of intercellular Ca2+ flashes. The relative Ca2+ flash anisotropy is principally explained by local cell shape anisotropy. Further, Ca2+ velocities are relatively uniform throughout the wing disc, irrespective of cell size or anisotropy. This can be explained by the opposing effects of cell diameter and cell elongation on intercellular Ca2+ propagation. Thus, intercellular Ca2+ transients follow lines of mechanical tension at velocities that are largely independent of tissue heterogeneity and reflect the mechanical state of the underlying tissue.

  15. Dynamic gap junctional communication: a delimiting model for tissue responses.

    PubMed Central

    Christ, G J; Brink, P R; Ramanan, S V

    1994-01-01

    Gap junctions are aqueous intercellular channels formed by a diverse class of membrane-spanning proteins, known as connexins. These aqueous pores provide partial cytoplasmic continuity between cells in most tissues, and are freely permeable to a host of physiologically relevant second messenger molecules/ionic species (e.g., Ca2+, IP3, cAMP, cGMP). Despite the fact that these second messenger molecules/ionic species have been shown to alter junctional patency, there is no clear basis for understanding how dynamic and transient changes in the intracellular concentration of second messenger molecules might modulate the extent of intercellular communication among coupled cells. Thus, we have modified the tissue monolayer model of Ramanan and Brink (1990) to account for both the up-regulatory and down-regulatory effects on junctions by second messenger molecules that diffuse through gap junctions. We have chosen the vascular wall as our morphological correlate because of its anisotropy and large investment of gap junctions. The model allows us to illustrate the putative behavior of gap junctions under a variety of physiologically relevant conditions. The modeling studies demonstrated that transient alterations in intracellular second messenger concentrations are capable of producing 50-125% changes in the number of cells recruited into a functional syncytial unit, after activation of a single cell. Moreover, the model conditions required to demonstrate such physiologically relevant changes in intercellular diffusion among coupled cells are commonly observed in intact tissues and cultured cells. Images FIGURE 2 PMID:7811948

  16. Development of intercellular communication during the epithelial reorganization of a renal cell line (LLC-PK1).

    PubMed

    Rabito, C A; Jarrell, J A; Abraham, E H

    1987-01-25

    Junctional permeability determinations after microinjection of the fluorescent tracer, Lucifer Yellow CH, show that the cells in confluent monolayers of the renal epithelial cell lines LLC-PK1 and A6 are interconnected by intercellular junctions. This cell-to-cell communication network permits the fluorescent dye to diffuse from the microinjected cell into multiple adjacent neighboring cells. Cell-to-cell diffusion of the fluorescent dye was not observed at pH 6.0. Full recovery occurred, however, when the pH of the extracellular medium was adjusted to 7.4. To provide a sensitive index of the averaged efficacy of junctional communication, we measured the number of cells that survived ouabain treatment in a 50% mixture of wild and ouabain-resistant mutant LLC-PK1 cells. Electron probe microanalysis in uncoupled cells showed that ouabain treatment produced two populations of cells, with totally different intracellular Na+ and K+ content. Under this condition, only 50% of the population survived after 48 h of treatment. When ouabain treatment was initiated 24 h after plating, however, 100% survival was observed, and the cells contained uniform intracellular Na+ and K+ concentration. This finding is consistent with the theory that this protective effect is mediated through the presence of the functional communicating intercellular junctions. When ouabain was applied at different times after plating, full protection is reached by 2 h. The early development of cell-to-cell communication, which precedes the development of the occluding junctions and several transport systems by several hours, is consistent with the involvement of the intercellular junctions in the synchronization of the polarization process.

  17. Characterizing Intercellular Signaling Peptides in Drug Addiction

    PubMed Central

    Romanova, Elena V.; Hatcher, Nathan G.; Rubakhin, Stanislav S.; Sweedler, Jonathan V.

    2009-01-01

    Intercellular signaling peptides (SPs) coordinate the activity of cells and influence organism behavior. SPs, a chemically and structurally diverse group of compounds responsible for transferring information between neurons, are broadly involved in neural plasticity, learning and memory, as well as in drug addiction phenomena. Historically, SP discovery and characterization has tracked advances in measurement capabilities. Today, a suite of analytical technologies is available to investigate individual SPs, as well as entire intercellular signaling complements, in samples ranging from individual cells to entire organisms. Immunochemistry and in situ hybridization are commonly used for following preselected SPs. Discovery-type investigations targeting the transcriptome and proteome are accomplished using high-throughput characterization technologies such as microarrays and mass spectrometry. By integrating directed approaches with discovery approaches, multiplatform studies fill critical gaps in our knowledge of drug-induced alterations in intercellular signaling. Throughout the past 35 years, the National Institute on Drug Abuse has made significant resources available to scientists that study the mechanisms of drug addiction. The roles of SPs in the addiction process are highlighted, as are the analytical approaches used to detect and characterize them. PMID:18722391

  18. Transient inter-cellular polymeric linker.

    PubMed

    Ong, Siew-Min; He, Lijuan; Thuy Linh, Nguyen Thi; Tee, Yee-Han; Arooz, Talha; Tang, Guping; Tan, Choon-Hong; Yu, Hanry

    2007-09-01

    Three-dimensional (3D) tissue-engineered constructs with bio-mimicry cell-cell and cell-matrix interactions are useful in regenerative medicine. In cell-dense and matrix-poor tissues of the internal organs, cells support one another via cell-cell interactions, supplemented by small amount of the extra-cellular matrices (ECM) secreted by the cells. Here we connect HepG2 cells directly but transiently with inter-cellular polymeric linker to facilitate cell-cell interaction and aggregation. The linker consists of a non-toxic low molecular-weight polyethyleneimine (PEI) backbone conjugated with multiple hydrazide groups that can aggregate cells within 30 min by reacting with the aldehyde handles on the chemically modified cell-surface glycoproteins. The cells in the cellular aggregates proliferated; and maintained the cortical actin distribution of the 3D cell morphology while non-aggregated cells died over 7 days of suspension culture. The aggregates lost distinguishable cell-cell boundaries within 3 days; and the ECM fibers became visible around cells from day 3 onwards while the inter-cellular polymeric linker disappeared from the cell surfaces over time. The transient inter-cellular polymeric linker can be useful for forming 3D cellular and tissue constructs without bulk biomaterials or extensive network of engineered ECM for various applications.

  19. Mechanically Induced Intercellular Calcium Communication in Confined Endothelial Structures

    PubMed Central

    Junkin, Michael; Lu, Yi; Long, Juexuan; Deymier, Pierre A.; Hoying, James B.; Wong, Pak Kin

    2012-01-01

    Calcium signaling in the diverse vascular structures is regulated by a wide range of mechanical and biochemical factors to maintain essential physiological functions of the vasculature. To properly transmit information, the intercellular calcium communication mechanism must be robust against various conditions in the cellular microenvironment. Using plasma lithography geometric confinement, we investigate mechanically induced calcium wave propagation in networks of human umbilical vein endothelial cells organized. Endothelial cell networks with confined architectures were stimulated at the single cell level, including using capacitive force probes. Calcium wave propagation in the network was observed using fluorescence calcium imaging. We show that mechanically induced calcium signaling in the endothelial networks is dynamically regulated against a wide range of probing forces and repeated stimulations. The calcium wave is able to propagate consistently in various dimensions from monolayers to individual cell chains, and in different topologies from linear patterns to cell junctions. Our results reveal that calcium signaling provides a robust mechanism for cell-cell communication in networks of endothelial cells despite the diversity of the microenvironmental inputs and complexity of vascular structures. PMID:23267827

  20. Mitotic cells form actin-based bridges with adjacent cells to provide intercellular communication during rounding.

    PubMed

    Fykerud, Tone A; Knudsen, Lars M; Totland, Max Z; Sørensen, Vigdis; Dahal-Koirala, Shiva; Lothe, Ragnhild A; Brech, Andreas; Leithe, Edward

    2016-11-01

    In order to achieve accurate chromosome segregation, eukaryotic cells undergo a dramatic change in morphology to obtain a spherical shape during mitosis. Interphase cells communicate directly with each other by exchanging ions and small molecules via gap junctions, which have important roles in controlling cell growth and differentiation. As cells round up during mitosis, the gap junctional communication between mitotic cells and adjacent interphase cells ceases. Whether mitotic cells use alternative mechanisms for mediating direct cell-cell communication during rounding is currently unknown. Here, we have studied the mechanisms involved in the remodeling of gap junctions during mitosis. We further demonstrate that mitotic cells are able to form actin-based plasma membrane bridges with adjacent cells during rounding. These structures, termed "mitotic nanotubes," were found to be involved in mediating the transport of cytoplasm, including Rab11-positive vesicles, between mitotic cells and adjacent cells. Moreover, a subpool of the gap-junction channel protein connexin43 localized in these intercellular bridges during mitosis. Collectively, the data provide new insights into the mechanisms involved in the remodeling of gap junctions during mitosis and identify actin-based plasma membrane bridges as a novel means of communication between mitotic cells and adjacent cells during rounding.

  1. Mitotic cells form actin-based bridges with adjacent cells to provide intercellular communication during rounding

    PubMed Central

    Fykerud, Tone A.; Knudsen, Lars M.; Totland, Max Z.; Dahal-Koirala, Shiva; Lothe, Ragnhild A.; Brech, Andreas; Leithe, Edward

    2016-01-01

    ABSTRACT In order to achieve accurate chromosome segregation, eukaryotic cells undergo a dramatic change in morphology to obtain a spherical shape during mitosis. Interphase cells communicate directly with each other by exchanging ions and small molecules via gap junctions, which have important roles in controlling cell growth and differentiation. As cells round up during mitosis, the gap junctional communication between mitotic cells and adjacent interphase cells ceases. Whether mitotic cells use alternative mechanisms for mediating direct cell-cell communication during rounding is currently unknown. Here, we have studied the mechanisms involved in the remodeling of gap junctions during mitosis. We further demonstrate that mitotic cells are able to form actin-based plasma membrane bridges with adjacent cells during rounding. These structures, termed “mitotic nanotubes,” were found to be involved in mediating the transport of cytoplasm, including Rab11-positive vesicles, between mitotic cells and adjacent cells. Moreover, a subpool of the gap-junction channel protein connexin43 localized in these intercellular bridges during mitosis. Collectively, the data provide new insights into the mechanisms involved in the remodeling of gap junctions during mitosis and identify actin-based plasma membrane bridges as a novel means of communication between mitotic cells and adjacent cells during rounding. PMID:27625181

  2. Target-Catalyzed DNA Four-Way Junctions for CRET Imaging of MicroRNA, Concatenated Logic Operations, and Self-Assembly of DNA Nanohydrogels for Targeted Drug Delivery.

    PubMed

    Bi, Sai; Xiu, Bao; Ye, Jiayan; Dong, Ying

    2015-10-21

    Here we report a target-catalyzed DNA four-way junction (DNA-4WJ) on the basis of toehold-mediated DNA strand displacement reaction (TM-SDR), which is readily applied in enzyme-free amplified chemiluminescence resonance energy transfer (CRET) imaging of microRNA. In this system, the introduction of target microRNA-let-7a (miR-let-7a) activates a cascade of assembly steps with four DNA hairpins, followed by a disassembly step in which the target microRNA is displaced and released from DNA-4WJ to catalyze the self-assembly of additional branched junctions. As a result, G-quadruplex subunit sequences and fluorophore fluorescein amidite (FAM) are encoded in DNA-4WJ in a close proximity, stimulating a CRET process in the presence of hemin/K(+) to form horseradish peroxidase (HRP)-mimicking DNAzyme that catalyzes the generation of luminol/H2O2 chemiluminescence (CL), which further transfers to FAM. The background signal is easily reduced using magnetic graphene oxide (MGO) to remove unreacted species through magnetic separation, which makes a great contribution to improve the detection sensitivity and achieves a detection limit as low as 6.9 fM microRNA-let-7a (miR-let-7a). In addition, four-input concatenated logic circuits with an automatic reset function have been successfully constructed relying on the architecture of the proposed DNA-4WJ. More importantly, DNA nanohydrogels are self-assembled using DNA-4WJs as building units after centrifugation, which are driven by liquid crystallization and dense packaging of building units. Moreover, the DNA nanohydrogels are readily functionalized by incorporating with aptamers, bioimaging agents, and drug loading sites, which thus are served as efficient nanocarriers for targeted drug delivery and cancer therapy with high loading capacity and excellent biocompatibility.

  3. Spatial organization of the extracellular matrix regulates cell–cell junction positioning

    PubMed Central

    Tseng, Qingzong; Duchemin-Pelletier, Eve; Deshiere, Alexandre; Balland, Martial; Guillou, Hervé; Filhol, Odile; Théry, Manuel

    2012-01-01

    The organization of cells into epithelium depends on cell interaction with both the extracellular matrix (ECM) and adjacent cells. The role of cell–cell adhesion in the regulation of epithelial topology is well-described. ECM is better known to promote cell migration and provide a structural scaffold for cell anchoring, but its contribution to multicellular morphogenesis is less well-understood. We developed a minimal model system to investigate how ECM affects the spatial organization of intercellular junctions. Fibronectin micropatterns were used to constrain the location of cell–ECM adhesion. We found that ECM affects the degree of stability of intercellular junction positioning and the magnitude of intra- and intercellular forces. Intercellular junctions were permanently displaced, and experienced large perpendicular tensional forces as long as they were positioned close to ECM. They remained stable solely in regions deprived of ECM, where they were submitted to lower tensional forces. The heterogeneity of the spatial organization of ECM induced anisotropic distribution of mechanical constraints in cells, which seemed to adapt their position to minimize both intra- and intercellular forces. These results uncover a morphogenetic role for ECM in the mechanical regulation of cells and intercellular junction positioning. PMID:22307605

  4. Structure, regulation and function of gap junctions in liver

    PubMed Central

    Maes, Michaël; Decrock, Elke; Wang, Nan; Leybaert, Luc; da Silva, Tereza Cristina; Veloso Alves Pereira, Isabel; Jaeschke, Hartmut; Cogliati, Bruno; Vinken, Mathieu

    2016-01-01

    Gap junctions are a specialized group of cell-to-cell junctions that mediate direct intercellular communication between cells. They arise from the interaction of 2 hemichannels of adjacent cells, which in turn are composed of 6 connexin proteins. In liver, gap junctions are predominantly found in hepatocytes and play critical roles in virtually all phases of the hepatic life cycle, including cell growth, differentiation, liver-specific functionality and cell death. Liver gap junctions are directed through a broad variety of mechanisms ranging from epigenetic control of connexin expression to posttranslational regulation of gap junction activity. This paper reviews established and novel aspects regarding the architecture, control and functional relevance of liver gap junctions. PMID:27001459

  5. Gap junctional communication during limb cartilage differentiation.

    PubMed

    Coelho, C N; Kosher, R A

    1991-03-01

    The onset of cartilage differentiation in the developing limb bud is characterized by a transient cellular condensation process in which prechondrogenic mesenchymal cells become closely apposed to one another prior to initiating cartilage matrix deposition. During this condensation process intimate cell-cell interactions occur which are necessary to trigger chondrogenic differentiation. In the present study, we demonstrate that extensive cell-cell communication via gap junctions as assayed by the intercellular transfer of lucifer yellow dye occurs during condensation and the onset of overt chondrogenesis in high density micromass cultures prepared from the homogeneous population of chondrogenic precursor cells comprising the distal subridge region of stage 25 embryonic chick wing buds. Furthermore, in heterogeneous micromass cultures prepared from the mesodermal cells of whole stage 23/24 limb buds, extensive gap junctional communication is limited to differentiating cartilage cells, while the nonchondrogenic cells of the cultures that are differentiating into the connective tissue lineage exhibit little or no intercellular communication via gap junctions. These results provide a strong incentive for considering and further investigating the possible involvement of cell-cell communication via gap junctions in the regulation of limb cartilage differentiation.

  6. Parametric analysis of intercellular ice propagation during cryosurgery, simulated using monte carlo techniques.

    PubMed

    Stott, Shannon L; Irimia, Daniel; Karlsson, Jens O M

    2004-04-01

    A microscale theoretical model of intracellular ice formation (IIF) in a heterogeneous tissue volume comprising a tumor mass and surrounding normal tissue is presented. Intracellular ice was assumed to form either by intercellular ice propagation or by processes that are not affected by the presence of ice in neighboring cells (e.g., nucleation or mechanical rupture). The effects of cryosurgery on a 2D tissue consisting of 10(4) cells were simulated using a lattice Monte Carlo technique. A parametric analysis was performed to assess the specificity of IIF-related cell damage and to identify criteria for minimization of collateral damage to the healthy tissue peripheral to the tumor. Among the parameters investigated were the rates of interaction-independent IIF and intercellular ice propagation in the tumor and in the normal tissue, as well as the characteristic length scale of thermal gradients in the vicinity of the cryosurgical probe. Model predictions suggest gap junctional intercellular communication as a potential new target for adjuvant therapies complementing the cryosurgical procedure.

  7. Molecular Diffusion through Cyanobacterial Septal Junctions

    PubMed Central

    Nieves-Morión, Mercedes

    2017-01-01

    ABSTRACT Heterocyst-forming cyanobacteria grow as filaments in which intercellular molecular exchange takes place. During the differentiation of N2-fixing heterocysts, regulators are transferred between cells. In the diazotrophic filament, vegetative cells that fix CO2 through oxygenic photosynthesis provide the heterocysts with reduced carbon and heterocysts provide the vegetative cells with fixed nitrogen. Intercellular molecular transfer has been traced with fluorescent markers, including calcein, 5-carboxyfluorescein, and the sucrose analogue esculin, which are observed to move down their concentration gradient. In this work, we used fluorescence recovery after photobleaching (FRAP) assays in the model heterocyst-forming cyanobacterium Anabaena sp. strain PCC 7120 to measure the temperature dependence of intercellular transfer of fluorescent markers. We find that the transfer rate constants are directly proportional to the absolute temperature. This indicates that the “septal junctions” (formerly known as “microplasmodesmata”) linking the cells in the filament allow molecular exchange by simple diffusion, without any activated intermediate state. This constitutes a novel mechanism for molecular transfer across the bacterial cytoplasmic membrane, in addition to previously characterized mechanisms for active transport and facilitated diffusion. Cyanobacterial septal junctions are functionally analogous to the gap junctions of metazoans. PMID:28049144

  8. The calcium ATPase SERCA2 regulates desmoplakin dynamics and intercellular adhesive strength through modulation of PKCα signaling

    PubMed Central

    Hobbs, Ryan P.; Amargo, Evangeline V.; Somasundaram, Agila; Simpson, Cory L.; Prakriya, Murali; Denning, Mitchell F.; Green, Kathleen J.

    2011-01-01

    Darier's disease (DD) is an inherited autosomal-dominant skin disorder characterized histologically by loss of adhesion between keratinocytes. DD is typically caused by mutations in sarcoendoplasmic reticulum Ca2+-ATPase isoform 2 (SERCA2), a major regulator of intracellular Ca2+ homeostasis in the skin. However, a defined role for SERCA2 in regulating intercellular adhesion remains poorly understood. We found that diminution of SERCA2 function by pharmacological inhibition or siRNA silencing in multiple human epidermal-derived cell lines was sufficient to disrupt desmosome assembly and weaken intercellular adhesive strength. Specifically, SERCA2-deficient cells exhibited up to a 60% reduction in border translocation of desmoplakin (DP), the desmosomal cytolinker protein necessary for intermediate filament (IF) anchorage to sites of robust cell-cell adhesion. In addition, loss of SERCA2 impaired the membrane translocation of protein kinase C α (PKCα), a known regulator of DP-IF association and desmosome assembly, to the plasma membrane by up to 70%. Exogenous activation of PKCα in SERCA2-deficient cells was sufficient to rescue the defective DP localization, desmosome assembly, and intercellular adhesive strength to levels comparable to controls. Our findings indicate that SERCA2-deficiency is sufficient to impede desmosome assembly and weaken intercellular adhesive strength via a PKCα-dependent mechanism, implicating SERCA2 as a novel regulator of PKCα signaling.—Hobbs, R. P., Amargo, E. V., Somasundaram, A., Simpson, C. L., Prakriya, M., Denning, M. F., Green, K. J. The calcium ATPase SERCA2 regulates desmoplakin dynamics and intercellular adhesive strength through modulation of PKCα signaling. PMID:21156808

  9. Intercellular Communication—Filling in the Gaps

    PubMed Central

    Meiners, Sally; Baron-Epel, Orna; Schindler, Melvin

    1988-01-01

    Coordination and synchrony of a variety of cellular activities in tissues of plants and animals occur as a consequence of the transfer of low molecular weight biosynthetic and signaling molecules through specialized structures (plasmodesmata in plant cells and gap junctions in mammalian cells) that form aqueous channels between contacting cells. Investigations with rat liver demonstrated that cell-cell communication is mediated by a 32 kilodalton polypeptide that forms a hexameric pore structure in the plasma membrane. Following association with the same structure in a contiguous cell, a trans-double membrane channel is created that has been termed a gap junction. In plant tissue, long tubelike structures called plasmodesmata are suggested to serve a similar cell-cell linking function between cytoplasmic compartments. Although morphologically distinct, dynamic observations suggest similarities in transport properties between gap junctions and plasmodesmata. Recent work now provides evidence that these functional similarities may reflect a more profound identity between the paradigm animal gap junction polypeptide (32 kilodalton rat liver polypeptide) and an immunologically homologous protein localized to plant plasma membrane/cell wall fractions that may be a component of plasmodesmata. PMID:16666225

  10. On the structural organization of isolated bovine lens fiber junctions.

    PubMed

    Zampighi, G; Simon, S A; Robertson, J D; McIntosh, T J; Costello, M J

    1982-04-01

    Junctions between fiber cells of bovine lenses have been isolated in milligram quantities, without using detergents or proteases. The structure of the isolated junctions has been studied by thin-section, negative-stain, and freeze-fracture electron microscopy and by x-ray diffraction. The junctions are large and most often have an undulating surface topology as determined by thin sectioning and freeze-fracture. These undulations resemble the tongue-and-groove interdigitations between lens fiber cells previously seen by others (D. H. Dickson and G. W. Crock, 1972, Invest. Ophthalmol. 11:809-815). In sections, the isolated junctions display a pentalamellar structure approximately 13-14 nm in overall thickness, which is significantly thinner than liver gap junctions. Each junctional membrane contains in the plane of the lipid bilayers distinct units arranged in a square lattice with a center-to-center spacing of 6.6 nm. Freeze-fracture replicas of the junctions fractured transversely show that the repeating units extend across the entire thickness of each membrane. Each unit is probably constructed from four identical subunits, with each subunit containing a protein of an apparent molecular weight of 27,000. We conclude that the lens junctions are structurally and chemically, different from gap junctions and could represent a new kind of intercellular contact, not simply another crystalline state of the gap junction protein.

  11. "Intercellular bridges" in a case of well differentiated squamous carcinoma.

    PubMed

    Nguyen, Michaela; Mikita, Geoffrey; Hoda, Rana S

    2016-02-01

    Intercellular bridges may aide in definitive identification of malignant cell origin, especially in squamous cell carcinoma. They are difficult to identify in routine cytologic specimens and are especially rare in smear preparations. Herein, we present images of intercellular bridges from a case of well differentiated squamous cell carcinoma of the esophagus in a cytologic specimen obtained from FNA of a paraesophageal lymph node.

  12. Establishment of cell-cell junctions depends on the oligomeric states of VE-cadherin

    PubMed Central

    Bibert, Stéphanie; Ayari, Hélène; Riveline, Daniel; Concord, Evelyne; Hermant, Bastien; Vernet, Thierry; Gulino-Debrac, Danièle

    2008-01-01

    Specifically expressed at intercellular adherens junctions of endothelial cells, VE-cadherin is a receptor that exhibits particular self-association properties. Indeed, in vitro studies demonstrated that the extracellular part of VE-cadherin elaborates Ca++-dependent hexameric structures. We hypothesized that this assembly could be at the basis of a new cadherin-mediated cell-cell adhesion mechanism. To verify this assumption, we first demonstrated that VE-cadherin can elaborate hexamers at the cell surface of confluent endothelial cells. Second, mutations were introduced within the extracellular part of VE-cadherin to destabilize the hexamer. Following an in vitro screening, three mutants were selected, among which, one is able to elaborate only dimers. The selected mutations were expressed as C-terminal Green Fluorescent Protein fusions in CHO cells. Despite their capacity to elaborate nascent cell-cell contacts, the mutants seem to be rapidly degraded and or internalized. Altogether, our results suggest that the formation of VE-cadherin hexamers protects this receptor and might allow the elaboration of mature endothelial cell-cell junctions. PMID:18343874

  13. STUDIES ON AN EPITHELIAL (GLAND) CELL JUNCTION

    PubMed Central

    Loewenstein, Werner R.; Kanno, Yoshinobu

    1964-01-01

    Membrane permeability of an epithelial cell junction (Drosophila salivary gland) was examined with intracellular microelectrodes and with fluorescent tracers. In contrast to the non-junctional cell membrane surface, which has a low permeability to ions (10-4 mho/cm2), the junctional membrane surface is highly permeable. In fact, it introduces no substantial restriction to ion flow beyond that in the cytoplasm; the resistance through a chain of cells (150 Ω cm) is only slightly greater than in extruded cytoplasm (100 Ω cm). The diffusion resistance along the intercellular space to the exterior, on the other hand, is very high. Here, there exists an ion barrier of, at least, 104Ω cm2. As a result, small ions and fluorescein move rather freely from one cell to the next, but do not leak appreciably through the intercellular space to the exterior. The organ here, rather than the single cell, appears to be the unit of ion environment. The possible underlying structural aspects are discussed. PMID:14206423

  14. Assembling carbon fiber–graphene–carbon fiber hetero-structures into 1D–2D–1D junction fillers and patterned structures for improved microwave absorption

    NASA Astrophysics Data System (ADS)

    Li, Huimin; Liu, Lin; Li, Hai-Bing; Song, Wei-Li; Bian, Xing-Ming; Zhao, Quan-Liang; Chen, Mingji; Yuan, Xujin; Chen, Haosen; Fang, Daining

    2017-04-01

    Since carbon-based structures of various dimensions, including one-dimensional (1D) carbon nanotubes, two-dimensional (2D) graphene and three-dimensional (3D) carbon foams, have attracted significant attention as microwave absorption fillers, we present an exceptional hetero-junction filler with a 1D–2D–1D feature, achieved by manipulating 2D graphene into 1D carbon fibers in the fiber-extruding process under the electric field. The as-fabricated 1D–2D–1D structural fillers exhibited much-improved dielectric properties and promoted microwave absorption performance in their composites, which is linked to the establishment of enhanced polarization capability, the generation of increased electric loss pathway and the creation of more favorable electromagnetic energy consumption conditions. The results suggest that employing 2D graphene in the 1D–2D–1D nanostructures played the critical role in tuning the electromagnetic response ability, because of its intrinsic electric advantages and dimensional features. To broaden the effective absorption bandwidth, periodic pattern-absorbing structures were designed, which showed combined absorption advantages for various thicknesses. Our strategy for fabricating 1D–2D–1D structural fillers illuminates a universal approach for manipulating dimensions and structures in the nanotechnology.

  15. Drosophila wing imaginal discs respond to mechanical injury via slow InsP3R-mediated intercellular calcium waves

    NASA Astrophysics Data System (ADS)

    Restrepo, Simon; Basler, Konrad

    2016-08-01

    Calcium signalling is a highly versatile cellular communication system that modulates basic functions such as cell contractility, essential steps of animal development such as fertilization and higher-order processes such as memory. We probed the function of calcium signalling in Drosophila wing imaginal discs through a combination of ex vivo and in vivo imaging and genetic analysis. Here we discover that wing discs display slow, long-range intercellular calcium waves (ICWs) when mechanically stressed in vivo or cultured ex vivo. These slow imaginal disc intercellular calcium waves (SIDICs) are mediated by the inositol-3-phosphate receptor, the endoplasmic reticulum (ER) calcium pump SERCA and the key gap junction component Inx2. The knockdown of genes required for SIDIC formation and propagation negatively affects wing disc recovery after mechanical injury. Our results reveal a role for ICWs in wing disc homoeostasis and highlight the utility of the wing disc as a model for calcium signalling studies.

  16. Drosophila wing imaginal discs respond to mechanical injury via slow InsP3R-mediated intercellular calcium waves

    PubMed Central

    Restrepo, Simon; Basler, Konrad

    2016-01-01

    Calcium signalling is a highly versatile cellular communication system that modulates basic functions such as cell contractility, essential steps of animal development such as fertilization and higher-order processes such as memory. We probed the function of calcium signalling in Drosophila wing imaginal discs through a combination of ex vivo and in vivo imaging and genetic analysis. Here we discover that wing discs display slow, long-range intercellular calcium waves (ICWs) when mechanically stressed in vivo or cultured ex vivo. These slow imaginal disc intercellular calcium waves (SIDICs) are mediated by the inositol-3-phosphate receptor, the endoplasmic reticulum (ER) calcium pump SERCA and the key gap junction component Inx2. The knockdown of genes required for SIDIC formation and propagation negatively affects wing disc recovery after mechanical injury. Our results reveal a role for ICWs in wing disc homoeostasis and highlight the utility of the wing disc as a model for calcium signalling studies. PMID:27503836

  17. Multisite electrophysiological recordings by self-assembled loose-patch-like junctions between cultured hippocampal neurons and mushroom-shaped microelectrodes

    PubMed Central

    Shmoel, Nava; Rabieh, Noha; Ojovan, Silviya M.; Erez, Hadas; Maydan, Eilon; Spira, Micha E.

    2016-01-01

    Substrate integrated planar microelectrode arrays is the “gold standard” method for millisecond-resolution, long-term, large-scale, cell-noninvasive electrophysiological recordings from mammalian neuronal networks. Nevertheless, these devices suffer from drawbacks that are solved by spike-detecting, spike-sorting and signal-averaging techniques which rely on estimated parameters that require user supervision to correct errors, merge clusters and remove outliers. Here we show that primary rat hippocampal neurons grown on micrometer sized gold mushroom-shaped microelectrodes (gMμE) functionalized simply by poly-ethylene-imine/laminin undergo self-assembly processes to form loose patch-like hybrid structures. More than 90% of the hybrids formed in this way record monophasic positive action potentials (APs). Of these, 34.5% record APs with amplitudes above 300 μV and up to 5,085 μV. This self-assembled neuron-gMμE configuration improves the recording quality as compared to planar MEA. This study characterizes and analyzes the electrophysiological signaling repertoire generated by the neurons-gMμE configuration, and discusses prospects to further improve the technology. PMID:27256971

  18. Intercellular communication lessons in heart failure.

    PubMed

    Bang, Claudia; Antoniades, Charalambos; Antonopoulos, Alexios S; Eriksson, Ulf; Franssen, Constantijn; Hamdani, Nazha; Lehmann, Lorenz; Moessinger, Christine; Mongillo, Marco; Muhl, Lars; Speer, Thimoteus; Thum, Thomas

    2015-11-01

    Cell-cell or inter-organ communication allows the exchange of information and messages, which is essential for the coordination of cell/organ functions and the maintenance of homeostasis. It has become evident that dynamic interactions of different cell types play a major role in the heart, in particular during the progression of heart failure, a leading cause of mortality worldwide. Heart failure is associated with compensatory structural and functional changes mostly in cardiomyocytes and cardiac fibroblasts, which finally lead to cardiomyocyte hypertrophy and fibrosis. Intercellular communication within the heart is mediated mostly via direct cell-cell interaction or the release of paracrine signalling mediators such as cytokines and chemokines. However, recent studies have focused on the exchange of genetic information via the packaging into vesicles as well as the crosstalk of lipids and other paracrine molecules within the heart and distant organs, such as kidney and adipose tissue, which might all contribute to the pathogenesis of heart failure. In this review, we discuss emerging communication networks and respective underlying mechanisms which could be involved in cardiovascular disease conditions and further emphasize promising therapeutic targets for drug development.

  19. Senescent cells communicate via intercellular protein transfer

    PubMed Central

    Biran, Anat; Perelmutter, Meirav; Gal, Hilah; Burton, Dominick G.A.; Ovadya, Yossi; Vadai, Ezra; Geiger, Tamar

    2015-01-01

    Mammalian cells mostly rely on extracellular molecules to transfer signals to other cells. However, in stress conditions, more robust mechanisms might be necessary to facilitate cell–cell communications. Cellular senescence, a stress response associated with permanent exit from the cell cycle and the development of an immunogenic phenotype, limits both tumorigenesis and tissue damage. Paradoxically, the long-term presence of senescent cells can promote tissue damage and aging within their microenvironment. Soluble factors secreted from senescent cells mediate some of these cell-nonautonomous effects. However, it is unknown whether senescent cells impact neighboring cells by other mechanisms. Here we show that senescent cells directly transfer proteins to neighboring cells and that this process facilitates immune surveillance of senescent cells by natural killer (NK) cells. We found that transfer of proteins to NK and T cells is increased in the murine preneoplastic pancreas, a site where senescent cells are present in vivo. Proteomic analysis and functional studies of the transferred proteins revealed that the transfer is strictly dependent on cell–cell contact and CDC42-regulated actin polymerization and is mediated at least partially by cytoplasmic bridges. These findings reveal a novel mode of intercellular communication by which senescent cells regulate their immune surveillance and might impact tumorigenesis and tissue aging. PMID:25854920

  20. Expression of claudins, occludin, junction adhesion molecule A and zona occludens 1 in canine organs

    PubMed Central

    Ahn, Changhwan; Shin, Da-Hye; Lee, Dongoh; Kang, Su-Myung; Seok, Ju-Hyung; Kang, Hee Young; Jeung, Eui-Bae

    2016-01-01

    Tight junctions are the outermost structures of intercellular junctions and are classified as transmembrane proteins. These factors form selective permeability barriers between cells, act as paracellular transporters and regulate structural and functional polarity of cells. Although tight junctions have been previously studied, comparison of the transcriptional-translational levels of these molecules in canine organs remains to be investigated. In the present study, organ-specific expression of the tight junction proteins, claudin, occludin, junction adhesion molecule A and zona occludens 1 was examined in the canine duodenum, lung, liver and kidney. Results of immunohistochemistry analysis demonstrated that the tight junctions were localized in intestinal villi and glands of the duodenum, bronchiolar epithelia and alveolar walls of the lung, endometrium and myometrium of the hepatocytes, and the distal tubules and glomeruli of the kidney. These results suggest that tight junctions are differently expressed in organs, and therefore may be involved in organ-specific functions to maintain physiological homeostasis. PMID:27600198

  1. Molecular mechanisms of intercellular communication: transmembrane signaling

    SciTech Connect

    Bitensky, M.W.; George, J.S.; Siegel, H.N.; McGregor, D.M.

    1982-01-01

    This short discussion of transmembrane signaling depicts a particular class of signaling devices whose functional characteristics may well be representative of broader classes of membrane switches. These multicomponent aggregates are characterized by tight organization of interacting components which function by conformational interactions to provide sensitive, amplified, rapid, and modulated responses. It is clear that the essential role of such switches in cell-cell interactions necessitated their appearance early in the history of the development of multicellular organisms. It also seems clear that once such devices made their appearance, the conformationally interactive moieties were firmly locked into a regulatory relationship. Since modification of interacting components could perturb or interfere with the functional integrity of the whole switch, genetic drift was only permitted at the input and outflow extremes. However, the GTP binding moiety and its interacting protein domains on contiguous portions of the receptor and readout components were highly conserved. The observed stringent evolutionary conservation of the molecular features of these membrane switches thus applies primarily to the central (GTP binding) elements. An extraordinary degree of variation was permitted within the domains of signal recognition and enzymatic output. Thus, time and evolution have adapted the central logic of the regulatory algorithm to serve a great variety of cellular purposes and to recognize a great variety of chemical and physical signals. This is exemplified by the richness of the hormonal and cellular dialogues found in primates such as man. Here the wealth of intercellular communiation can support the composition and performance of symphonies and the study of cellular immunology.

  2. Mitochondria Know No Boundaries: Mechanisms and Functions of Intercellular Mitochondrial Transfer

    PubMed Central

    Torralba, Daniel; Baixauli, Francesc; Sánchez-Madrid, Francisco

    2016-01-01

    Mitochondria regulate multiple cell processes, including calcium signaling, apoptosis and cell metabolism. Mitochondria contain their own circular genome encoding selected subunits of the oxidative phosphorylation complexes. Recent findings reveal that, in addition to being maternally inherited, mitochondria can traverse cell boundaries and thus be horizontally transferred between cells. Although, the physiological relevance of this phenomenon is still under debate, mitochondria uptake rescues mitochondrial respiration defects in recipient cells and regulates signaling, proliferation or chemotherapy resistance in vitro and in vivo. In this review, we outline the pathophysiological consequences of horizontal mitochondrial transfer and offer a perspective on the cellular and molecular mechanisms mediating their intercellular transmission, including tunneling nanotubes, extracellular vesicles, cellular fusion, and GAP junctions. The physiological relevance of mitochondrial transfer and the potential therapeutic application of this exchange for treating mitochondrial-related diseases are discussed. PMID:27734015

  3. Intercellular Ca(2+) wave propagation in human retinal pigment epithelium cells induced by mechanical stimulation.

    PubMed

    Abu Khamidakh, A E; Juuti-Uusitalo, K; Larsson, K; Skottman, H; Hyttinen, J

    2013-03-01

    Ca(2+) signaling is vitally important in cellular physiological processes and various drugs also affect Ca(2+) signaling. Thus, knowledge of Ca(2+) dynamics is important toward understanding cell biology, as well as the development of drug-testing assays. ARPE-19 cells are widely used for modeling human retinal pigment epithelium functions and drug-testing, but intercellular communication has not been assessed in these cells. In this study, we investigated intercellular Ca(2+) communication induced by mechanical stimulation in ARPE-19 cells. An intercellular Ca(2+) wave was induced in ARPE-19 monolayer by point mechanical stimulation of a single cell. Dynamic changes of intracellular Ca(2+) concentration ([Ca(2+)](i)) in the monolayer were tracked with fluorescence microscopy imaging using Ca(2+)-sensitive fluorescent dye fura-2 in presence and absence of extracellular Ca(2+), after depletion of intracellular Ca(2+) stores with thapsigargin, and after application of gap junction blocker α-glycyrrhetinic acid and P2-receptor blocker suramin. Normalized fluorescence values, reflecting amplitude of [Ca(2+)](i) increase, and percentage of responsive cells were calculated to quantitatively characterize Ca(2+) wave propagation. Mechanical stimulation of a single cell within a confluent monolayer of ARPE-19 cells initiated an increase in [Ca(2+)](i), which propagated to neighboring cells in a wave-like manner. Ca(2+) wave propagated to up to 14 cell tiers in control conditions. The absence of extracellular Ca(2+) reduced [Ca(2+)](i) increase in the cells close to the site of mechanical stimulation, whereas the depletion of intracellular Ca(2+) stores with thapsigargin blocked the wave spreading to distant cells. The gap junction blocker α-glycyrrhetinic acid reduced [Ca(2+)](i) increase in the cell tiers close to the site of mechanical stimulation, indicating involvement of gap junctions in Ca(2+) wave propagation. The P2-receptor blocker suramin reduced the percentage

  4. Rickettsia Sca4 Reduces Vinculin-Mediated Intercellular Tension to Promote Spread.

    PubMed

    Lamason, Rebecca L; Bastounis, Effie; Kafai, Natasha M; Serrano, Ricardo; Del Álamo, Juan C; Theriot, Julie A; Welch, Matthew D

    2016-10-20

    Spotted fever group (SFG) rickettsiae are human pathogens that infect cells in the vasculature. They disseminate through host tissues by a process of cell-to-cell spread that involves protrusion formation, engulfment, and vacuolar escape. Other bacterial pathogens rely on actin-based motility to provide a physical force for spread. Here, we show that SFG species Rickettsia parkeri typically lack actin tails during spread and instead manipulate host intercellular tension and mechanotransduction to promote spread. Using transposon mutagenesis, we identified surface cell antigen 4 (Sca4) as a secreted effector of spread that specifically promotes protrusion engulfment. Sca4 interacts with the cell-adhesion protein vinculin and blocks association with vinculin's binding partner, α-catenin. Using traction and monolayer stress microscopy, we show that Sca4 reduces vinculin-dependent mechanotransduction at cell-cell junctions. Our results suggest that Sca4 relieves intercellular tension to promote protrusion engulfment, which represents a distinctive strategy for manipulating cytoskeletal force generation to enable spread.

  5. Intercellular transfer along the trichomes of the invasive terminal heterocyst forming cyanobacterium Cylindrospermopsis raciborskii CS-505.

    PubMed

    Plominsky, Álvaro M; Delherbe, Nathalie; Mandakovic, Dinka; Riquelme, Brenda; González, Karen; Bergman, Birgitta; Mariscal, Vicente; Vásquez, Mónica

    2015-03-01

    Cylindrospermopsis raciborskii CS-505 is an invasive freshwater filamentous cyanobacterium that when grown diazotrophically may develop trichomes of up to 100 vegetative cells while differentiating only two end heterocysts, the sole sites for their N2-fixation process. We examined the diazotrophic growth and intercellular transfer mechanisms in C. raciborskii CS-505. Subjecting cultures to a combined-nitrogen-free medium to elicit N2 fixation, the trichome length remained unaffected while growth rates decreased. The structures and proteins for intercellular communication showed that while a continuous periplasmic space was apparent along the trichomes, the putative septal junction sepJ gene is divided into two open reading frames and lacks several transmembrane domains unlike the situation in Anabaena, differentiating a 5-fold higher frequency of heterocysts. FRAP analyses also showed that the dyes calcein and 5-CFDA were taken up by heterocysts and vegetative cells, and that the transfer from heterocysts and 'terminal' vegetative cells showed considerably higher transfer rates than that from vegetative cells located in the middle of the trichomes. The data suggest that C. raciborskii CS-505 compensates its low-frequency heterocyst phenotype by a highly efficient transfer of the fixed nitrogen towards cells in distal parts of the trichomes (growing rapidly) while cells in central parts suffers (slow growth).

  6. Regulation of Endothelial Adherens Junctions by Tyrosine Phosphorylation

    PubMed Central

    Adam, Alejandro Pablo

    2015-01-01

    Endothelial cells form a semipermeable, regulated barrier that limits the passage of fluid, small molecules, and leukocytes between the bloodstream and the surrounding tissues. The adherens junction, a major mechanism of intercellular adhesion, is comprised of transmembrane cadherins forming homotypic interactions between adjacent cells and associated cytoplasmic catenins linking the cadherins to the cytoskeleton. Inflammatory conditions promote the disassembly of the adherens junction and a loss of intercellular adhesion, creating openings or gaps in the endothelium through which small molecules diffuse and leukocytes transmigrate. Tyrosine kinase signaling has emerged as a central regulator of the inflammatory response, partly through direct phosphorylation and dephosphorylation of the adherens junction components. This review discusses the findings that support and those that argue against a direct effect of cadherin and catenin phosphorylation in the disassembly of the adherens junction. Recent findings indicate a complex interaction between kinases, phosphatases, and the adherens junction components that allow a fine regulation of the endothelial permeability to small molecules, leukocyte migration, and barrier resealing. PMID:26556953

  7. Homotypic gap junctional communication associated with metastasis increases suppression increases with PKA kinase activity and is unaffected by P13K inhibition

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Loss of gap junctional intercellular communication (GJIC) between cancer cells is a common characteristic of malignant transformation. This communication is mediated by connexin proteins that make up the functional units of gap junctions. Connexins are highly regulated at the protein level and phosp...

  8. Methoxychlor and Vinclozolin Induce Rapid Changes in Intercellular and Intracellular Signaling in Liver Progenitor Cells.

    PubMed

    Babica, Pavel; Zurabian, Rimma; Kumar, Esha R; Chopra, Rajus; Mianecki, Maxwell J; Park, Joon-Suk; Jaša, Libor; Trosko, James E; Upham, Brad L

    2016-09-01

    Methoxychlor (MXC) and vinclozolin (VIN) are well-recognized endocrine disrupting chemicals known to alter epigenetic regulations and transgenerational inheritance; however, non-endocrine disruption endpoints are also important. Thus, we determined the effects of MXC and VIN on the dysregulation of gap junctional intercellular communication (GJIC) and activation of mitogen-activated protein kinases (MAPKs) in WB-F344 rat liver epithelial cells. Both chemicals induced a rapid dysregulation of GJIC at non-cytotoxic doses, with 30 min EC50 values for GJIC inhibition being 10 µM for MXC and 126 µM for VIN. MXC inhibited GJIC for at least 24 h, while VIN effects were transient and GJIC recovered after 4 h. VIN induced rapid hyperphosphorylation and internalization of gap junction protein connexin43, and both chemicals also activated MAPK ERK1/2 and p38. Effects on GJIC were not prevented by MEK1/2 inhibitor, but by an inhibitor of phosphatidylcholine-specific phospholipase C (PC-PLC), resveratrol, and in the case of VIN, also, by a p38 inhibitor. Estrogen (ER) and androgen receptor (AR) modulators (estradiol, ICI 182,780, HPTE, testosterone, flutamide, VIN M2) did not attenuate MXC or VIN effects on GJIC. Our data also indicate that the effects were elicited by the parental compounds of MXC and VIN. Our study provides new evidence that MXC and VIN dysregulate GJIC via mechanisms involving rapid activation of PC-PLC occurring independently of ER- or AR-dependent genomic signaling. Such alterations of rapid intercellular and intracellular signaling events involved in regulations of gene expression, tissue development, function and homeostasis, could also contribute to transgenerational epigenetic effects of endocrine disruptors.

  9. Cell Cycle Checkpoint Proteins p21 and Hus1 Regulating Intercellular Signaling Induced By Alpha Particle Irradiation

    NASA Astrophysics Data System (ADS)

    Wu, Lijun; Zhao, Ye; Wang, Jun; Hang, Haiying

    In recent years, the attentions for radiation induced bystander effects (RIBE) have been paid on the intercellular signaling events connecting the irradiated and non-irradiated cells. p21 is a member of the Cip/Kip family and plays essential roles in cell cycle progression arrest after cellular irradiation. DNA damage checkpoint protein Hus1 is a member of the Rad9-Rad1-Hus1 complex and functions as scaffold at the damage sites to facilitate the activation of downstream effectors. Using the medium trasfer method and the cells of MEF, MEF (p21-/-), MEF (p21-/-Hus1-/-) as either medium donor or receptor cells, it was found that with 5cGy alpha particle irradiation, the bystander cells showed a significant induction of -H2AX for normal MEFs (p¡0.05). However, the absence of p21 resulted in deficiency in inducing bystander effects. Further results indicated p21 affected the intercellular DNA damage signaling mainly through disrupting the production or release of the damage signals from irradiated cells. When Hus1 and p21 were both knocked out, an obvious induction of -H2AX recurred in bystander cells and the induction of -H2AX was GJIC (gap junction-mediated intercellular communication) dependent, indicating the interrelationship between p21 and Hus1 regulated the production and relay of DNA damage signals from irradiated cells to non-irradiated bystander cells.

  10. The Role of Chemical Inhibition of Gap-Junctional Intercellular Communication in Toxicology

    DTIC Science & Technology

    1989-03-01

    Communication Between Rat Liver zoithelial Cells M.S. Rczabek, J.t. Trosko, C. Jone, and S.D. Sleiqht. FireMaster BP-6 (FM), a mixture of polybrominated...V79 cells." Cell Biol. Toxicol., 3:1-15, 1987. T.J. Kavanagh, C.C. Chang, and J.E. Trosko, "Effect of the polybrominated biphenyls, Firemaster BP

  11. Chemical Tumor Promoters, Oncogenes and Growth Factors: Modulators of Gap Junctional Intercellular Communication

    DTIC Science & Technology

    1991-01-01

    Bennett and D.C. Spray, Cold Spring Harbor Laboratory, Cold Spring Harbor , NY, 1985, p. 289. 5. Madhukar, B.V., J.E. Trosko and C.C. Chang. in Cell...P.D. Hoerger. Cold Spring Harbor Labo- ratory, Cold Spring Harbor , N.Y., 1988. p. 139. 42. Lacey, S.W.. Amer. 1. Med. Sci. 29:39 (1986). 43

  12. Slit Diaphragms Contain Tight Junction Proteins

    PubMed Central

    Fukasawa, Hirotaka; Bornheimer, Scott; Kudlicka, Krystyna; Farquhar, Marilyn G.

    2009-01-01

    Slit diaphragms are essential components of the glomerular filtration apparatus, as changes in these junctions are the hallmark of proteinuric diseases. Slit diaphragms, considered specialized adherens junctions, contain both unique membrane proteins (e.g., nephrin, podocin, and Neph1) and typical adherens junction proteins (e.g., P-cadherin, FAT, and catenins). Whether slit diaphragms also contain tight junction proteins is unknown. Here, immunofluorescence, immunogold labeling, and cell fractionation demonstrated that rat slit diaphragms contain the tight junction proteins JAM-A (junctional adhesion molecule A), occludin, and cingulin. We found these proteins in the same protein complexes as nephrin, podocin, CD2AP, ZO-1, and Neph1 by cosedimentation, coimmunoprecipitation, and pull-down assays. PAN nephrosis increased the protein levels of JAM-A, occludin, cingulin, and ZO-1 several-fold in glomeruli and loosened their attachment to the actin cytoskeleton. These data extend current information about the molecular composition of slit diaphragms by demonstrating the presence of tight junction proteins, although slit diaphragms lack the characteristic morphologic features of tight junctions. The contribution of these proteins to the assembly of slit diaphragms and potential signaling cascades requires further investigation. PMID:19478094

  13. The structural organization and protein composition of lens fiber junctions

    PubMed Central

    1989-01-01

    The structural organization and protein composition of lens fiber junctions isolated from adult bovine and calf lenses were studied using combined electron microscopy, immunolocalization with monoclonal and polyclonal anti-MIP and anti-MP70 (two putative gap junction-forming proteins), and freeze-fracture and label-fracture methods. The major intrinsic protein of lens plasma membranes (MIP) was localized in single membranes and in an extensive network of junctions having flat and undulating surface topologies. In wavy junctions, polyclonal and monoclonal anti-MIPs labeled only the cytoplasmic surface of the convex membrane of the junction. Label-fracture experiments demonstrated that the convex membrane contained MIP arranged in tetragonal arrays 6-7 nm in unit cell dimension. The apposing concave membrane of the junction displayed fracture faces without intramembrane particles or pits. Therefore, wavy junctions are asymmetric structures composed of MIP crystals abutted against particle-free membranes. In thin junctions, anti-MIP labeled the cytoplasmic surfaces of both apposing membranes with varying degrees of asymmetry. In thin junctions, MIP was found organized in both small clusters and single membranes. These small clusters also abut against particle-free apposing membranes, probably in a staggered or checkerboard pattern. Thus, the structure of thin and wavy junctions differed only in the extent of crystallization of MIP, a property that can explain why this protein can produce two different antibody-labeling patterns. A conclusion of this study is that wavy and thin junctions do not contain coaxially aligned channels, and, in these junctions, MIP is unlikely to form gap junction-like channels. We suggest MIP may behave as an intercellular adhesion protein which can also act as a volume-regulating channel to collapse the lens extracellular space. Junctions constructed of MP70 have a wider overall thickness (18-20 nm) and are abundant in the cortical regions

  14. Non-invasive microfluidic gap junction assay.

    PubMed

    Chen, Sisi; Lee, Luke P

    2010-03-01

    Gap junctions are protein channels between cells that allow direct electrical and metabolic coupling via the exchange of biomolecules and ions. Their expression, though ubiquitous in most mammalian cell types, is especially important for the proper functioning of cardiac and neuronal systems. Many existing methods for studying gap junction communication suffer from either unquantifiable data or difficulty of use. Here, we measure the extent of dye spread and effective diffusivities through gap junction connected cells using a quantitative microfluidic cell biology platform. After loading dye by hydrodynamic focusing of calcein/AM, dye transfer dynamics into neighboring, unexposed cells can be monitored via timelapse fluorescent microscopy. By using a selective microfluidic dye loading over a confluent layer of cells, we found that high expression of gap junctions in C6 cells transmits calcein across the monolayer with an effective diffusivity of 3.4 x 10(-13) m(2)/s, which are highly coupled by Cx43. We also found that the gap junction blocker 18alpha-GA works poorly in the presence of serum even at high concentrations (50 microM); however, it is highly effective down to 2.5 microM in the absence of serum. Furthermore, when the drug is washed out, dye spread resumes rapidly within 1 min for all doses, indicating the drug does not affect transcriptional regulation of connexins in these Cx43+ cells, in contrast to previous studies. This integrated microfluidic platform enables the in situ monitoring of gap junction communication, yielding dynamic information about intercellular molecular transfer and pharmacological inhibition and recovery.

  15. Intestinal epithelial barrier function and tight junction proteins with heat and exercise.

    PubMed

    Dokladny, Karol; Zuhl, Micah N; Moseley, Pope L

    2016-03-15

    A single layer of enterocytes and tight junctions (intercellular multiprotein complexes) form the intestinal epithelial barrier that controls transport of molecules through transcellular and paracellular pathways. A dysfunctional or "leaky" intestinal tight junction barrier allows augmented permeation of luminal antigens, endotoxins, and bacteria into the blood stream. Various substances and conditions have been shown to affect the maintenance of the intestinal epithelial tight junction barrier. The primary focus of the present review is to analyze the effects of exertional or nonexertional (passive hyperthermia) heat stress on tight junction barrier function in in vitro and in vivo (animals and humans) models. Our secondary focus is to review changes in tight junction proteins in response to exercise or hyperthermic conditions. Finally, we discuss some pharmacological or nutritional interventions that may affect the cellular mechanisms involved in maintaining homeostasis of the intestinal epithelial tight junction barrier during heat stress or exercise.

  16. Intestinal epithelial barrier function and tight junction proteins with heat and exercise

    PubMed Central

    Zuhl, Micah N.; Moseley, Pope L.

    2015-01-01

    A single layer of enterocytes and tight junctions (intercellular multiprotein complexes) form the intestinal epithelial barrier that controls transport of molecules through transcellular and paracellular pathways. A dysfunctional or “leaky” intestinal tight junction barrier allows augmented permeation of luminal antigens, endotoxins, and bacteria into the blood stream. Various substances and conditions have been shown to affect the maintenance of the intestinal epithelial tight junction barrier. The primary focus of the present review is to analyze the effects of exertional or nonexertional (passive hyperthermia) heat stress on tight junction barrier function in in vitro and in vivo (animals and humans) models. Our secondary focus is to review changes in tight junction proteins in response to exercise or hyperthermic conditions. Finally, we discuss some pharmacological or nutritional interventions that may affect the cellular mechanisms involved in maintaining homeostasis of the intestinal epithelial tight junction barrier during heat stress or exercise. PMID:26359485

  17. Steroid hormone effects on intercellular communication between term pregnant human myometrial cells before labor.

    PubMed

    Ciray, H N; Bäckström, T; Ulmsten, U; Roomans, G M

    1996-08-01

    The appearance of gap junctions (GJs) between myometrial smooth muscle cells is one of the major events associated with the onset of labor. We have employed dye-coupling and electrical-current injection techniques to study the mechanisms by which steroid hormones regulate GJs in term pregnant myometrium of women before labor. Progesterone (P4) did not alter the input resistance (Ro) of the tissues when added to Tyrode's solution, which was used as control treatment. Octanol, the putative gap junctional uncoupling agent, increased the Ro of the cells compared to the control and P4-treated groups. The membrane potential (Em) did not differ between these groups. However, when P4 was applied after the tissue was perfused with estradiol (E2), the results changed dramatically: the Em hyperpolarized, and the Ro increased. Octanol increased the Ro in E2-treated tissues, but did not affect the Em. Consecutive application of E2, octanol, E2, and P4 resulted in rapid changes in the Ro of the cells. Dye-coupling was mostly detected between cells from controls and E2-treated tissues. These results indicate that P4 exerts its effects in the presence of E2 and that P4 has rapid effects on the intercellular communication between human myometrial cells.

  18. Dioscin augments HSV-tk-mediated suicide gene therapy for melanoma by promoting connexin-based intercellular communication

    PubMed Central

    Li, Bin; Wu, Yingya; Liu, Xijuan; Tan, Yuhui; Du, Biaoyan

    2017-01-01

    Suicide gene therapy is a promising strategy against melanoma. However, the low efficiency of the gene transfer technique can limit its application. Our preliminary data showed that dioscin, a glucoside saponin, could upregulate the expression of connexins Cx26 and Cx43, major components of gap junctions, in melanoma cells. We hypothesized that dioscin may increase the bystander effect of herpes simplex virus thymidine kinase/ganciclovir (HSV-tk/GCV) through increasing the formation of gap junctions. Further analysis showed that dioscin indeed could increase the gap junctional intercellular communication in B16 melanoma cells, resulting in more efficient GCV-induced bystander killing in B16tk cells. By contrast, overexpression of dominant negative Cx43 impaired the cell-cell communication of B16 cells and subsequently weakened the bystander effect of HSV-tk/GCV gene therapy. In vivo, combination treatment with dioscin and GCV of tumor-bearing mice with 30% positive B16tk cells and 70% wild-type B16 cells caused a significant reduction in tumor volume and weight compared to treatment with GCV or dioscin alone. Taken together, these results demonstrated that dioscin could augment the bystander effect of the HSV-tk/GCV system through increasing connexin-mediated gap junction coupling. PMID:27903977

  19. Galectin-3 Regulates Desmoglein-2 and Intestinal Epithelial Intercellular Adhesion*

    PubMed Central

    Jiang, Kun; Rankin, Carl R.; Nava, Porfirio; Sumagin, Ronen; Kamekura, Ryuta; Stowell, Sean R.; Feng, Mingli; Parkos, Charles A.; Nusrat, Asma

    2014-01-01

    The desmosomal cadherins, desmogleins, and desmocollins mediate strong intercellular adhesion. Human intestinal epithelial cells express the desmoglein-2 isoform. A proteomic screen for Dsg2-associated proteins in intestinal epithelial cells identified a lectin referred to as galectin-3 (Gal3). Gal3 bound to N-linked β-galactosides in Dsg2 extracellular domain and co-sedimented with caveolin-1 in lipid rafts. Down-regulation of Gal3 protein or incubation with lactose, a galactose-containing disaccharide that competitively inhibits galectin binding to Dsg2, decreased intercellular adhesion in intestinal epithelial cells. In the absence of functional Gal3, Dsg2 protein was internalized from the plasma membrane and degraded in the proteasome. These results report a novel role of Gal3 in stabilizing a desmosomal cadherin and intercellular adhesion in intestinal epithelial cells. PMID:24567334

  20. Plasmodesmata: intercellular tunnels facilitating transport of macromolecules in plants.

    PubMed

    Kragler, Friedrich

    2013-04-01

    In plants, intercellular structures named plasmodesmata (PD) form a continuous cytoplasmic network between neighboring cells. PD pores provide channels for intercellular symplasmic (cell-to-cell) transport throughout most tissues of the plant body. Cell-defining proteins, such as transcription factors, and regulatory non-coding sequences, such as short interfering RNA, micro RNA, protein-encoding messenger RNAs, viroids, and viral RNA/DNA genomes move via PD channels to adjacent cells. PD-mediated intercellular transport of macromolecules is a regulated process depending on the tissue, developmental stage, and nature of the transported macromolecule. In this review, PD channels and their similarity to tunneling nanotubes present in animals are highlighted. In addition, homeodomain protein movement and cellular components regulating transport are discussed.

  1. Functional assessment of gap junctions in monolayer and three-dimensional cultures of human tendon cells using fluorescence recovery after photobleaching

    NASA Astrophysics Data System (ADS)

    Kuzma-Kuzniarska, Maria; Yapp, Clarence; Pearson-Jones, Thomas W.; Jones, Andrew K.; Hulley, Philippa A.

    2014-01-01

    Gap junction-mediated intercellular communication influences a variety of cellular activities. In tendons, gap junctions modulate collagen production, are involved in strain-induced cell death, and are involved in the response to mechanical stimulation. The aim of the present study was to investigate gap junction-mediated intercellular communication in healthy human tendon-derived cells using fluorescence recovery after photobleaching (FRAP). The FRAP is a noninvasive technique that allows quantitative measurement of gap junction function in living cells. It is based on diffusion-dependent redistribution of a gap junction-permeable fluorescent dye. Using FRAP, we showed that human tenocytes form functional gap junctions in monolayer and three-dimensional (3-D) collagen I culture. Fluorescently labeled tenocytes following photobleaching rapidly reacquired the fluorescent dye from neighboring cells, while HeLa cells, which do not communicate by gap junctions, remained bleached. Furthermore, both 18 β-glycyrrhetinic acid and carbenoxolone, standard inhibitors of gap junction activity, impaired fluorescence recovery in tendon cells. In both monolayer and 3-D cultures, intercellular communication in isolated cells was significantly decreased when compared with cells forming many cell-to-cell contacts. In this study, we used FRAP as a tool to quantify and experimentally manipulate the function of gap junctions in human tenocytes in both two-dimensional (2-D) and 3-D cultures.

  2. DIFFERENTIATION OF THE JUNCTIONAL COMPLEX OF SURFACE CELLS IN THE DEVELOPING FUNDULUS BLASTODERM

    PubMed Central

    Lentz, Thomas L.; Trinkaus, J. P.

    1971-01-01

    The structure of the junctional complex between surface cells was investigated in blastula, mid gastrula, late gastrula, and early embryo of the teleost fish Fundulus heteroclitus. In blastulae, the intercellular complex is simple and consists of an apical region where the adjacent membranes are closely apposed (40–60 A) and in places touch, an intermediate zone with a wider intercellular space (> 100 A), and incipient desmosomes. In gastrulae, there are frequent points of fusion of membranes along the apical zone of the complex. Dilatations and an increased number of desmosomes in different stages of development are found along the intermediate zone. In mid gastrula, a close or gap junction with an intercellular space of 20 A occurs below the level of the desmosomes. In late gastrula, the gap junction is reduced in extent and desmosomes are better developed. In the early embryo, the basic organization of the complex is the same, although the deeply situated close junctions are no longer apparent and desmosomes and their associated system of filaments are well developed. At this time, the junctional complex is comparable to that of many epithelia and consists of an apical zonula occludens, a short zonula adherens, and deeply situated maculae adherentes. PMID:5545331

  3. Intercellular carbon nanotube translocation assessed by flow cytometry imaging.

    PubMed

    Marangon, Iris; Boggetto, Nicole; Ménard-Moyon, Cécilia; Venturelli, Enrica; Béoutis, Marie-Lys; Péchoux, Christine; Luciani, Nathalie; Wilhelm, Claire; Bianco, Alberto; Gazeau, Florence

    2012-09-12

    The fate of carbon nanotubes in the organism is still controversial. Here, we propose a statistical high-throughput imaging method to localize and quantify functionalized multiwalled carbon nanotubes in cells. We give the first experimental evidence of an intercellular translocation of carbon nanotubes. This stress-induced longitudinal transfer of nanomaterials is mediated by cell-released microvesicles known as vectors for intercellular communication. This finding raises new critical issues for nanotoxicology, since carbon nanotubes could be disseminated by circulating extracellular cell-released vesicles and visiting several cells in the course of their passage into the organism.

  4. Microtubules regulate disassembly of epithelial apical junctions

    PubMed Central

    Ivanov, Andrei I; McCall, Ingrid C; Babbin, Brian; Samarin, Stanislav N; Nusrat, Asma; Parkos, Charles A

    2006-01-01

    Background Epithelial tight junction (TJ) and adherens junction (AJ) form the apical junctional complex (AJC) which regulates cell-cell adhesion, paracellular permeability and cell polarity. The AJC is anchored on cytoskeletal structures including actin microfilaments and microtubules. Such cytoskeletal interactions are thought to be important for the assembly and remodeling of apical junctions. In the present study, we investigated the role of microtubules in disassembly of the AJC in intestinal epithelial cells using a model of extracellular calcium depletion. Results Calcium depletion resulted in disruption and internalization of epithelial TJs and AJs along with reorganization of perijunctional F-actin into contractile rings. Microtubules reorganized into dense plaques positioned inside such F-actin rings. Depolymerization of microtubules with nocodazole prevented junctional disassembly and F-actin ring formation. Stabilization of microtubules with either docetaxel or pacitaxel blocked contraction of F-actin rings and attenuated internalization of junctional proteins into a subapical cytosolic compartment. Likewise, pharmacological inhibition of microtubule motors, kinesins, prevented contraction of F-actin rings and attenuated disassembly of apical junctions. Kinesin-1 was enriched at the AJC in cultured epithelial cells and it also accumulated at epithelial cell-cell contacts in normal human colonic mucosa. Furthermore, immunoprecipitation experiments demonstrated association of kinesin-1 with the E-cadherin-catenin complex. Conclusion Our data suggest that microtubules play a role in disassembly of the AJC during calcium depletion by regulating formation of contractile F-actin rings and internalization of AJ/TJ proteins. PMID:16509970

  5. Internalization of adhesion junction proteins and their association with recycling endosome marker proteins in rat seminiferous epithelium.

    PubMed

    Young, J'Nelle S; Takai, Yoshimi; Kojic, Katarina L; Vogl, A Wayne

    2012-03-01

    Tubulobulbar complexes (TBCs) are elaborate cytoskeleton-related structures that are formed in association with intercellular junctions in the seminiferous epithelium. They consist of a cylindrical double-membrane core composed of the plasma membranes of the two attached cells, cuffed by a dendritic network of actin filaments. TBCs are proposed to be subcellular machines that internalize intercellular junctions during the extensive junction remodeling that occurs during spermatogenesis. At the apical sites of attachment between Sertoli cells and spermatids, junction disassembly is part of the sperm release mechanism. In this study, we used immunological probes to explore junction internalization and recycling at apical TBCs in the rat seminiferous epithelium. We demonstrate that β1-integrin and nectin 2 were concentrated at the ends of TBCs and for the first time show that the early endosome marker RAB5A was also distinctly localized at the ends of TBCs that appear to be the 'bulbar' regions of the complexes. Significantly, we also demonstrate that the 'long-loop' recycling endosome marker RAB11A was co-distributed with nectin 2 at junctions with early spermatids deeper in the epithelium. Our results are consistent with the hypothesis that TBCs associated with late spermatids internalize adhesion junctions and also indicate that some of the internalized junction proteins may be recycled to form junctions with the next generation of spermatids.

  6. Cyanobacterial clock, a stable phase oscillator with negligible intercellular coupling

    PubMed Central

    Amdaoud, M.; Vallade, M.; Weiss-Schaber, C.; Mihalcescu, I.

    2007-01-01

    Accuracy in cellular function has to be achieved despite random fluctuations (noise) in the concentrations of different molecular constituents inside and outside the cell. The circadian oscillator in cyanobacteria is an example of resilience to noise. This resilience could be either the consequence of intercellular communication or the intrinsic property of the built-in biochemical network. Here we investigate the intercellular coupling hypothesis. A short theoretical depiction of interacting noisy phase oscillators, confirmed by numerical simulations, allows us to discriminate the effect of coupling from noise. Experimentally, by studying the phase of concurrent populations of different initial phases, we evaluate a very small upper limit of the intercellular coupling strength. In addition, in situ entrainment experiments confirm our ability to detect a coupling of the circadian oscillator to an external force and to describe explicitly the dynamic change of the mean phase. We demonstrate, therefore, that the cyanobacterial clock stability is a built-in property as the intercellular coupling effect is negligible. PMID:17438272

  7. Coordination of Intercellular Ca2+ Signaling in Endothelial Cell Tubes of Mouse Resistance Arteries

    PubMed Central

    Socha, Matthew J.; Domeier, Timothy L.; Behringer, Erik J.; Segal, Steven S.

    2012-01-01

    Objective To test the hypothesis that Ca2+ responses to G-protein coupled receptor (GPCR) activation are coordinated between neighboring endothelial cells of resistance arteries. Methods Endothelial cell tubes were freshly isolated from superior epigastric arteries of C57BL/6 mice. Intercellular coupling was tested using microinjection of propidium iodide. Following loading with fluo-4 dye, intracellular Ca2+ responses to ACh were imaged with confocal microscopy. Results Cell-to-cell transfer of propidium iodide confirmed functional gap junction channels. 1 μM ACh evoked Ca2+ responses [9.8±0.8/min, (F/F0)=3.11±0.2] which pseudo-linescan analysis revealed to be composed of Ca2+ waves and spatially-restricted Ca2+ release events. 100 nM ACh induced Ca2+ responses of lower frequency (4.5±0.7/min) and amplitude (F/F0=1.95±0.11) composed primarily of spatially-restricted events. The interval between Ca2+ waves in Adjacent cells (0.79±0.12 s) was shorter (P<0.05) than between Nonadjacent cells (1.56±0.25 s). Spatially-restricted Ca2+ release events had similar frequencies and latencies between Adjacent and Nonadjacent cells. Inhibiting intracellular Ca2+ release with 2-APB, Xestospongin C or thapsigargin eliminated Ca2+ responses. Conclusions With moderate GPCR stimulation, localized Ca2+ release events predominate among cells. Greater GPCR stimulation evokes coordinated intercellular Ca2+ waves via the endoplasmic reticulum. Calcium signaling during GPCR activation is complex among cells, varying with stimulus intensity and proximity to actively signaling cells. PMID:22860994

  8. Epithelial adhesive junctions

    PubMed Central

    Capaldo, Christopher T.; Farkas, Attila E.

    2014-01-01

    Epithelial adhesive cell-to-cell contacts contain large, plasma membrane-spanning multiprotein aggregates that perform vital structural and signaling functions. Three prominent adhesive contacts are the tight junction, adherens junction, and the desmosome. Each junction type has unique cellular functions and a complex molecular composition. In this review, we comment on recent and exciting advances in our understanding of junction composition and function. PMID:24592313

  9. Wideband rotating junctions

    NASA Astrophysics Data System (ADS)

    Pochernyaev, V. N.

    1993-06-01

    Rotating junctions of coaxial-waveguide and waveguide type with a traveling wave coefficient exceeding 0.8 in a wide frequency range are considered. The design of these junctions is based on a method of the theory of electrodynamic circuits. Numerical results are obtained for rotating junctions of partially filled rectangular waveguide type and their particular cases.

  10. Oocyte triplet pairing for electrophysiological investigation of gap junctional coupling

    PubMed Central

    Hayar, Abdallah; Charlesworth, Amanda; Garcia-Rill, Edgar

    2010-01-01

    Gap junctions formed by expressing connexin subunits in Xenopus oocytes provide a valuable tool for revealing the gating properties of intercellular gap junctions in electrically coupled cells. We describe a new method that consists of simultaneous triple recordings from 3 apposed oocytes expressing exogenous connexins. The advantages of this method is that in one single experiment, one oocyte serves as control while a pair of oocytes, which have been manipulated differently, may be tested for different gap junctional properties. Moreover, we can study simultaneously the gap junctional coupling of 3 different pairs of oocytes in the same preparation. If the experiment consists of testing the effect of a single drug, this approach will reduce the time required, as background coupling in control pairs of oocytes does not need to be measured separately as with the conventional 2 oocyte pairing. The triplet approach also increases confidence that any changes seen in junctional communication are due to the experimental treatment and not variation in the preparation of oocytes or execution of the experiment. In this study, we show the example of testing the gap junctional properties among three oocytes, two of which are expressing rat connexin36. PMID:20230857

  11. Different roles of cadherins in the assembly and structural integrity of the desmosome complex

    PubMed Central

    Lowndes, Molly; Rakshit, Sabyasachi; Shafraz, Omer; Borghi, Nicolas; Harmon, Robert M.; Green, Kathleen J.; Sivasankar, Sanjeevi; Nelson, W. James

    2014-01-01

    ABSTRACT Adhesion between cells is established by the formation of specialized intercellular junctional complexes, such as desmosomes. Desmosomes contain isoforms of two members of the cadherin superfamily of cell adhesion proteins, desmocollins (Dsc) and desmogleins (Dsg), but their combinatorial roles in desmosome assembly are not understood. To uncouple desmosome assembly from other cell–cell adhesion complexes, we used micro-patterned substrates of Dsc2aFc and/or Dsg2Fc and collagen IV; we show that Dsc2aFc, but not Dsg2Fc, was necessary and sufficient to recruit desmosome-specific desmoplakin into desmosome puncta and produce strong adhesive binding. Single-molecule force spectroscopy showed that monomeric Dsc2a, but not Dsg2, formed Ca2+-dependent homophilic bonds, and that Dsg2 formed Ca2+-independent heterophilic bonds with Dsc2a. A W2A mutation in Dsc2a inhibited Ca2+-dependent homophilic binding, similar to classical cadherins, and Dsc2aW2A, but not Dsg2W2A, was excluded from desmosomes in MDCK cells. These results indicate that Dsc2a, but not Dsg2, is required for desmosome assembly through homophilic Ca2+- and W2-dependent binding, and that Dsg2 might be involved later in regulating a switch to Ca2+-independent adhesion in mature desmosomes. PMID:24610950

  12. Different roles of cadherins in the assembly and structural integrity of the desmosome complex

    SciTech Connect

    Lowndes, M; Rakshit, S; Shafraz, O; Borghi, N; Harmon, R M; Green, K J; Sivasankar, S; Nelson, W J

    2014-05-15

    Adhesion between cells is established by the formation of specialized intercellular junctional complexes, such as desmosomes. Desmosomes comprise two members of the cadherin superfamily of cell adhesion proteins, desmocollin (Dsc) and desmoglein (Dsg), but their combinatorial roles in desmosome assembly is not understood. To uncouple desmosome assembly from other cell-cell adhesion complexes, we used micro-patterned substrates of Dsc2aFc and/or Dsg2Fc and collagen IV; we show that Dsc2aFc, but not Dsg2Fc, was necessary and sufficient to recruit desmosome-specific desmoplakin into desmosome puncta and produce strong adhesive binding. Single Molecule Force Spectroscopy showed that monomeric Dsc2a, but not Dsg2, formed Ca2+-dependent homophilic bonds, and that Dsg2 formed Ca2+-independent heterophilic bonds with Dsc2a. A W2A mutation in Dsc2a inhibited Ca2+-dependent homophilic binding, similar to classical cadherins, and Dsc2aW2A, but not Dsg2W2A, was excluded from desmosomes in MDCK cells. These results indicate that Dsc2a, not Dsg2, is required for desmosome assembly via homophilic Ca2+- and W2/strand swap-dependent binding, and that Dsg2 may be involved later in regulating a switch to Ca2+-independent adhesion in mature desmosomes.

  13. MicroRNAs regulate tight junction proteins and modulate epithelial/endothelial barrier functions.

    PubMed

    Cichon, Christoph; Sabharwal, Harshana; Rüter, Christian; Schmidt, M Alexander

    2014-01-01

    Tightly controlled epithelial and endothelial barriers are a prerequisite for life as these barriers separate multicellular organisms from their environment and serve as first lines of defense. Barriers between neighboring epithelial cells are formed by multiple intercellular junctions including the 'apical junctional complex-AJC' with tight junctions (TJ), adherens junctions (AJ), and desmosomes. TJ consist of tetraspan transmembrane proteins like occludin, various claudins that directly control paracellular permeability, and the 'Junctional Adhesion Molecules' (JAMs). For establishing tight barriers TJ are essential but at the same time have to allow also selective permeability. For this, TJ need to be tightly regulated and controlled. This is organized by a variety of adaptor molecules, i.e., protein kinases, phosphatases and GTPases, which in turn are regulated and fine-tuned involving microRNAs (miRNAs). In this review we summarize available data on the role and targeting of miRNAs in the maintenance of epithelial and/or endothelial barriers.

  14. The role of gap junctions in inflammatory and neoplastic disorders (Review)

    PubMed Central

    Wong, Pui; Laxton, Victoria; Srivastava, Saurabh; Chan, Yin Wah Fiona; Tse, Gary

    2017-01-01

    Gap junctions are intercellular channels made of connexin proteins, mediating both electrical and biochemical signals between cells. The ability of gap junction proteins to regulate immune responses, cell proliferation, migration, apoptosis and carcinogenesis makes them attractive therapeutic targets for treating inflammatory and neoplastic disorders in different organ systems. Alterations in gap junction profile and expression levels are observed in hyperproliferative skin disorders, lymphatic vessel diseases, inflammatory lung diseases, liver injury and neoplastic disorders. It is now recognized that the therapeutic effects mediated by traditional pharmacological agents are dependent upon gap junction communication and may even act by influencing gap junction expression or function. Novel strategies for modulating the function or expression of connexins, such as the use of synthetic mimetic peptides and siRNA technology are considered. PMID:28098880

  15. ZO-1 controls endothelial adherens junctions, cell-cell tension, angiogenesis, and barrier formation.

    PubMed

    Tornavaca, Olga; Chia, Minghao; Dufton, Neil; Almagro, Lourdes Osuna; Conway, Daniel E; Randi, Anna M; Schwartz, Martin A; Matter, Karl; Balda, Maria S

    2015-03-16

    Intercellular junctions are crucial for mechanotransduction, but whether tight junctions contribute to the regulation of cell-cell tension and adherens junctions is unknown. Here, we demonstrate that the tight junction protein ZO-1 regulates tension acting on VE-cadherin-based adherens junctions, cell migration, and barrier formation of primary endothelial cells, as well as angiogenesis in vitro and in vivo. ZO-1 depletion led to tight junction disruption, redistribution of active myosin II from junctions to stress fibers, reduced tension on VE-cadherin and loss of junctional mechanotransducers such as vinculin and PAK2, and induced vinculin dissociation from the α-catenin-VE-cadherin complex. Claudin-5 depletion only mimicked ZO-1 effects on barrier formation, whereas the effects on mechanotransducers were rescued by inhibition of ROCK and phenocopied by JAM-A, JACOP, or p114RhoGEF down-regulation. ZO-1 was required for junctional recruitment of JACOP, which, in turn, recruited p114RhoGEF. ZO-1 is thus a central regulator of VE-cadherin-dependent endothelial junctions that orchestrates the spatial actomyosin organization, tuning cell-cell tension, migration, angiogenesis, and barrier formation.

  16. Capturing intercellular sugar-mediated ligand-receptor recognitions via a simple yet highly biospecific interfacial system.

    PubMed

    Li, Zhen; Deng, Si-Si; Zang, Yi; Gu, Zhen; He, Xiao-Peng; Chen, Guo-Rong; Chen, Kaixian; James, Tony D; Li, Jia; Long, Yi-Tao

    2013-01-01

    Intercellular ligand-receptor recognitions are crucial natural interactions that initiate a number of biological and pathological events. We present here the simple construction of a unique class of biomimetic interfaces based on a graphene-mediated self-assembly of glycosyl anthraquinones to a screen-printed electrode for the detection of transmembrane glycoprotein receptors expressed on a hepatoma cell line. We show that an electroactive interface confined with densely clustered galactosyl ligands is able to ingeniously recognize the asialoglycoprotein receptors on live Hep-G2 cells employing simple electrochemical techniques. The only facility used is a personal laptop in connection with a cheap and portable electrochemical workstation.

  17. Capturing intercellular sugar-mediated ligand-receptor recognitions via a simple yet highly biospecific interfacial system

    NASA Astrophysics Data System (ADS)

    Li, Zhen; Deng, Si-Si; Zang, Yi; Gu, Zhen; He, Xiao-Peng; Chen, Guo-Rong; Chen, Kaixian; James, Tony D.; Li, Jia; Long, Yi-Tao

    2013-07-01

    Intercellular ligand-receptor recognitions are crucial natural interactions that initiate a number of biological and pathological events. We present here the simple construction of a unique class of biomimetic interfaces based on a graphene-mediated self-assembly of glycosyl anthraquinones to a screen-printed electrode for the detection of transmembrane glycoprotein receptors expressed on a hepatoma cell line. We show that an electroactive interface confined with densely clustered galactosyl ligands is able to ingeniously recognize the asialoglycoprotein receptors on live Hep-G2 cells employing simple electrochemical techniques. The only facility used is a personal laptop in connection with a cheap and portable electrochemical workstation.

  18. Transfection of C6 Glioma Cells with Connexin 43 cDNA: Analysis of Expression, Intercellular Coupling, and Cell Proliferation

    NASA Astrophysics Data System (ADS)

    Zhu, D.; Caveney, S.; Kidder, G. M.; Naus, C. C. G.

    1991-03-01

    C6 glioma cells express low levels of the gap junction protein connexin 43 and its mRNA and display very weak dye coupling. When implanted into the rat cerebrum, these cells quickly give rise to a large glioma. To investigate the role of gap junctions in the tumor characteristics of these cells, we have used Lipofectin-mediated transfection to introduce a full-length cDNA encoding connexin 43. Several transfected clones were obtained that exhibited various amounts of connexin 43 mRNA transcribed from the inserted cDNA. Immunocytochemical analysis revealed an increase in the amount of connexin 43 immunoreactivity in the transfected cells, being localized at areas of intercellular contact as well as in the cytoplasm. The level of dye coupling was also assessed and found to correlate with the amount of connexin 43 mRNA. When cell proliferation was followed over several days, cells expressing the transfected cDNA grew more slowly than nontransfected cells. These transfected cells will be useful in examining the role of gap junctions in tumorigenesis.

  19. Structural basis for the selective permeability of channels made of communicating junction proteins

    PubMed Central

    Ek-Vitorin, Jose F.; Burt, Janis M.

    2012-01-01

    The open state(s) of gap junction channels is evident from their permeation by small ions in response to an applied intercellular (transjunctional/transchannel) voltage gradient. That an open channel allows variable amounts of current to transit from cell-to-cell in the face of a constant intercellular voltage difference indicates channel open/closing can be complete or partial. The physiological significance of such open state options is, arguably, the main concern of junctional regulation. Because gap junctions are permeable to many substances, it is sensible to inquire whether and how each open state influences the intercellular diffusion of molecules as valuable as, but less readily detected than current-carrying ions. Presumably, structural changes perceived as shifts in channel conductivity would significantly alter the transjunctional diffusion of molecules whose limiting diameter approximates the pore’s limiting diameter. Moreover, changes in junctional permeability to some molecules might occur without evident changes in conductivity, either at macroscopic or single channel level. Open gap junction channels allow the exchange of cytoplasmic permeants between contacting cells by simple diffusion. The identity of such permeants, and the functional circumstances and consequences of their junctional exchange presently constitute the most urgent (and demanding) themes of the field. Here, we consider the necessity for regulating this exchange, the possible mechanism(s) and structural elements likely involved in such regulation, and how regulatory phenomena could be perceived as changes in chemical vs. electrical coupling; an overall reflection on our collective knowledge of junctional communication is then applied to suggest new avenues of research. PMID:22342665

  20. Gap Junction Enhancer Potentiates Cytotoxicity of Cisplatin in Breast Cancer Cells.

    PubMed

    Ding, Ying; Nguyen, Thu Annelise

    2012-11-01

    Cisplatin is one of the most widely used anti-cancer drugs due to its ability to damage DNA and induce apoptosis. However, increasing reports of side effects and drug resistance indicate the limitation of cisplatin in cancer therapeutics. Recent studies showed that inhibition of gap junctions diminishes the cytotoxic effect and contributes to drug resistance. Therefore, identification of molecules that counteract gap junctional inhibition without decreasing the anti-cancer effect of cisplatin could be used in combinational treatment, potentiating cisplatin efficacy and preventing resistance. This study investigates the effects of combinational treatment of cisplatin and PQ1, a gap junction enhancer, in T47D breast cancer cells. Our results showed that combinational treatment of PQ1 and cisplatin increased gap junctional intercellular communication (GJIC) as well as expressions of connexins (Cx26, Cx32 and Cx43), and subsequently decreased cell viability. Ki67, a proliferation marker, was decreased by 75% with combinational treatment. Expressions of pro-apoptotic factors (cleaved caspase-3/-8/-9 and bax) were increased by the combinational treatment with PQ1 and cisplatin; whereas, the pro-survival factor, bcl-2, was decreased by the combinational treatment. Our study demonstrates for the first time that the combinational treatment with gap junction enhancers can counteract cisplatin induced inhibition of gap junctional intercellular communication and reduction of connexin expression, thereby increasing the efficacy of cisplatin in cancer cells.

  1. Inhibition of connexin43 gap junction channels by the endocrine disruptor ioxynil

    SciTech Connect

    Leithe, Edward; Kjenseth, Ane; Bruun, Jarle; Sirnes, Solveig; Rivedal, Edgar

    2010-08-15

    Gap junctions are intercellular plasma membrane domains containing channels that mediate transport of ions, metabolites and small signaling molecules between adjacent cells. Gap junctions play important roles in a variety of cellular processes, including regulation of cell growth and differentiation, maintenance of tissue homeostasis and embryogenesis. The constituents of gap junction channels are a family of trans-membrane proteins called connexins, of which the best-studied is connexin43. Connexin43 functions as a tumor suppressor protein in various tissue types and is frequently dysregulated in human cancers. The pesticide ioxynil has previously been shown to act as an endocrine disrupting chemical and has multiple effects on the thyroid axis. Furthermore, both ioxynil and its derivative ioxynil octanoate have been reported to induce tumors in animal bioassays. However, the molecular mechanisms underlying the possible tumorigenic effects of these compounds are unknown. In the present study we show that ioxynil and ioxynil octanoate are strong inhibitors of connexin43 gap junction channels. Both compounds induced rapid loss of connexin43 gap junctions at the plasma membrane and increased connexin43 degradation. Ioxynil octanoate, but not ioxynil, was found to be a strong activator of ERK1/2. The compounds also had different effects on the phosphorylation status of connexin43. Taken together, the data show that ioxynil and ioxynil octanoate are potent inhibitors of intercellular communication via gap junctions.

  2. Inter-Cellular Forces Orchestrate Contact Inhibition of Locomotion

    PubMed Central

    Davis, John R.; Luchici, Andrei; Mosis, Fuad; Thackery, James; Salazar, Jesus A.; Mao, Yanlan; Dunn, Graham A.; Betz, Timo; Miodownik, Mark; Stramer, Brian M.

    2015-01-01

    Summary Contact inhibition of locomotion (CIL) is a multifaceted process that causes many cell types to repel each other upon collision. During development, this seemingly uncoordinated reaction is a critical driver of cellular dispersion within embryonic tissues. Here, we show that Drosophila hemocytes require a precisely orchestrated CIL response for their developmental dispersal. Hemocyte collision and subsequent repulsion involves a stereotyped sequence of kinematic stages that are modulated by global changes in cytoskeletal dynamics. Tracking actin retrograde flow within hemocytes in vivo reveals synchronous reorganization of colliding actin networks through engagement of an inter-cellular adhesion. This inter-cellular actin-clutch leads to a subsequent build-up in lamellar tension, triggering the development of a transient stress fiber, which orchestrates cellular repulsion. Our findings reveal that the physical coupling of the flowing actin networks during CIL acts as a mechanotransducer, allowing cells to haptically sense each other and coordinate their behaviors. PMID:25799385

  3. Intercellular trafficking and protein delivery by a herpesvirus structural protein.

    PubMed

    Elliott, G; O'Hare, P

    1997-01-24

    We show that the HSV-1 structural protein VP22 has the remarkable property of intercellular transport, which is so efficient that following expression in a subpopulation the protein spreads to every cell in a monolayer, where it concentrates in the nucleus and binds chromatin. VP22 movement was observed both after delivery of DNA by transfection or microinjection and during virus infection. Moreover, we demonstrate that VP22 trafficking occurs via a nonclassical Golgi-independent mechanism. Sensitivity to cytochalasin D treatment suggests that VP22 utilizes a novel trafficking pathway that involves the actin cytoskeleton. In addition, we demonstrate intercellular transport of a VP22 fusion protein after endogenous synthesis or exogenous application, indicating that VP22 may have potential in the field of protein delivery.

  4. Bulk-boundary correspondence from the intercellular Zak phase

    NASA Astrophysics Data System (ADS)

    Rhim, Jun-Won; Behrends, Jan; Bardarson, Jens H.

    2017-01-01

    The Zak phase γ , the generalization of the Berry phase to Bloch wave functions in solids, is often used to characterize inversion-symmetric one-dimensional (1D) topological insulators. Due to its dependence on the real-space origin and unit cell, however, there is an ambiguity in its use in a bulk-boundary correspondence. Here, we extract an origin-independent part of γ , the so-called intercellular Zak phase γinter, and show that it is a bulk quantity that unambiguously predicts the number of surface modes. Specifically, a neutral finite 1D tight-binding system has ns=γinter/π (mod 2) in-gap surface modes below the Fermi level if there exists a commensurate inversion-symmetric bulk unit cell. We demonstrate this in two steps: First, we verify that ±e γinter/2 π (mod e ) equals the extra charge accumulation in the surface region in a terminated system of a translationally invariant 1D insulator, while the remnant part of γ , the intracellular Zak phase γintra, corresponds to the electronic part of the bulk's unit-cell dipole moment. Second, we show that the extra charge accumulation is related to the number of surface modes when the unit cell is inversion symmetric. We study several tight-binding models to quantitatively check both the relation between the extra charge accumulation and the intercellular Zak phase, and the bulk-boundary correspondence using the intercellular Zak phase.

  5. Exosomes as mediators of intercellular communication: clinical implications.

    PubMed

    Nazimek, Katarzyna; Bryniarski, Krzysztof; Santocki, Michał; Ptak, Włodzimierz

    2015-01-01

    Cells of multicellular organisms exchange informative signals by diverse mechanisms. Recent findings uncovered the special role of extracellular vesicles, especially exosomes, in intercellular communication. Exosomes, present in all tested human bodily fluids, carry various functional compounds including proteins, lipids, and diverse RNA molecules. The composition of exosome cargo in vivo is likely formed by a regulated selection of specific components and can express the current status of the exosome-secreting cell. Therefore, particular emphasis is now placed on the extremely high potential of exosomes as essentially noninvasive prognostic and diagnostic biomarkers, but also as therapeutic nanocarriers, especially after the discovery that their cargo as well as cell-targeting specificity could be shaped in vitro. In addition, targeting the exosomes mediating pathological intercellular communication may also express high therapeutic potential. Hence, numerous studies are conducted to explore the profile and function of exosomes and their cargo in health and disease and to shape their properties to facilitate their clinical application. The present review summarizes the current knowledge on the role of exosomes in different physiological and pathological mechanisms of intercellular communication with a particular focus on the use of exosomes in the diagnosis and treatment of various inflammatory, cardiovascular, metabolic, and neurodegenerative disorders as well as malignant neoplasms.

  6. Activation of the δ-opioid receptor promotes cutaneous wound healing by affecting keratinocyte intercellular adhesion and migration

    PubMed Central

    Bigliardi, P L; Neumann, C; Teo, Y L; Pant, A; Bigliardi-Qi, M

    2015-01-01

    BACKGROUND AND PURPOSE In addition to its analgesic functions, the peripheral opioid receptor system affects skin homeostasis by influencing cell differentiation, migration and adhesion; also, wound healing is altered in δ-opioid receptor knockout mice (DOPr–/–). Hence, we investigated δ-opioid receptor effects on the expression of several proteins of the desmosomal junction complex and on the migratory behaviour of keratinocytes. EXPERIMENTAL APPROACH Expression levels of desmosomal cadherins in wild-type and DOPr–/– mice, and the morphology of intercellular adhesion in human keratinocytes were analysed by immunofluorescence. To investigate the δ-opioid receptor activation pathway, protein expression was studied using Western blot and its effect on cellular migration determined by in vitro live cell migration recordings from human keratinocytes. KEY RESULTS Expression of the desmosomal cadherins, desmogleins 1 and 4, was up-regulated in skin from DOPr–/– mice, and down-regulated in δ-opioid receptor-overexpressing human keratinocytes. The localization of desmoplakin expression was rearranged from linear arrays emanating from cell borders to puncta in cell periphery, resulting in less stable intercellular adhesion. Migration and wound recovery were enhanced in human keratinocyte monolayers overexpressing δ-opioid receptors in vitro. These δ-opioid receptor effects were antagonized by specific PKCα/β inhibition indicating they were mediated through the PKC signalling pathway. Finally, cells overexpressing δ-opioid receptors developed characteristically long but undirected protrusions containing filamentous actin and δ-opioid receptors, indicating an enhanced migratory phenotype. CONCLUSION AND IMPLICATIONS Opioid receptors affect intercellular adhesion and wound healing mechanisms, underlining the importance of a cutaneous neuroendocrine system in wound healing and skin homeostasis. LINKED ARTICLES This article is part of a themed section on

  7. β-Catenin Serves as a Clutch between Low and High Intercellular E-Cadherin Bond Strengths

    PubMed Central

    Bajpai, Saumendra; Feng, Yunfeng; Wirtz, Denis; Longmore, Gregory D.

    2013-01-01

    A wide range of invasive pathological outcomes originate from the loss of epithelial phenotype and involve either loss of function or downregulation of transmembrane adhesive receptor complexes, including Ecadherin (Ecad) and binding partners β-catenin and α-catenin at adherens junctions. Cellular pathways regulating wild-type β-catenin level, or direct mutations in β-catenin that affect the turnover of the protein have been shown to contribute to cancer development, through induction of uncontrolled proliferation of transformed tumor cells, particularly in colon cancer. Using single-molecule force spectroscopy, we show that depletion of β-catenin or the prominent cancer-related S45 deletion mutation in β-catenin present in human colon cancers both weaken tumor intercellular Ecad/Ecad bond strength and diminishes the capacity of specific extracellular matrix proteins—including collagen I, collagen IV, and laminin V—to modulate intercellular Ecad/Ecad bond strength through α-catenin and the kinase activity of glycogen synthase kinase 3 (GSK-3β). Thus, in addition to regulating tumor cell proliferation, cancer-related mutations in β-catenin can influence tumor progression by weakening the adhesion of tumor cells to one another through reduced individual Ecad/Ecad bond strength and cellular adhesion to specific components of the extracellular matrix and the basement membrane. PMID:24268141

  8. Gap Junctions in the Control of Vascular Function

    PubMed Central

    Duling, Brian R.

    2009-01-01

    Abstract Direct intercellular communication via gap junctions is critical in the control and coordination of vascular function. In the cardiovascular system, gap junctions are made up of one or more of four connexin proteins: Cx37, Cx40, Cx43, and Cx45. The expression of more than one gap-junction protein in the vasculature is not redundant. Rather, vascular connexins work in concert, first during the development of the cardiovascular system, and then in integrating smooth muscle and endothelial cell function, and in coordinating cell function along the length of the vessel wall. In addition, connexin-based channels have emerged as an important signaling pathway in the astrocyte-mediated neurovascular coupling. Direct electrical communication between endothelial cells and vascular smooth muscle cells via gap junctions is thought to play a relevant role in the control of vasomotor tone, providing the signaling pathway known as endothelium-derived hyperpolarizing factor (EDHF). Consistent with the importance of gap junctions in the regulation of vasomotor tone and arterial blood pressure, the expression of connexins is altered in diseases associated with vascular complications. In this review, we discuss the participation of connexin-based channels in the control of vascular function in physiologic and pathologic conditions, with a special emphasis on hypertension and diabetes. Antioxid. Redox Signal. 11, 251–266. PMID:18831678

  9. Connexin expression and intercellular communication in two- and three-dimensional in vitro cultures of human bladder carcinoma.

    PubMed Central

    Knuechel, R.; Siebert-Wellnhofer, A.; Traub, O.; Dermietzel, R.

    1996-01-01

    The identification of gap-junctional proteins (connexins) and the preparation of related antibodies provides new tools to study patterns of intercellular communication in tumors. Focusing on the biology of human bladder carcinoma, we compared the expression of gap-junctional proteins (connexins Cx26, Cx32, and Cx43) with a dye-coupling assay for gap-junctional intercellular communication in three cell lines with different urothelial differentiation. The cell lines HCV-29, RT4, and J82 were initially grown as monolayers of different ages. Connexin expression was found mostly positive over the time of culture and found constantly negative only in J82 cells for Cx26 and HCV-29 cells for Cx32. In HCV-29 cells, Cx26 increased in positivity over the time of culture. Western blotting with the antibodies confirmed the findings. Comparisons of dye transfer using Lucifer Yellow showed an increase of coupling in the normal urothelial cell line HCV-29 in contrast to a decrease of coupling in the tumor cell lines. Data were extended by multicellular spheroid (MCS) co-cultures with the stromal fibroblast line N1. In three-dimensional cultures as MCSs, Cx26 was increased in proximity of RT4 tumor cells to fibroblasts, and positivity was maintained in J82 cells. E-cadherin expression in cell lines showed no change in dependence of growth state. The data suggest that Cx26 plays a role in negative growth control or differentiation of urothelial cells. Preliminary comparative data on normal and neoplastic urothelium show all three connexins in normal urothelium, in contrast to varying amounts of Cx43 and low amounts of Cx32 in tumors and evident loss of Cx26 in low-grade tumors. Discrepancies between monolayer and MCS cultures are most likely due to higher differentiation in MCSs, and the continuation of systematic work with heterologous MCSs is indicated for more information on the role of gap-junctional proteins in human tumors. Images Figure 1 Figure 2 Figure 3 Figure 5 Figure 6

  10. An electrostatic mechanism for Ca2+-mediated regulation of gap junction channels

    PubMed Central

    Bennett, Brad C.; Purdy, Michael D.; Baker, Kent A.; Acharya, Chayan; McIntire, William E.; Stevens, Raymond C.; Zhang, Qinghai; Harris, Andrew L.; Abagyan, Ruben; Yeager, Mark

    2016-01-01

    Gap junction channels mediate intercellular signalling that is crucial in tissue development, homeostasis and pathologic states such as cardiac arrhythmias, cancer and trauma. To explore the mechanism by which Ca2+ blocks intercellular communication during tissue injury, we determined the X-ray crystal structures of the human Cx26 gap junction channel with and without bound Ca2+. The two structures were nearly identical, ruling out both a large-scale structural change and a local steric constriction of the pore. Ca2+ coordination sites reside at the interfaces between adjacent subunits, near the entrance to the extracellular gap, where local, side chain conformational rearrangements enable Ca2+chelation. Computational analysis revealed that Ca2+-binding generates a positive electrostatic barrier that substantially inhibits permeation of cations such as K+ into the pore. Our results provide structural evidence for a unique mechanism of channel regulation: ionic conduction block via an electrostatic barrier rather than steric occlusion of the channel pore. PMID:26753910

  11. Tricellular Tight Junctions in the Inner Ear

    PubMed Central

    2016-01-01

    Tight junctions (TJs) are structures that seal the space between the epithelial cell sheets. In the inner ear, the barrier function of TJs is indispensable for the separation of the endolymphatic and perilymphatic spaces, which is essential for the generation and maintenance of the endocochlear potential (EP). TJs are formed by the intercellular binding of membrane proteins, known as claudins, and mutations in these proteins cause deafness in humans and mice. Within the epithelial cell sheet, however, a bound structure is present at the site where the corners of three cells meet (tricellular tight junctions (tTJs)), and the maintenance of the barrier function at this location cannot be explained by the claudins alone. Tricellulin and the angulin family of proteins (angulin-1/LSR, angulin-2/ILDR1, and angulin-3/ILDR2) have been identified as tTJ-associated proteins. Tricellulin and ILDR1 are localized at the tTJ and alterations in these proteins have been reported to be involved in deafness. In this review, we will present the current state of knowledge for tTJs. PMID:27195292

  12. Role of gap junction channel in the development of beat-to-beat action potential repolarization variability and arrhythmias.

    PubMed

    Magyar, Janos; Banyasz, Tamas; Szentandrassy, Norbert; Kistamas, Kornel; Nanasi, Peter P; Satin, Jonathan

    2015-01-01

    The short-term beat-to-beat variability of cardiac action potential duration (SBVR) occurs as a random alteration of the ventricular repolarization duration. SBVR has been suggested to be more predictive of the development of lethal arrhythmias than the action potential prolongation or QT prolongation of ECG alone. The mechanism underlying SBVR is not completely understood but it is known that SBVR depends on stochastic ion channel gating, intracellular calcium handling and intercellular coupling. Coupling of single cardiomyocytes significantly decreases the beat-to-beat changes in action potential duration (APD) due to the electrotonic current flow between neighboring cells. The magnitude of this electrotonic current depends on the intercellular gap junction resistance. Reduced gap junction resistance causes greater electrotonic current flow between cells, and reduces SBVR. Myocardial ischaemia (MI) is known to affect gap junction channel protein expression and function. MI increases gap junction resistance that leads to slow conduction, APD and refractory period dispersion, and an increase in SBVR. Ultimately, development of reentry arrhythmias and fibrillation are associated post-MI. Antiarrhythmic drugs have proarrhythmic side effects requiring alternative approaches. A novel idea is to target gap junction channels. Specifically, the use of gap junction channel enhancers and inhibitors may help to reveal the precise role of gap junctions in the development of arrhythmias. Since cell-to-cell coupling is represented in SBVR, this parameter can be used to monitor the degree of coupling of myocardium.

  13. Modulation of metabolic communication through gap junction channels by transjunctional voltage; synergistic and antagonistic effects of gating and ionophoresis

    PubMed Central

    Palacios-Prado, Nicolás; Bukauskas, Feliksas F.

    2011-01-01

    Gap junction (GJ) channels assembled from connexin (Cx) proteins provide a structural basis for direct electrical and metabolic cell-cell communication. Here, we focus on gating and permeability properties of Cx43/Cx45 heterotypic GJs exhibiting asymmetries of both voltage-gating and transjunctional flux (Jj) of fluorescent dyes depending on transjunctional voltage (Vj). Relatively small differences in the resting potential of communicating cells can substantially reduce or enhance this flux at relative negativity or positivity on Cx45 side, respectively. Similarly, series of Vj pulses resembling bursts of action potentials (APs) reduce Jj when APs initiate in the cell expressing Cx43 and increase Jj when APs initiate in the cell expressing Cx45. Jj of charged fluorescent dyes is affected by ionophoresis and Vj-gating and the asymmetry of Jj-Vj dependence in heterotypic GJs is enhanced or reduced when ionophoresis and Vj-gating work in a synergistic or antagonistic manner, respectively. Modulation of cell-to-cell transfer of metabolites and signaling molecules by Vj may occur in excitable as well as non-excitable tissues and may be more expressed in the border between normal and pathological regions where intercellular gradients of membrane potential and concentration of ions are substantially altered. PMID:21930112

  14. Indian Ocean Triple Junction

    SciTech Connect

    Tapscott, C.R.; Patriat, P.; Fisher, R.L.; Sclater, J.G.; Hoskins, H.; Parsons, B.

    1980-09-10

    The boundaries of three major plates (Africa, India, and Antarctica) meet in a triple junction in the Indian Ocean near 25 /sup 0/S, 70 /sup 0/E. Using observed bathymetry and magnetic anomalies, we locate the junction to within 5 km and show that it is a ridge-ridge-ridge type. Relative plate motion is N60 /sup 0/E at 50 mm/yr (full rate) across the Central Indian Ridge, N47 /sup 0/E at 60 mm/yr across the Southeast Indian Ridge, and N3 /sup 0/W at 15 mm/yr across te Southwest Indian Ridge; the observed velocity triangle is closed. Poles of instantaneous relative plate motion are determined for all plate pairs. The data in the South Atlantic and Indian oceans are consistent with a rigid African plate without significant internal deformation. Two of the ridges at the triple junction are normal midocean spreading centers with well-defined median valleys. The Southwest Indian Ridge, however, has a peculiar morphology near the triple junction, that of an elongate triangular deep, with the triple junction at its apex. The floor of the deep represents crust formed at the Southwest Indian Ridge, and the morphology is a consequence of the evolution of the triple junction and is similar to that at the Galapagos Triple Junction. Though one cannot determine with precision the stability conditions at the triple junction, the development of the junction over the last 10 m.y. can be mapped, and the topographic expressions of the triple junction traces may be detected on the three plates.

  15. Evidence for a minigap in YBCO grain boundary Josephson junctions.

    PubMed

    Lucignano, P; Stornaiuolo, D; Tafuri, F; Altshuler, B L; Tagliacozzo, A

    2010-10-01

    Self-assembled YBaCuO diffusive grain boundary submicron Josephson junctions offer a realization of a special regime of the proximity effect, where normal state coherence prevails on the superconducting coherence in the barrier region. Resistance oscillations from the current-voltage characteristic encode mesoscopic information on the junction and more specifically on the minigap induced in the barrier. Their persistence at large voltages is evidence of the long lifetime of the antinodal (high energy) quasiparticles.

  16. Differential regulation of the levels of three gap junction mRNAs in Xenopus embryos

    PubMed Central

    1990-01-01

    Xenopus mRNAs that potentially encode gap junction proteins in the oocyte and early embryo have been identified by low-stringency screening of cDNA libraries with cloned mammalian gap junction cDNAs. The levels of these mRNAs show strikingly different temporal regulation and tissue distribution. Using a nomenclature designed to stress important structural similarities of distinct gap junction gene products, the deduced polypeptides have been designated the Xenopus alpha 1 and alpha 2 gap junction proteins. The alpha 2 gap junction mRNA is a maternal transcript that disappears by the late gastrula stage. It is not detected in any organ of the adult except the ovary, and resides primarily, if not exclusively, in the oocytes and early embryos. The alpha 1 gap junction mRNA appears during organogenesis, and is detected in RNA from a wide variety of organs. It is also found in full-grown oocytes, but is rapidly degraded upon oocyte maturation, both in vivo and in vitro. The alpha 1 and alpha 2 mRNAs encode proteins with different degrees of amino acid sequence similarity to the predominant gap junction subunit of the mammalian heart (connexin 43). Together with our earlier report of a mid-embryonic (beta 1) gap junction mRNA, the results suggest that intercellular communication during oocyte growth and postfertilization development is a complex phenomenon involving the coordinated regulation of several genes. PMID:2155241

  17. Molecular basis of gap junctional communication in the CNS of the leech Hirudo medicinalis.

    PubMed

    Dykes, Iain M; Freeman, Fiona M; Bacon, Jonathan P; Davies, Jane A

    2004-01-28

    Gap junctions are intercellular channels that allow the passage of ions and small molecules between cells. In the nervous system, gap junctions mediate electrical coupling between neurons. Despite sharing a common topology and similar physiology, two unrelated gap junction protein families exist in the animal kingdom. Vertebrate gap junctions are formed by members of the connexin family, whereas invertebrate gap junctions are composed of innexin proteins. Here we report the cloning of two innexins from the leech Hirudo medicinalis. These innexins show a differential expression in the leech CNS: Hm-inx1 is expressed by every neuron in the CNS but not in glia, whereas Hm-inx2 is expressed in glia but not neurons. Heterologous expression in the paired Xenopus oocyte system demonstrated that both innexins are able to form functional homotypic gap junctions. Hm-inx1 forms channels that are not strongly gated. In contrast, Hm-inx2 forms channels that are highly voltage-dependent; these channels demonstrate properties resembling those of a double rectifier. In addition, Hm-inx1 and Hm-inx2 are able to cooperate to form heterotypic gap junctions in Xenopus oocytes. The behavior of these channels is primarily that predicted from the properties of the constituent hemichannels but also demonstrates evidence of an interaction between the two. This work represents the first demonstration of a functional gap junction protein from a Lophotrochozoan animal and supports the hypothesis that connexin-based communication is restricted to the deuterostome clade.

  18. Regulation and roles for claudin-family tight junction proteins

    PubMed Central

    Findley, Mary K.; Koval, Michael

    2009-01-01

    Transmembrane proteins known as claudins play a critical role in tight junctions by regulating paracellular barrier permeability. The control of claudin assembly into tight junctions requires a complex interplay between several classes of claudins, other transmembrane proteins and scaffold proteins. Claudins are also subject to regulation by post-translational modifications including phosphorylation and palmitoylation. Several human diseases have been linked to claudin mutations, underscoring the physiologic function of these proteins. Roles for claudins in regulating cell phenotype and growth control also are beginning to emerge, suggesting a multifaceted role for claudins in regulation of cells beyond serving as a simple structural element of tight junctions. PMID:19319969

  19. Intercellular salicylic acid accumulation during compatible and incompatible Arabidopsis-Pseudomonas syringae interactions.

    PubMed

    Wilson, Daniel C; Carella, Philip; Cameron, Robin K

    2014-01-01

    The phytohormone salicylic acid (SA) plays an important role in several disease resistance responses. During the Age-Related Resistance (ARR) response that occurs in mature Arabidopsis responding to Pseudomonas syringae pv tomato (Pst), SA accumulates in the intercellular space where it may act as an antimicrobial agent. Recently we measured intracellular and intercellular SA levels in young, ARR-incompetent plants responding to virulent and avirulent strains of Pst to determine if intercellular SA accumulation is a component of additional defense responses to Pst. In young plants virulent Pst suppressed both intra- and intercellular SA accumulation in a coronatine-dependent manner. In contrast, high levels of intra- and intercellular SA accumulated in response to avirulent Pst. Our results support the idea that SA accumulation in the intercellular space is an important component of multiple defense responses. Future research will include understanding how mature plants counteract the effects of coronatine during the ARR response.

  20. POLARIZED INTERCELLULAR BRIDGES IN OVARIAN FOLLICLES OF THE CECROPIA MOTH

    PubMed Central

    Woodruff, Richard I.; Telfer, William H.

    1973-01-01

    Fluorescein-labeled rabbit serum globulin was injected into vitellogenic oocytes of the cecropia moth. Though the label spread throughout the ooplasm in less than 30 min, it was unable even after 2 h to cross the complex of intercellular bridges connecting the oocyte to its seven nurse cells. After injection into a single nurse cell, fluorescence was detected in the oocyte adjacent to the bridge complex within 3 min and had spread throughout the ooplasm in 30 min. Here also, the cell bodies of the six uninjected nurse cells remained nonfluorescent. Four of the nurse cells are not bridged directly to the oocyte but only through the apical ends of their siblings. Unidirectional movement must therefore occur in the apical cytoplasm of the nurse cells, as well as in the intercellular bridges. The nurse cells of healthy follicles had an intracellular electrical potential -40 mV relative to blood or dissecting solution, while oocytes measured -30 mV. A mV difference was also detected by direct comparison between a ground electrode in one cell and a recording electrode in the other. Three conditions were found in which the 10 mV difference was reduced or reversed in polarity. In all three cases fluorescent globulin was able in some degree to cross the bridges from the oocyte to the nurse cells. PMID:4125369

  1. Sticky Matrix: Adhesion Mechanism of the Staphylococcal Polysaccharide Intercellular Adhesin.

    PubMed

    Formosa-Dague, Cécile; Feuillie, Cécile; Beaussart, Audrey; Derclaye, Sylvie; Kucharíková, Soňa; Lasa, Iñigo; Van Dijck, Patrick; Dufrêne, Yves F

    2016-03-22

    The development of bacterial biofilms on surfaces leads to hospital-acquired infections that are difficult to fight. In Staphylococci, the cationic polysaccharide intercellular adhesin (PIA) forms an extracellular matrix that connects the cells together during biofilm formation, but the molecular forces involved are unknown. Here, we use advanced force nanoscopy techniques to unravel the mechanism of PIA-mediated adhesion in a clinically relevant methicillin-resistant Staphylococcus aureus (MRSA) strain. Nanoscale multiparametric imaging of the structure, adhesion, and elasticity of bacteria expressing PIA shows that the cells are surrounded by a soft and adhesive matrix of extracellular polymers. Cell surface softness and adhesion are dramatically reduced in mutant cells deficient for the synthesis of PIA or under unfavorable growth conditions. Single-cell force spectroscopy demonstrates that PIA promotes cell-cell adhesion via the multivalent electrostatic interaction with polyanionic teichoic acids on the S. aureus cell surface. This binding mechanism rationalizes, at the nanoscale, the well-known ability of PIA to strengthen intercellular adhesion in staphylococcal biofilms. Force nanoscopy offers promising prospects for understanding the fundamental forces in antibiotic-resistant biofilms and for designing anti-adhesion compounds targeting matrix polymers.

  2. Intercellular communication in malignant pleural mesothelioma: properties of tunneling nanotubes

    PubMed Central

    Ady, Justin W.; Desir, Snider; Thayanithy, Venugopal; Vogel, Rachel I.; Moreira, André L.; Downey, Robert J.; Fong, Yuman; Manova-Todorova, Katia; Moore, Malcolm A. S.; Lou, Emil

    2014-01-01

    Malignant pleural mesothelioma is a particularly aggressive and locally invasive malignancy with a poor prognosis despite advances in understanding of cancer cell biology and development of new therapies. At the cellular level, cultured mesothelioma cells present a mesenchymal appearance and a strong capacity for local cellular invasion. One important but underexplored area of mesothelioma cell biology is intercellular communication. Our group has previously characterized in multiple histological subtypes of mesothelioma a unique cellular protrusion known as tunneling nanotubes (TnTs). TnTs are long, actin filament-based, narrow cytoplasmic extensions that are non-adherent when cultured in vitro and are capable of shuttling cellular cargo between connected cells. Our prior work confirmed the presence of nanotube structures in tumors resected from patients with human mesothelioma. In our current study, we quantified the number of TnTs/cell among various mesothelioma subtypes and normal mesothelial cells using confocal microscopic techniques. We also examined changes in TnT length over time in comparison to cell proliferation. We further examined potential approaches to the in vivo study of TnTs in animal models of cancer. We have developed novel approaches to study TnTs in aggressive solid tumor malignancies and define fundamental characteristics of TnTs in malignant mesothelioma. There is mounting evidence that TnTs play an important role in intercellular communication in mesothelioma and thus merit further investigation of their role in vivo. PMID:25400582

  3. E-cadherin and Src associate with extradesmosomal Dsg3 and modulate desmosome assembly and adhesion.

    PubMed

    Rötzer, Vera; Hartlieb, Eva; Vielmuth, Franziska; Gliem, Martin; Spindler, Volker; Waschke, Jens

    2015-12-01

    Desmosomes provide strong intercellular cohesion essential for the integrity of cells and tissues exposed to continuous mechanical stress. For desmosome assembly, constitutively synthesized desmosomal cadherins translocate to the cell-cell border, cluster and mature in the presence of Ca(2+) to stable cell contacts. As adherens junctions precede the formation of desmosomes, we investigated in this study the relationship between the classical cadherin E-cadherin and the desmosomal cadherin Desmoglein 3 (Dsg3), the latter of which is indispensable for cell-cell adhesion in keratinocytes. By using autoantibodies from patients with the blistering skin disease pemphigus vulgaris (PV), we showed in loss of function studies that E-cadherin compensates for effects of desmosomal disassembly. Overexpression of E-cadherin reduced the loss of cell cohesion induced by PV autoantibodies and attenuated activation of p38 MAPK. Silencing of E-cadherin abolished the localization of Dsg3 at the membrane and resulted in a shift of Dsg3 from the cytoskeletal to the non-cytoskeletal protein pool which conforms to the notion that E-cadherin regulates desmosome assembly. Mechanistically, we identified a complex consisting of extradesmosomal Dsg3, E-cadherin, β-catenin and Src and that the stability of this complex is regulated by Src. Moreover, Dsg3 and E-cadherin are phosphorylated on tyrosine residues in a Src-dependent manner and Src activity is required for recruiting Dsg3 to the cytoskeletal pool as well as for desmosome maturation towards a Ca(2+)-insensitive state. Our data provide new insights into the role of E-cadherin and the contribution of Src signaling for formation and maintenance of desmosomal junctions.

  4. Identification and characterization of RBM44 as a novel intercellular bridge protein.

    PubMed

    Iwamori, Tokuko; Lin, Yi-Nan; Ma, Lang; Iwamori, Naoki; Matzuk, Martin M

    2011-02-25

    Intercellular bridges are evolutionarily conserved structures that connect differentiating germ cells. We previously reported the identification of TEX14 as the first essential intercellular bridge protein, the demonstration that intercellular bridges are required for male fertility, and the finding that intercellular bridges utilize components of the cytokinesis machinery to form. Herein, we report the identification of RNA binding motif protein 44 (RBM44) as a novel germ cell intercellular bridge protein. RBM44 was identified by proteomic analysis after intercellular bridge enrichment using TEX14 as a marker protein. RBM44 is highly conserved between mouse and human and contains an RNA recognition motif of unknown function. RBM44 mRNA is enriched in testis, and immunofluorescence confirms that RBM44 is an intercellular bridge component. However, RBM44 only partially localizes to TEX14-positive intercellular bridges. RBM44 is expressed most highly in pachytene and secondary spermatocytes, but disappears abruptly in spermatids. We discovered that RBM44 interacts with itself and TEX14 using yeast two-hybrid, mammalian two-hybrid, and immunoprecipitation. To define the in vivo function of RBM44, we generated a targeted deletion of Rbm44 in mice. Rbm44 null male mice produce somewhat increased sperm, and show enhanced fertility of unknown etiology. Thus, although RBM44 localizes to intercellular bridges during meiosis, RBM44 is not required for fertility in contrast to TEX14.

  5. [Gap junctions: A new therapeutic target in major depressive disorder?].

    PubMed

    Sarrouilhe, D; Dejean, C

    2015-11-01

    Major depressive disorder is a multifactorial chronic and debilitating mood disease with high lifetime prevalence and is associated with excess mortality, especially from cardiovascular diseases and through suicide. The treatments of this disease with tricyclic antidepressants and monoamine oxidase inhibitors are poorly tolerated and those that selectively target serotonin and norepinephrine re-uptake are not effective in all patients, showing the need to find new therapeutic targets. Post-mortem studies of brains from patients with major depressive disorders described a reduced expression of the gap junction-forming membrane proteins connexin 30 and connexin 43 in the prefrontal cortex and the locus coeruleus. The use of chronic unpredictable stress, a rodent model of depression, suggests that astrocytic gap junction dysfunction contributes to the pathophysiology of major depressive disorder. Chronic treatments of rats with fluoxetine and of rat cultured cortical astrocytes with amitriptyline support the hypothesis that the upregulation of gap junctional intercellular communication between brain astrocytes could be a novel mechanism for the therapeutic effect of antidepressants. In conclusion, astrocytic gap junctions are emerging as a new potential therapeutic target for the treatment of patients with major depressive disorder.

  6. The psychostimulant modafinil enhances gap junctional communication in cortical astrocytes.

    PubMed

    Liu, Xinhe; Petit, Jean-Marie; Ezan, Pascal; Gyger, Joël; Magistretti, Pierre; Giaume, Christian

    2013-12-01

    Sleep-wake cycle is characterized by changes in neuronal network activity. However, for the last decade there is increasing evidence that neuroglial interaction may play a role in the modulation of sleep homeostasis and that astrocytes have a critical impact in this process. Interestingly, astrocytes are organized into communicating networks based on their high expression of connexins, which are the molecular constituents of gap junction channels. Thus, neuroglial interactions should also be considered as the result of the interplay between neuronal and astroglial networks. Here, we investigate the effect of modafinil, a wakefulness-promoting agent, on astrocyte gap junctional communication. We report that in the cortex modafinil injection increases the expression of mRNA and protein of connexin 30 but not those of connexin 43, the other major astroglial connexin. These increases are correlated with an enhancement of intercellular dye coupling in cortical astrocytes, which is abolished when neuronal activity is silenced by tetrodotoxin. Moreover, gamma-hydroxybutyric acid, which at a millimolar concentration induces sleep, has an opposite effect on astroglial gap junctions in an activity-independent manner. These results support the proposition that astroglia may play an important role in complex physiological brain functions, such as sleep regulation, and that neuroglial networking interaction is modified during sleep-wake cycle. This article is part of the Special Issue Section entitled 'Current Pharmacology of Gap Junction Channels and Hemichannels'.

  7. The performances of silicon solar cell with core-shell p-n junctions of micro-nano pillars fabricated by cesium chloride self-assembly and dry etching

    NASA Astrophysics Data System (ADS)

    Liu, Jing; Zhang, Xinshuai; Dong, Gangqiang; Liao, Yuanxun; Wang, Bo; Zhang, Tianchong; Yi, Futing

    2014-03-01

    Silicon micro-nano pillars are cost-efficiently integrated using twice cesium chloride (CsCl) islands lithography technique and dry etching for solar cell applications. The micro PMMA islands are fabricated by inductively coupled plasma (ICP) dry etching with micro CsCl islands as masks, and the nano CsCl islands with nano sizes then are made on the surface of micro PMMA islands and silicon. By ICP dry etching with the mask of micro PMMA islands and nano CsCl islands, the micro-nano silicon pillars are made and certain height micro pillars are randomly positioned between dense arrays of nano pillars with different morphologies by controlling etching conditions. With 300 nm depth p-n junction detected by secondary-ion mass spectrometry (SIMS), the micro pillars of the diameter about 1 μm form the core-shell p-n junction to maximize utility of p-n junction interface and enable efficient free carrier collection, and the nano tapered pillars of 150 nm diameter are used to decrease reflection by a graded-refractive-index. Compared to single micro or nano pillar arrayed cells, the co-integrated solar cell with micro and nano pillars demonstrates improved photovoltaic characteristic that is a photovoltaic conversion efficiency (PCE) of 15.35 % with a short circuit current density ( J sc) of 38.40 mA/cm2 and an open circuit voltage ( V oc) of 555.7 mV, which benefits from the advantages of micro-nano pillar structures and can be further improved upon process optimization.

  8. Formation of adherens junctions leads to the emergence of a tissue-level tension in epithelial monolayers

    PubMed Central

    Harris, Andrew R.; Daeden, Alicia; Charras, Guillaume T.

    2014-01-01

    ABSTRACT Adherens junctions and desmosomes integrate the cytoskeletons of adjacent cells into a mechanical syncitium. In doing so, intercellular junctions endow tissues with the strength needed to withstand the mechanical stresses encountered in normal physiology and to coordinate tension during morphogenesis. Though much is known about the biological mechanisms underlying junction formation, little is known about how tissue-scale mechanical properties are established. Here, we use deep atomic force microscopy (AFM) indentation to measure the apparent stiffness of epithelial monolayers reforming from dissociated cells and examine which cellular processes give rise to tissue-scale mechanics. We show that the formation of intercellular junctions coincided with an increase in the apparent stiffness of reforming monolayers that reflected the generation of a tissue-level tension. Tension rapidly increased, reaching a maximum after 150 min, before settling to a lower level over the next 3 h as monolayers established homeostasis. The emergence of tissue tension correlated with the formation of adherens junctions but not desmosomes. As a consequence, inhibition of any of the molecular mechanisms participating in adherens junction initiation, remodelling and maturation significantly impeded the emergence of tissue-level tension in monolayers. PMID:24659804

  9. Quantum junction solar cells.

    PubMed

    Tang, Jiang; Liu, Huan; Zhitomirsky, David; Hoogland, Sjoerd; Wang, Xihua; Furukawa, Melissa; Levina, Larissa; Sargent, Edward H

    2012-09-12

    Colloidal quantum dot solids combine convenient solution-processing with quantum size effect tuning, offering avenues to high-efficiency multijunction cells based on a single materials synthesis and processing platform. The highest-performing colloidal quantum dot rectifying devices reported to date have relied on a junction between a quantum-tuned absorber and a bulk material (e.g., TiO(2)); however, quantum tuning of the absorber then requires complete redesign of the bulk acceptor, compromising the benefits of facile quantum tuning. Here we report rectifying junctions constructed entirely using inherently band-aligned quantum-tuned materials. Realizing these quantum junction diodes relied upon the creation of an n-type quantum dot solid having a clean bandgap. We combine stable, chemically compatible, high-performance n-type and p-type materials to create the first quantum junction solar cells. We present a family of photovoltaic devices having widely tuned bandgaps of 0.6-1.6 eV that excel where conventional quantum-to-bulk devices fail to perform. Devices having optimal single-junction bandgaps exhibit certified AM1.5 solar power conversion efficiencies of 5.4%. Control over doping in quantum solids, and the successful integration of these materials to form stable quantum junctions, offers a powerful new degree of freedom to colloidal quantum dot optoelectronics.

  10. Acidic pH of the lateral intercellular spaces of MDCK cells cultured on permeable supports.

    PubMed

    Chatton, J Y; Spring, K R

    1994-06-01

    The pH of the lateral intercellular space (LIS) of Madin-Darby canine kidney (MDCK) cell monolayers grown on permeable supports was investigated by microspectrofluorimetry using BCECF (2',7'-bis(carboxyethyl)-5,6-carboxyfluorescein). The permeability of the support was selectively reduced by growing Zn-Al-silicate crystals inside its pores. The diffusion of BCECF across the filter was sufficiently retarded to allow measurements of fluorescence in the LIS. The LIS pH and intracellular pH of the cells surrounding them were determined in HEPES-buffered solutions. When the perfusate pH was 7.4, the LIS pH was more acidic (7.06 +/- 0.02) and equaled the cytoplasmic pH (7.08 +/- 0.05). When perfusate was changed to pH 7.0 or 7.8, the LIS changed linearly by about half the magnitude of the perfusate pH. Intracellular pH followed LIS pH variations between perfusate pH 7.0 and 7.4 but was significantly higher when perfusate pH was 7.8. Tight junctional H+ permeability was undetectably low. The low steady-state pH in the LIS was not altered by inhibitors of acid transport or low temperature. Rapid perturbations of pH in the LIS showed that protons were not immobilized in the LIS. The acidic microenvironment within the LIS may be the result of buffering by the cell surface proteins.

  11. Metal-free molecular junctions on ITO via amino-silane binding-towards optoelectronic molecular junctions.

    PubMed

    Sergani, S; Furmansky, Y; Visoly-Fisher, I

    2013-11-15

    Light control over currents in molecular junctions is desirable as a non-contact input with high spectral and spatial resolution provided by the photonic input and the molecular electronics element, respectively. Expanding the study of molecular junctions to non-metallic transparent substrates, such as indium tin oxide (ITO), is vital for the observation of molecular optoelectronic effects. Non-metallic electrodes are expected to decrease the probability of quenching of molecular photo-excited states, light-induced plasmonic effects, or significant electrode expansion under visible light. We have developed micron-sized, metal free, optically addressable ITO molecular junctions with a conductive polymer serving as the counter-electrode. The electrical transport was shown to be dominated by the nature of the self-assembled monolayer (SAM). The use of amino-silane (APTMS) as the chemical binding scheme to ITO was found to be significant in determining the transport properties of the junctions. APTMS allows high junction yields and the formation of dense molecular layers preventing electrical short. However, polar amino-silane binding to the ITO significantly decreased the conductance compared to thiol-bound SAMs, and caused tilted geometry and disorder in the molecular layer. As the effect of the molecular structure on transport properties is clearly observed in our junctions, such metal-free junctions are suitable for characterizing the optoelectronic properties of molecular junctions.

  12. Metal-free molecular junctions on ITO via amino-silane binding—towards optoelectronic molecular junctions

    NASA Astrophysics Data System (ADS)

    Sergani, S.; Furmansky, Y.; Visoly-Fisher, I.

    2013-11-01

    Light control over currents in molecular junctions is desirable as a non-contact input with high spectral and spatial resolution provided by the photonic input and the molecular electronics element, respectively. Expanding the study of molecular junctions to non-metallic transparent substrates, such as indium tin oxide (ITO), is vital for the observation of molecular optoelectronic effects. Non-metallic electrodes are expected to decrease the probability of quenching of molecular photo-excited states, light-induced plasmonic effects, or significant electrode expansion under visible light. We have developed micron-sized, metal free, optically addressable ITO molecular junctions with a conductive polymer serving as the counter-electrode. The electrical transport was shown to be dominated by the nature of the self-assembled monolayer (SAM). The use of amino-silane (APTMS) as the chemical binding scheme to ITO was found to be significant in determining the transport properties of the junctions. APTMS allows high junction yields and the formation of dense molecular layers preventing electrical short. However, polar amino-silane binding to the ITO significantly decreased the conductance compared to thiol-bound SAMs, and caused tilted geometry and disorder in the molecular layer. As the effect of the molecular structure on transport properties is clearly observed in our junctions, such metal-free junctions are suitable for characterizing the optoelectronic properties of molecular junctions.

  13. META2: Intercellular DNA Methylation Pairwise Annotation and Integrative Analysis

    PubMed Central

    2016-01-01

    Genome-wide deciphering intercellular differential DNA methylation as well as its roles in transcriptional regulation remains elusive in cancer epigenetics. Here we developed a toolkit META2 for DNA methylation annotation and analysis, which aims to perform integrative analysis on differentially methylated loci and regions through deep mining and statistical comparison methods. META2 contains multiple versatile functions for investigating and annotating DNA methylation profiles. Benchmarked with T-47D cell, we interrogated the association within differentially methylated CpG (DMC) and region (DMR) candidate count and region length and identified major transition zones as clues for inferring statistically significant DMRs; together we validated those DMRs with the functional annotation. Thus META2 can provide a comprehensive analysis approach for epigenetic research and clinical study. PMID:28116291

  14. Does Pseudomonas aeruginosa use intercellular signalling to build biofilm communities?

    PubMed

    Kirisits, Mary Jo; Parsek, Matthew R

    2006-12-01

    Pseudomonas aeruginosa is a Gram-negative bacterial species that causes several opportunistic human infections. This organism is also found in the environment, where it is renowned (like other Pseudomonads) for its ability to use a wide variety of compounds as carbon and energy sources. It is a model species for studying group-related behaviour in bacteria. Two types of group behaviour it engages in are intercellular signalling, or quorum sensing, and the formation of surface-associated communities called biofilms. Both quorum sensing and biofilm formation are important in the pathogenesis of P. aeruginosa infections. Quorum sensing regulates the expression of several secreted virulence factors and quorum sensing mutant strains are attenuated for virulence in animal models. Biofilms have been implicated in chronic infections. Two examples are the chronic lung infections afflicting people suffering from cystic fibrosis and colonization of indwelling medical devices. This review will discuss quorum sensing and biofilm formation and studies that link these two processes.

  15. Extracellular vesicles shuffling intercellular messages: for good or for bad.

    PubMed

    Lo Cicero, Alessandra; Stahl, Philip D; Raposo, Graça

    2015-08-01

    The release of extracellular vesicles (EVs) is a highly conserved process exploited by diverse organisms as a mode of intercellular communication. Vesicles of sizes ranging from 30 to 1000nm, or even larger, are generated by blebbing of the plasma membrane (microvesicles) or formed in multivesicular endosomes (MVEs) to be secreted by exocytosis as exosomes. Exosomes, microvesicles and other EVs contain membrane and cytosolic components that include proteins, lipids and RNAs, a composition that differs related to their site of biogenesis. Several mechanisms are involved in vesicle formation at the plasma membrane or in endosomes, which is reflected in their heterogeneity, size and composition. EVs have significant promise for therapeutics and diagnostics and for understanding physiological and pathological processes all of which have boosted research to find modulators of their composition, secretion and targeting.

  16. [Role of the Notch receptors in intercellular communication].

    PubMed

    Seugnet, L; Simpson, P

    1996-04-01

    The Notch gene was discovered in Drosophila at the beginning of the century and is currently the subject of intensive investigation, not only in invertebrates but also in vertebrates where remarkably well conserved homologues have been recently found. Notch encodes a new kind of cellular receptor whose functioning is still unclear and plays a role in a large number of cell interactions throughout development and in tissue renewal in the adult. Detailed study in invertebrates of some of these interactions has led to the identification of other genes required for transduction of the signal initiated by the receptor. Notch is always involved in processes where cells have the potential to choose between several different programmes of differentiation. Cells adopt a specific developmental pathway as a result of the inhibition of some programmes through Notch signalling. In this review we discuss the contribution of different experimental models to an understanding of the role of Notch in intercellular signalling.

  17. Identification and Characterization of Tunneling Nanotubes for Intercellular Trafficking.

    PubMed

    Abounit, Saïda; Delage, Elise; Zurzolo, Chiara

    2015-06-01

    Tunneling nanotubes (TNTs) are thin membranous channels providing direct cytoplasmic connection between remote cells. They are commonly observed in different cell cultures and increasing evidence supports their role in intercellular communication and pathogen transfer. However, the study of TNTs presents several pitfalls (e.g., difficulty in preserving such delicate structures, possible confusion with other protrusions, structural and functional heterogeneity, etc.) and therefore requires thoroughly designed approaches. The methods described in this unit represent a guideline for the characterization of TNTs (or TNT-like structures) in cell culture. Specifically, optimized protocols to (1) identify TNTs and the cytoskeletal elements present inside them; (2) evaluate TNT frequency in cell culture; (3) unambiguously distinguish them from other cellular connections or protrusions; and (4) monitor their formation in living cells are provided. Finally, this unit describes how to assess TNT-mediated cell-to-cell transfer of cellular components, which is a fundamental criterion for identifying functional TNTs.

  18. Structure of the tight junctions of the human eccrine sweat gland.

    PubMed

    Briggman, J V; Bank, H L; Bigelow, J B; Graves, J S; Spicer, S S

    1981-12-01

    The human eccrine sweat gland contains two anatomically and functionally discrete segments: the secretory coil, which produces an isotonic or slightly hypertonic precursor fluid, and the coiled duct, which reabsorbs Na+ and Cl- to yield a hypotonic sweat. We examined the freeze-fracture morphology of tight junctions from isolated secretory coil and coiled duct segments to assess indirectly the contribution of paracellular ion transport in secretion and resorption in the sweat gland. In the secretory coil, tight junctions of the intercellular canaliculus and main lumen consisted of approximately 9 and 6, closely spaced, parallel or anastomosing elements, respectively. Tight junctions of the coiled duct were similar in appearance to those at the main lumen of the secretory coil. In both the secretory coil and coiled duct, and average of 2 to 3, widely spaced junctional elements were usually observed basolateral to the closely spaced junctional elements in the region corresponding to the location of the zonula adherens in Epon sections. The complexity of the tight junctions of the secretory coil exceeded what we expected for an epithelium secreting an isosmotic fluid. The elaborate tight junctions of the coiled duct support other evidence for an intermediate to high transepithelial resistance.

  19. Role of tight junction proteins in gastroesophageal reflux disease

    PubMed Central

    2012-01-01

    Background Gastroesophageal reflux disease (GERD) is associated with impaired epithelial barrier function that is regulated by cell-cell contacts. The aim of the study was to investigate the expression pattern of selected components involved in the formation of tight junctions in relation to GERD. Methods Eighty-four patients with GERD-related symptoms with endoscopic signs (erosive: n = 47) or without them (non-erosive: n = 37) as well as 26 patients lacking GERD-specific symptoms as controls were included. Endoscopic and histological characterization of esophagitis was performed according to the Los Angeles and adapted Ismeil-Beigi criteria, respectively. Mucosal biopsies from distal esophagus were taken for analysis by histopathology, immunohistochemistry and quantitative reverse-transcription polymerase chain reaction (RT-PCR) of five genes encoding tight junction components [Occludin, Claudin-1, -2, Zona occludens (ZO-1, -2)]. Results Histopathology confirmed GERD-specific alterations as dilated intercellular spaces in the esophageal mucosa of patients with GERD compared to controls (P < 0.05). Claudin-1 and −2 were 2- to 6-fold upregulation on transcript (P < 0.01) and in part on protein level (P < 0.015) in GERD, while subgroup analysis of revealed this upregulation for ERD only. In both erosive and non-erosive reflux disease, expression levels of Occludin and ZO-1,-2 were not significantly affected. Notably, the induced expression of both claudins did not correlate with histopathological parameters (basal cell hyperplasia, dilated intercellular spaces) in patients with GERD. Conclusions Taken together, the missing correlation between the expression of tight junction-related components and histomorphological GERD-specific alterations does not support a major role of the five proteins studied in the pathogenesis of GERD. PMID:22994974

  20. Catalase protects tumor cells from apoptosis induction by intercellular ROS signaling.

    PubMed

    Bechtel, Wibke; Bauer, Georg

    2009-11-01

    Transformed cells are subject to intercellular induction of apoptosis by neighbouring nontransformed cells and to autocrine apoptotic self-destruction. Both processes depend on extracellular superoxide anion generation by the transformed cells and on the release of peroxidase from both nontransformed and transformed cells. This concerted action results in HOCl synthesis, HOCl-superoxide anion interaction and generation of apoptosis-inducing hydroxyl radicals. In contrast to transformed cells, ex vivo tumor cells are resistant against intercellular induction of apoptosis and autocrine apoptotic self-destruction. Resistance of tumor cells against intercellular ROS signaling depends on interference through catalase expression on the membrane. Intercellular ROS signaling of tumor cells can be restored when i) exogenous HOCl is added; ii) exogenous hydrogen peroxide is supplied, or iii) catalase is inhibited. These findings define the biochemical basis for specific apoptosis induction in tumor cells through re-establishment of intercellular ROS signaling, a potential novel approach in tumor prevention and therapy.

  1. Nuclear–cytoskeletal linkages facilitate cross talk between the nucleus and intercellular adhesions

    PubMed Central

    Stewart, Rachel M.; Zubek, Amanda E.; Rosowski, Kathryn A.; Schreiner, Sarah M.

    2015-01-01

    The linker of nucleoskeleton and cytoskeleton (LINC) complex allows cells to actively control nuclear position by coupling the nucleus to the cytoplasmic cytoskeleton. Nuclear position responds to the formation of intercellular adhesions through coordination with the cytoskeleton, but it is not known whether this response impacts adhesion function. In this paper, we demonstrate that the LINC complex component SUN2 contributes to the mechanical integrity of intercellular adhesions between mammalian epidermal keratinocytes. Mice deficient for Sun2 exhibited irregular hair follicle intercellular adhesions, defective follicle structure, and alopecia. Primary mouse keratinocytes lacking Sun2 displayed aberrant nuclear position in response to adhesion formation, altered desmosome distribution, and mechanically defective adhesions. This dysfunction appeared rooted in a failure of Sun2-null cells to reorganize their microtubule network to support coordinated intercellular adhesion. Together, these results suggest that cross talk between the nucleus, cytoskeleton, and intercellular adhesions is important for epidermal tissue integrity. PMID:25963820

  2. Four-junction superconducting circuit

    NASA Astrophysics Data System (ADS)

    Qiu, Yueyin; Xiong, Wei; He, Xiao-Ling; Li, Tie-Fu; You, J. Q.

    2016-06-01

    We develop a theory for the quantum circuit consisting of a superconducting loop interrupted by four Josephson junctions and pierced by a magnetic flux (either static or time-dependent). In addition to the similarity with the typical three-junction flux qubit in the double-well regime, we demonstrate the difference of the four-junction circuit from its three-junction analogue, including its advantages over the latter. Moreover, the four-junction circuit in the single-well regime is also investigated. Our theory provides a tool to explore the physical properties of this four-junction superconducting circuit.

  3. Dot junction solar cells

    NASA Technical Reports Server (NTRS)

    Daud, T.; Crotty, G. T.

    1986-01-01

    A design of solar cells with reduced junction area on the cell surface is investigated for reduction of saturation current and increase in open-circuit voltage. Equidiameter dot junctions distributed across the surface of the cell offer an efficient alternative, with variations in dot diameter and in the spacing between dots giving the required variations in the ratio of junction area to total surface area. A simplified analysis for short-circuit current and other cell parameters, which enables cell design optimization, is presented. Experimental solar-cell performance results, as functions of different area ratios, are presented and compared with the model. It is shown that saturation current reduction is possible for achieving efficiencies as high as 18 percent in flat-plate terrestrial applications.

  4. Perspective: Thermal and thermoelectric transport in molecular junctions

    NASA Astrophysics Data System (ADS)

    Cui, Longji; Miao, Ruijiao; Jiang, Chang; Meyhofer, Edgar; Reddy, Pramod

    2017-03-01

    With the advent of molecular electronics, tremendous attention has been paid towards understanding the structure-function relationship of molecular junctions. Understanding how heat is transported, dissipated, and converted into electricity in molecular junctions is of great importance for designing thermally robust molecular circuits and high-performance energy conversion devices. Further, the study of thermal and thermoelectric phenomena in molecular junctions provides novel insights into the limits of applicability of classical laws. Here, we present a review of the computational and experimental progress made in probing thermoelectric effects, thermal conduction, heat dissipation, and local heating/cooling in self-assembled monolayer and single molecule junctions. We also discuss some outstanding challenges and potential future directions.

  5. 16. VIEW OF JUNCTION BETWEEN CABLE CHASE AND SHIELDING TANK. ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    16. VIEW OF JUNCTION BETWEEN CABLE CHASE AND SHIELDING TANK. SHOWS CABLES AND LINES IN THE TRENCH, POLE OF FRAME ASSEMBLY, AND EQUIPMENT IN CONCRETE BOX ADJACENT TO CABLE CHASE. INEL PHOTO NUMBER 65-6178, TAKEN NOVEMBER 10, 1965. - Idaho National Engineering Laboratory, Advanced Reentry Vehicle Fusing System, Scoville, Butte County, ID

  6. Epithelial junctions and Rho family GTPases: the zonular signalosome

    PubMed Central

    Citi, Sandra; Guerrera, Diego; Spadaro, Domenica; Shah, Jimit

    2014-01-01

    The establishment and maintenance of epithelial cell-cell junctions is crucially important to regulate adhesion, apico-basal polarity and motility of epithelial cells, and ultimately controls the architecture and physiology of epithelial organs. Junctions are supported, shaped and regulated by cytoskeletal filaments, whose dynamic organization and contractility are finely tuned by GTPases of the Rho family, primarily RhoA, Rac1 and Cdc42. Recent research has identified new molecular mechanisms underlying the cross-talk between these GTPases and epithelial junctions. Here we briefly summarize the current knowledge about the organization, molecular evolution and cytoskeletal anchoring of cell-cell junctions, and we comment on the most recent advances in the characterization of the interactions between Rho GTPases and junctional proteins, and their consequences with regards to junction assembly and regulation of cell behavior in vertebrate model systems. The concept of “zonular signalosome” is proposed, which highlights the close functional relationship between proteins of zonular junctions (zonulae occludentes and adhaerentes) and the control of cytoskeletal organization and signaling through Rho GTPases, transcription factors, and their effectors. PMID:25483301

  7. Regulation of Hemichannels and Gap Junction Channels by Cytokines in Antigen-Presenting Cells

    PubMed Central

    Shoji, Kenji F.; Aguirre, Adam; Sáez, Juan C.

    2014-01-01

    Autocrine and paracrine signals coordinate responses of several cell types of the immune system that provide efficient protection against different challenges. Antigen-presenting cells (APCs) coordinate activation of this system via homocellular and heterocellular interactions. Cytokines constitute chemical intercellular signals among immune cells and might promote pro- or anti-inflammatory effects. During the last two decades, two membrane pathways for intercellular communication have been demonstrated in cells of the immune system. They are called hemichannels (HCs) and gap junction channels (GJCs) and provide new insights into the mechanisms of the orchestrated response of immune cells. GJCs and HCs are permeable to ions and small molecules, including signaling molecules. The direct intercellular transfer between contacting cells can be mediated by GJCs, whereas the release to or uptake from the extracellular milieu can be mediated by HCs. GJCs and HCs can be constituted by two protein families: connexins (Cxs) or pannexins (Panxs), which are present in almost all APCs, being Cx43 and Panx1 the most ubiquitous members of each protein family. In this review, we focus on the effects of different cytokines on the intercellular communication mediated by HCs and GJCs in APCs and their impact on purinergic signaling. PMID:25301274

  8. Roles of gap junctions, connexins, and pannexins in epilepsy

    PubMed Central

    Mylvaganam, Shanthini; Ramani, Meera; Krawczyk, Michal; Carlen, Peter L.

    2014-01-01

    Enhanced gap junctional communication (GJC) between neurons is considered a major factor underlying the neuronal synchrony driving seizure activity. In addition, the hippocampal sharp wave ripple complexes, associated with learning and seizures, are diminished by GJC blocking agents. Although gap junctional blocking drugs inhibit experimental seizures, they all have other non-specific actions. Besides interneuronal GJC between dendrites, inter-axonal and inter-glial GJC is also considered important for seizure generation. Interestingly, in most studies of cerebral tissue from animal seizure models and from human patients with epilepsy, there is up-regulation of glial, but not neuronal gap junctional mRNA and protein. Significant changes in the expression and post-translational modification of the astrocytic connexin Cx43, and Panx1 were observed in an in vitro Co++ seizure model, further supporting a role for glia in seizure-genesis, although the reasons for this remain unclear. Further suggesting an involvement of astrocytic GJC in epilepsy, is the fact that the expression of astrocytic Cx mRNAs (Cxs 30 and 43) is several fold higher than that of neuronal Cx mRNAs (Cxs 36 and 45), and the number of glial cells outnumber neuronal cells in mammalian hippocampal and cortical tissue. Pannexin expression is also increased in both animal and human epileptic tissues. Specific Cx43 mimetic peptides, Gap 27 and SLS, inhibit the docking of astrocytic connexin Cx43 proteins from forming intercellular gap junctions (GJs), diminishing spontaneous seizures. Besides GJs, Cx membrane hemichannels in glia and Panx membrane channels in neurons and glia are also inhibited by traditional gap junctional pharmacological blockers. Although there is no doubt that connexin-based GJs and hemichannels, and pannexin-based membrane channels are related to epilepsy, the specific details of how they are involved and how we can modulate their function for therapeutic purposes remain to be

  9. Astrocytic gap junctional networks suppress cellular damage in an in vitro model of ischemia

    SciTech Connect

    Shinotsuka, Takanori; Yasui, Masato; Nuriya, Mutsuo

    2014-02-07

    Highlights: • Astrocytes exhibit characteristic changes in [Ca{sup 2+}]{sub i} under OGD. • Astrocytic [Ca{sup 2+}]{sub i} increase is synchronized with a neuronal anoxic depolarization. • Gap junctional couplings protect neurons as well as astrocytes during OGD. - Abstract: Astrocytes play pivotal roles in both the physiology and the pathophysiology of the brain. They communicate with each other via extracellular messengers as well as through gap junctions, which may exacerbate or protect against pathological processes in the brain. However, their roles during the acute phase of ischemia and the underlying cellular mechanisms remain largely unknown. To address this issue, we imaged changes in the intracellular calcium concentration ([Ca{sup 2+}]{sub i}) in astrocytes in mouse cortical slices under oxygen/glucose deprivation (OGD) condition using two-photon microscopy. Under OGD, astrocytes showed [Ca{sup 2+}]{sub i} oscillations followed by larger and sustained [Ca{sup 2+}]{sub i} increases. While the pharmacological blockades of astrocytic receptors for glutamate and ATP had no effect, the inhibitions of gap junctional intercellular coupling between astrocytes significantly advanced the onset of the sustained [Ca{sup 2+}]{sub i} increase after OGD exposure. Interestingly, the simultaneous recording of the neuronal membrane potential revealed that the onset of the sustained [Ca{sup 2+}]{sub i} increase in astrocytes was synchronized with the appearance of neuronal anoxic depolarization. Furthermore, the blockade of gap junctional coupling resulted in a concurrent faster appearance of neuronal depolarizations, which remain synchronized with the sustained [Ca{sup 2+}]{sub i} increase in astrocytes. These results indicate that astrocytes delay the appearance of the pathological responses of astrocytes and neurons through their gap junction-mediated intercellular network under OGD. Thus, astrocytic gap junctional networks provide protection against tissue damage

  10. Turbine vane plate assembly

    DOEpatents

    Schiavo Jr., Anthony L.

    2006-01-10

    A turbine vane assembly includes a turbine vane having first and second shrouds with an elongated airfoil extending between. Each end of the airfoil transitions into a shroud at a respective junction. Each of the shrouds has a plurality of cooling passages, and the airfoil has a plurality of cooling passages extending between the first and second shrouds. A substantially flat inner plate and an outer plate are coupled to each of the first and second shrouds so as to form inner and outer plenums. Each inner plenum is defined between at least the junction and the substantially flat inner plate; each outer plenum is defined between at least the substantially flat inner plate and the outer plate. Each inner plenum is in fluid communication with a respective outer plenum through at least one of the cooling passages in the respective shroud.

  11. Contribution of Folate Biosynthesis to Ralstonia solanacearum Proliferation in Intercellular Spaces

    PubMed Central

    Shinohara, Rena; Kanda, Ayami; Ohnishi, Kouhei; Kiba, Akinori; Hikichi, Yasufumi

    2005-01-01

    The vigorous proliferation of Ralstonia solanacearum OE1-1 in host intercellular spaces after the invasion of host plants is necessary for the virulence of this bacterium. A folate auxotroph, RM, in which a mini-Tn5 transposon was inserted into pabB encoding para-aminobenzoate synthase component I, lost its ability to vigorously proliferate in intercellular spaces along with its systemic infectivity and virulence after inoculation into roots and infiltration into leaves of tobacco plants. Complementation of RM with the pabB gene allowed the mutant to multiply in intercellular spaces and to cause disease. In tobacco plants that were pretreated with folate, RM was able to vigorously proliferate in the intercellular spaces and cause disease. Interestingly, when it was inoculated through cut stems, the mutant multiplied in the plants and was virulent. Moreover, the mutant multiplied well in stem fluids but not in intercellular fluids, suggesting that the folate concentration within intercellular spaces may be a limiting factor for bacterial proliferation. Therefore, folate biosynthesis contributes to the vigorous proliferation of bacteria in intercellular spaces and leads to systemic infectivity resulting in virulence. PMID:15640216

  12. Exosomes Mediate the Intercellular Communication after Myocardial Infarction

    PubMed Central

    Yuan, Ming-Jie; Maghsoudi, Taneen; Wang, Tao

    2016-01-01

    The mechanisms of cardiac repair after myocardial infarction (MI) are complicated and not well-understood currently. It is known that exosomes are released from most cells, recognized as new candidates with important roles in intercellular and tissue-level communication. Cells can package proteins and RNA messages into exosome and secret to recipient cells, which regulate gene expression in recipient cells. The research on exosomes in cardiovascular disease is just emerging. It is well-known that exosomes from cardiomyocyte can transfect endothelial cells, stem cells, fibroblasts and smooth muscle cells to induce cellular changes. After myocardial infarction (MI), the exosomes play important roles in local and distant microcommunication. Nowadays, exosomal microRNAs transportation has been found to deliver signals to mediate cardiac repair after MI. However, the exosomes quality and quantities are variable under different pathological conditions. Therefore, we speculate that the monitoring of the quality and quantity of exosomes may serve as diagnosis and prognosis biomarkers of MI, and the study of exosomes will provide insights for the new therapeutics to cardiac remodeling after MI. PMID:26941569

  13. Reconfigurable microfluidics with integrated aptasensors for monitoring intercellular communication.

    PubMed

    Kwa, Timothy; Zhou, Qing; Gao, Yandong; Rahimian, Ali; Kwon, Lydia; Liu, Ying; Revzin, Alexander

    2014-05-21

    We report the development of a microsystem integrating anti-TNF-α aptasensors with vacuum-actuatable microfluidic devices that may be used to monitor intercellular communications. Actuatable chambers were used to expose to mitogen a group of ~600 cells while not stimulating another group of monocytes only 600 μm away. Co-localizing groups of cells with miniature 300 μm diameter aptamer-modified electrodes enabled monitoring of TNF-α release from each group independently. The microsystem allowed observation of the sequence of events that included 1) mitogenic activation of the first group of monocytes to produce TNF-α, 2) diffusion of TNF-α to the location of the second group of cells and 3) activation of the second group of cells resulting in the production of TNF-α by these cells. Thus, we were able to experimentally verify reciprocal paracrine crosstalk between the two groups of cells secreting the same signalling molecule. Given the prevalence of such cellular communications during injury, cancer or immune response and the dearth of available monitoring techniques, the microsystem described here is envisioned to have significant impact on cell biology.

  14. Exosomes Mediate the Intercellular Communication after Myocardial Infarction.

    PubMed

    Yuan, Ming-Jie; Maghsoudi, Taneen; Wang, Tao

    2016-01-01

    The mechanisms of cardiac repair after myocardial infarction (MI) are complicated and not well-understood currently. It is known that exosomes are released from most cells, recognized as new candidates with important roles in intercellular and tissue-level communication. Cells can package proteins and RNA messages into exosome and secret to recipient cells, which regulate gene expression in recipient cells. The research on exosomes in cardiovascular disease is just emerging. It is well-known that exosomes from cardiomyocyte can transfect endothelial cells, stem cells, fibroblasts and smooth muscle cells to induce cellular changes. After myocardial infarction (MI), the exosomes play important roles in local and distant microcommunication. Nowadays, exosomal microRNAs transportation has been found to deliver signals to mediate cardiac repair after MI. However, the exosomes quality and quantities are variable under different pathological conditions. Therefore, we speculate that the monitoring of the quality and quantity of exosomes may serve as diagnosis and prognosis biomarkers of MI, and the study of exosomes will provide insights for the new therapeutics to cardiac remodeling after MI.

  15. Plasmodesmata: channels for intercellular signaling during plant growth and development.

    PubMed

    Sevilem, Iris; Yadav, Shri Ram; Helariutta, Ykä

    2015-01-01

    Plants have evolved strategies for short- and long-distance communication to coordinate plant development and to adapt to changing environmental conditions. Plasmodesmata (PD) are intercellular nanochannels that provide an effective pathway for both selective and nonselective movement of various molecules that function in diverse biological processes. Numerous non-cell-autonomous proteins (NCAP) and small RNAs have been identified that have crucial roles in cell fate determination and organ patterning during development. Both the density and aperture size of PD are developmentally regulated, allowing formation of spatial symplastic domains for establishment of tissue-specific developmental programs. The PD size exclusion limit (SEL) is controlled by reversible deposition of callose, as well as by some PD-associated proteins. Although a large number of PD-associated proteins have been identified, many of their functions remain unknown. Despite the fact that PD are primarily membranous structures, surprisingly very little is known about their lipid composition. Thus, future studies in PD biology will provide deeper insights into the high-resolution structure and tightly regulated functions of PD and the evolution of PD-mediated cell-to-cell communication in plants.

  16. M-cadherin-mediated intercellular interactions activate satellite cell division.

    PubMed

    Marti, Merce; Montserrat, Núria; Pardo, Cristina; Mulero, Lola; Miquel-Serra, Laia; Rodrigues, Alexandre Miguel Cavaco; Andrés Vaquero, José; Kuebler, Bernd; Morera, Cristina; Barrero, María José; Izpisua Belmonte, Juan Carlos

    2013-11-15

    Adult muscle stem cells and their committed myogenic precursors, commonly referred to as the satellite cell population, are involved in both muscle growth after birth and regeneration after damage. It has been previously proposed that, under these circumstances, satellite cells first become activated, divide and differentiate, and only later fuse to the existing myofiber through M-cadherin-mediated intercellular interactions. Our data show that satellite cells fuse with the myofiber concomitantly to cell division, and only when the nuclei of the daughter cells are inside the myofiber, do they complete the process of differentiation. Here we demonstrate that M-cadherin plays an important role in cell-to-cell recognition and fusion, and is crucial for cell division activation. Treatment of satellite cells with M-cadherin in vitro stimulates cell division, whereas addition of anti-M-cadherin antibodies reduces the cell division rate. Our results suggest an alternative model for the contribution of satellite cells to muscle development, which might be useful in understanding muscle regeneration, as well as muscle-related dystrophies.

  17. Myosin VI facilitates connexin 43 gap junction accretion

    PubMed Central

    Waxse, Bennett J.

    2017-01-01

    ABSTRACT In this study, we demonstrate myosin VI enrichment at Cx43 (also known as GJA1)-containing gap junctions (GJs) in heart tissue, primary cardiomyocytes and cell culture models. In primary cardiac tissue and in fibroblasts from the myosin VI-null mouse as well as in tissue culture cells transfected with siRNA against myosin VI, we observe reduced GJ plaque size with a concomitant reduction in intercellular communication, as shown by fluorescence recovery after photobleaching (FRAP) and a new method of selective calcein administration. Analysis of the molecular role of myosin VI in Cx43 trafficking indicates that myosin VI is dispensable for the delivery of Cx43 to the cell surface and connexon movement in the plasma membrane. Furthermore, we cannot corroborate clathrin or Dab2 localization at gap junctions and we do not observe a function for the myosin-VI–Dab2 complex in clathrin-dependent endocytosis of annular gap junctions. Instead, we found that myosin VI was localized at the edge of Cx43 plaques by using total internal reflection fluorescence (TIRF) microscopy and use FRAP to identify a plaque accretion defect as the primary manifestation of myosin VI loss in Cx43 homeostasis. A fuller understanding of this derangement may explain the cardiomyopathy or gliosis associated with the loss of myosin VI. PMID:28096472

  18. Victory Junction Gang Camp

    ERIC Educational Resources Information Center

    Shell, Ryan

    2007-01-01

    This article describes the Victory Junction Gang Camp, a not-for-profit, NASCAR-themed camp for children with chronic medical conditions that serves 24 different disease groups. The mission of the camp is to give children life-changing camping experiences that are exciting, fun, and empowering in a safe and medically sound environment. While doing…

  19. Supramolecular Systems and Chemical Reactions in Single-Molecule Break Junctions.

    PubMed

    Li, Xiaohui; Hu, Duan; Tan, Zhibing; Bai, Jie; Xiao, Zongyuan; Yang, Yang; Shi, Jia; Hong, Wenjing

    2017-04-01

    The major challenges of molecular electronics are the understanding and manipulation of the electron transport through the single-molecule junction. With the single-molecule break junction techniques, including scanning tunneling microscope break junction technique and mechanically controllable break junction technique, the charge transport through various single-molecule and supramolecular junctions has been studied during the dynamic fabrication and continuous characterization of molecular junctions. This review starts from the charge transport characterization of supramolecular junctions through a variety of noncovalent interactions, such as hydrogen bond, π-π interaction, and electrostatic force. We further review the recent progress in constructing highly conductive molecular junctions via chemical reactions, the response of molecular junctions to external stimuli, as well as the application of break junction techniques in controlling and monitoring chemical reactions in situ. We suggest that beyond the measurement of single molecular conductance, the single-molecule break junction techniques provide a promising access to study molecular assembly and chemical reactions at the single-molecule scale.

  20. Heterotypic gap junctions at glutamatergic mixed synapses are abundant in goldfish brain

    PubMed Central

    Rash, John E.; Kamasawa, Naomi; Vanderpool, Kimberly G.; Yasumura, Thomas; O'Brien, John; Nannapaneni, Srikant; Pereda, Alberto E.; Nagy, James I.

    2014-01-01

    Gap junctions provide for direct intercellular electrical and metabolic coupling. The abundance of gap junctions at “large myelinated club ending” synapses on Mauthner cells of the teleost brain provided a convenient model to correlate anatomical and physiological properties of electrical synapses. There, presynaptic action potentials were found to evoke short-latency electrical “pre-potentials” immediately preceding their accompanying glutamate-induced depolarizations, making these the first unambiguously identified “mixed” (i.e., chemical plus electrical) synapses in the vertebrate CNS. We recently showed that gap junctions at these synapses exhibit asymmetric electrical resistance (i.e., electrical rectification), which we correlated with total molecular asymmetry of connexin composition in their apposing gap junction hemiplaques, with Cx35 restricted to axon terminal hemiplaques and Cx34.7 restricted to apposing Mauthner cell plasma membranes. We now show that similarly heterotypic neuronal gap junctions are abundant throughout goldfish brain, with labeling exclusively for Cx35 in presynaptic hemiplaques and exclusively for Cx34.7 in postsynaptic hemiplaques. Moreover, the vast majority of these asymmetric gap junctions occur at glutamatergic axon terminals. The widespread distribution of heterotypic gap junctions at glutamatergic mixed synapses throughout goldfish brain and spinal cord implies that pre- vs. postsynaptic asymmetry at electrical synapses evolved early in the chordate lineage. We propose that the advantages of the molecular and functional asymmetry of connexins at electrical synapses that are so prominently expressed in the teleost CNS are unlikely to have been abandoned in higher vertebrates. However, to create asymmetric coupling in mammals, where most gap junctions are composed of Cx36 on both sides, would require some other mechanism, such as differential phosphorylation of connexins on opposite sides of the same gap junction or

  1. Heterotypic gap junctions at glutamatergic mixed synapses are abundant in goldfish brain.

    PubMed

    Rash, J E; Kamasawa, N; Vanderpool, K G; Yasumura, T; O'Brien, J; Nannapaneni, S; Pereda, A E; Nagy, J I

    2015-01-29

    Gap junctions provide for direct intercellular electrical and metabolic coupling. The abundance of gap junctions at "large myelinated club ending (LMCE)" synapses on Mauthner cells (M-cells) of the teleost brain provided a convenient model to correlate anatomical and physiological properties of electrical synapses. There, presynaptic action potentials were found to evoke short-latency electrical "pre-potentials" immediately preceding their accompanying glutamate-induced depolarizations, making these the first unambiguously identified "mixed" (i.e., chemical plus electrical) synapses in the vertebrate CNS. We recently showed that gap junctions at these synapses exhibit asymmetric electrical resistance (i.e., electrical rectification), which we correlated with total molecular asymmetry of connexin composition in their apposing gap junction hemiplaques, with connexin35 (Cx35) restricted to axon terminal hemiplaques and connexin34.7 (Cx34.7) restricted to apposing M-cell plasma membranes. We now show that similarly heterotypic neuronal gap junctions are abundant throughout goldfish brain, with labeling exclusively for Cx35 in presynaptic hemiplaques and exclusively for Cx34.7 in postsynaptic hemiplaques. Moreover, the vast majority of these asymmetric gap junctions occur at glutamatergic axon terminals. The widespread distribution of heterotypic gap junctions at glutamatergic mixed synapses throughout goldfish brain and spinal cord implies that pre- vs. postsynaptic asymmetry at electrical synapses evolved early in the chordate lineage. We propose that the advantages of the molecular and functional asymmetry of connexins at electrical synapses that are so prominently expressed in the teleost CNS are unlikely to have been abandoned in higher vertebrates. However, to create asymmetric coupling in mammals, where most gap junctions are composed of connexin36 (Cx36) on both sides, would require some other mechanism, such as differential phosphorylation of connexins on

  2. Polysaccharide intercellular adhesin in biofilm: structural and regulatory aspects

    PubMed Central

    Arciola, Carla Renata; Campoccia, Davide; Ravaioli, Stefano; Montanaro, Lucio

    2015-01-01

    Staphylococcus aureus and Staphylococcus epidermidis are the leading etiologic agents of implant-related infections. Biofilm formation is the main pathogenetic mechanism leading to the chronicity and irreducibility of infections. The extracellular polymeric substances of staphylococcal biofilms are the polysaccharide intercellular adhesin (PIA), extracellular-DNA, proteins, and amyloid fibrils. PIA is a poly-β(1-6)-N-acetylglucosamine (PNAG), partially deacetylated, positively charged, whose synthesis is mediated by the icaADBC locus. DNA sequences homologous to ica locus are present in many coagulase-negative staphylococcal species, among which S. lugdunensis, however, produces a biofilm prevalently consisting of proteins. The product of icaA is an N-acetylglucosaminyltransferase that synthetizes PIA oligomers from UDP-N-acetylglucosamine. The product of icaD gives optimal efficiency to IcaA. The product of icaC is involved in the externalization of the nascent polysaccharide. The product of icaB is an N-deacetylase responsible for the partial deacetylation of PIA. The expression of ica locus is affected by environmental conditions. In S. aureus and S. epidermidis ica-independent alternative mechanisms of biofilm production have been described. S. epidermidis and S. aureus undergo to a phase variation for the biofilm production that has been ascribed, in turn, to the transposition of an insertion sequence in the icaC gene or to the expansion/contraction of a tandem repeat naturally harbored within icaC. A role is played by the quorum sensing system, which negatively regulates biofilm formation, favoring the dispersal phase that disseminates bacteria to new infection sites. Interfering with the QS system is a much debated strategy to combat biofilm-related infections. In the search of vaccines against staphylococcal infections deacetylated PNAG retained on the surface of S. aureus favors opsonophagocytosis and is a potential candidate for immune-protection. PMID

  3. Intercellular Adhesion Molecule 1 Knockout Abrogates Radiation Induced Pulmonary Inflammation

    NASA Astrophysics Data System (ADS)

    Hallahan, Dennis E.; Virudachalam, Subbulakshmi

    1997-06-01

    Increased expression of intercellular adhesion molecule 1 (ICAM-1; CD54) is induced by exposure to ionizing radiation. The lung was used as a model to study the role of ICAM-1 in the pathogenesis of the radiation-induced inflammation-like response. ICAM-1 expression increased in the pulmonary microvascular endothelium and not in the endothelium of larger pulmonary vessels following treatment of mice with thoracic irradiation. To quantify radiation-induced ICAM-1 expression, we utilized fluorescence-activated cell sorting analysis of anti-ICAM-1 antibody labeling of pulmonary microvascular endothelial cells from human cadaver donors (HMVEC-L cells). Fluorochrome conjugates and UV microscopy were used to quantify the fluorescence intensity of ICAM in the irradiated lung. These studies showed a dose- and time-dependent increase in ICAM-1 expression in the pulmonary microvascular endothelium. Peak expression occurred at 24 h, while threshold dose was as low as 2 Gy. To determine whether ICAM-1 is required for inflammatory cell infiltration into the irradiated lung, the anti-ICAM-1 blocking antibody was administered by tail vein injection to mice following thoracic irradiation. Inflammatory cells were quantified by immunofluorescence for leukocyte common antigen (CD45). Mice treated with the anti-ICAM-1 blocking antibody showed attenuation of inflammatory cell infiltration into the lung in response to ionizing radiation exposure. To verify the requirement of ICAM-1 in the inflammation-like radiation response, we utilized the ICAM-1 knockout mouse. ICAM-1 was not expressed in the lungs of ICAM-1-deficient mice following treatment with thoracic irradiation. ICAM-1 knockout mice had no increase in the inflammatory cell infiltration into the lung in response to thoracic irradiation. These studies demonstrate a radiation dose-dependent increase in ICAM-1 expression in the pulmonary microvascular endothelium, and show that ICAM-1 is required for inflammatory cell infiltration

  4. Thermoelectric generator with hinged assembly for fins

    DOEpatents

    Purdy, David L.; Shapiro, Zalman M.; Hursen, Thomas F.; Maurer, Gerould W.

    1976-11-02

    A cylindrical casing has a central shielded capsule of radioisotope fuel. A plurality of thermonuclear modules are axially arranged with their hot junctions resiliently pressed toward the shield and with their cold junctions adjacent a transition member having fins radiating heat to the environment. For each module, the assembly of transition member and fins is hinged to the casing for swinging to permit access to and removal of such module. A ceramic plate having gold layers on opposite faces prevents diffusion bonding of the hot junction to the shield.

  5. Gap junction-mediated electrical transmission: regulatory mechanisms and plasticity

    PubMed Central

    Pereda, Alberto E.; Curti, Sebastian; Hoge, Gregory; Cachope, Roger; Flores, Carmen E.; Rash, John E.

    2012-01-01

    The term synapse applies to cellular specializations that articulate the processing of information within neural circuits by providing a mechanism for the transfer of information between two different neurons. There are two main modalities of synaptic transmission: chemical and electrical. While most efforts have been dedicated to the understanding of the properties and modifiability of chemical transmission, less is still known regarding the plastic properties of electrical synapses, whose structural correlate is the gap junction. A wealth of data indicates that, rather than passive intercellular channels, electrical synapses are more dynamic and modifiable than was generally perceived. This article will discuss the factors determining the strength of electrical transmission and review current evidence demonstrating its dynamic properties. Like their chemical counterparts, electrical synapses can also be plastic and modifiable. PMID:22659675

  6. Trends in drug delivery through tissue barriers containing tight junctions

    PubMed Central

    Tscheik, Christian; Blasig, Ingolf E.; Winkler, Lars

    2013-01-01

    A limitation in the uptake of many drugs is the restricted permeation through tissue barriers. There are two general ways to cross barriers formed by cell layers: by transcytosis or by diffusion through the intercellular space. In the latter, tight junctions (TJs) play the decisive role in the regulation of the barrier permeability. Thus, transient modulation of TJs is a potent strategy to improve drug delivery. There have been extensive studies on surfactant-like absorption enhancers. One of the most effective enhancers found is sodium caprate. However, this modulates TJs in an unspecific fashion. A novel approach would be the specific modulation of TJ-associated marvel proteins and claudins, which are the main structural components of the TJs. Recent studies have identified synthetic peptidomimetics and RNA interference techniques to downregulate the expression of targeted TJ proteins. This review summarizes current progress and discusses the impact on TJs' barrier function. PMID:24665392

  7. Morphology and intercellular communication in glial cells of intramural ganglia from the guinea-pig urinary bladder.

    PubMed

    Hanani, M; Maudlej, N; Härtig, W

    1999-04-16

    Neurons in most peripheral ganglia are surrounded by satellite glial cells (SCs), but these cells have so far received little attention. We used immunohistochemistry and intracellular injections of tracers to characterize SCs in the intramural ganglia of the guinea-pig urinary bladder, which are part of the parasympathetic system. Intracellular injections of horseradish peroxidase (HRP) revealed two morphological types: cells that surrounded neurons and are SCs proper, and bipolar cells with processes that projected into the nerve fiber bundles connecting the ganglia. SCs were immunopositive for glutamine synthetase (GS) and S100beta and immunonegative for glial fibrillary acidic protein (GFAP). Injections of Lucifer yellow (LY) or biocytin (molecules known to cross gap junctions) into single SCs showed that these cells have a very low degree of intercellular coupling. A mean of 0.31 and 0.71 cells were coupled to the injected cells, using LY and biocytin, respectively. It appears that SCs in the bladder ganglia are distinct from central and enteric glial cells in the small degree of their coupling and in the absence of GFAP immunostaining.

  8. Activity-Dependent Neuronal Control of Gap-Junctional Communication in Astrocytes

    PubMed Central

    Rouach, Nathalie; Glowinski, Jacques; Giaume, Christian

    2000-01-01

    A typical feature of astrocytes is their high degree of intercellular communication through gap junction channels. Using different models of astrocyte cultures and astrocyte/neuron cocultures, we have demonstrated that neurons upregulate gap-junctional communication and the expression of connexin 43 (Cx43) in astrocytes. The propagation of intercellular calcium waves triggered in astrocytes by mechanical stimulation was also increased in cocultures. This facilitation depends on the age and number of neurons, indicating that the state of neuronal differentiation and neuron density constitute two crucial factors of this interaction. The effects of neurons on astrocytic communication and Cx43 expression were reversed completely after neurotoxic treatments. Moreover, the neuronal facilitation of glial coupling was suppressed, without change in Cx43 expression, after prolonged pharmacological treatments that prevented spontaneous synaptic activity. Altogether, these results demonstrate that neurons exert multiple and differential controls on astrocytic gap-junctional communication. Since astrocytes have been shown to facilitate synaptic efficacy, our findings suggest that neuronal and astrocytic networks interact actively through mutual setting of their respective modes of communication. PMID:10871289

  9. Diurnal Variation of Tight Junction Integrity Associates Inversely with Matrix Metalloproteinase Expression in Xenopus laevis Corneal Epithelium: Implications for Circadian Regulation of Homeostatic Surface Cell Desquamation

    PubMed Central

    Wiechmann, Allan F.; Ceresa, Brian P.; Howard, Eric W.

    2014-01-01

    Background and Objectives The corneal epithelium provides a protective barrier against pathogen entrance and abrasive forces, largely due to the intercellular junctional complexes between neighboring cells. After a prescribed duration at the corneal surface, tight junctions between squamous surface cells must be disrupted to enable them to desquamate as a component of the tissue homeostatic renewal. We hypothesize that matrix metalloproteinase (MMPs) are secreted by corneal epithelial cells and cleave intercellular junctional proteins extracellularly at the epithelial surface. The purpose of this study was to examine the expression of specific MMPs and tight junction proteins during both the light and dark phases of the circadian cycle, and to assess their temporal and spatial relationships in the Xenopus laevis corneal epithelium. Methodology/Principal Findings Expression of MMP-2, tissue inhibitor of MMP-2 (TIMP-2), membrane type 1-MMP (MT1-MMP) and the tight junction proteins occludin and claudin-4 were examined by confocal double-label immunohistochemistry on corneas obtained from Xenopus frogs at different circadian times. Occludin and claudin-4 expression was generally uniformly intact on the surface corneal epithelial cell lateral membranes during the daytime, but was frequently disrupted in small clusters of cells at night. Concomitantly, MMP-2 expression was often elevated in a mosaic pattern at nighttime and associated with clusters of desquamating surface cells. The MMP-2 binding partners, TIMP-2 and MT1-MMP were also localized to surface corneal epithelial cells during both the light and dark phases, with TIMP-2 tending to be elevated during the daytime. Conclusions/Significance MMP-2 protein expression is elevated in a mosaic pattern in surface corneal epithelial cells during the nighttime in Xenopus laevis, and may play a role in homeostatic surface cell desquamation by disrupting intercellular junctional proteins. The sequence of MMP secretion and

  10. Fractional order junctions

    NASA Astrophysics Data System (ADS)

    Machado, J. Tenreiro

    2015-01-01

    Gottfried Leibniz generalized the derivation and integration, extending the operators from integer up to real, or even complex, orders. It is presently recognized that the resulting models capture long term memory effects difficult to describe by classical tools. Leon Chua generalized the set of lumped electrical elements that provide the building blocks in mathematical models. His proposal of the memristor and of higher order elements broadened the scope of variables and relationships embedded in the development of models. This paper follows the two directions and proposes a new logical step, by generalizing the concept of junction. Classical junctions interconnect system elements using simple algebraic restrictions. Nevertheless, this simplistic approach may be misleading in the presence of unexpected dynamical phenomena and requires including additional "parasitic" elements. The novel γ -junction includes, as special cases, the standard series and parallel connections and allows a new degree of freedom when building models. The proposal motivates the search for experimental and real world manifestations of the abstract conjectures.

  11. Thermoelectricity in molecular junctions.

    PubMed

    Reddy, Pramod; Jang, Sung-Yeon; Segalman, Rachel A; Majumdar, Arun

    2007-03-16

    By trapping molecules between two gold electrodes with a temperature difference across them, the junction Seebeck coefficients of 1,4-benzenedithiol (BDT), 4,4'-dibenzenedithiol, and 4,4''-tribenzenedithiol in contact with gold were measured at room temperature to be +8.7 +/- 2.1 microvolts per kelvin (muV/K), +12.9 +/- 2.2 muV/K, and +14.2 +/- 3.2 muV/K, respectively (where the error is the full width half maximum of the statistical distributions). The positive sign unambiguously indicates p-type (hole) conduction in these heterojunctions, whereas the Au Fermi level position for Au-BDT-Au junctions was identified to be 1.2 eV above the highest occupied molecular orbital level of BDT. The ability to study thermoelectricity in molecular junctions provides the opportunity to address these fundamental unanswered questions about their electronic structure and to begin exploring molecular thermoelectric energy conversion.

  12. Automated array assembly, phase 2

    NASA Technical Reports Server (NTRS)

    Carbajal, B. G.

    1979-01-01

    Tasks of scaling up the tandem junction cell (TJC) from 2 cm x 2 cm to 6.2 cm and the assembly of several modules using these large area TJC's are described. The scale-up of the TJC was based on using the existing process and doing the necessary design activities to increase the cell area to an acceptably large area. The design was carried out using available device models. The design was verified and sample large area TJCs were fabricated. Mechanical and process problems occurred causing a schedule slippage that resulted in contract expiration before enough large-area TJCs were fabricated to populate the sample tandem junction modules (TJM). A TJM design was carried out in which the module interconnects served to augment the current collecting buses on the cell. No sample TJMs were assembled due to a shortage of large-area TJCs.

  13. Changes in intestinal tight junction permeability associated with industrial food additives explain the rising incidence of autoimmune disease.

    PubMed

    Lerner, Aaron; Matthias, Torsten

    2015-06-01

    The incidence of autoimmune diseases is increasing along with the expansion of industrial food processing and food additive consumption. The intestinal epithelial barrier, with its intercellular tight junction, controls the equilibrium between tolerance and immunity to non-self-antigens. As a result, particular attention is being placed on the role of tight junction dysfunction in the pathogenesis of AD. Tight junction leakage is enhanced by many luminal components, commonly used industrial food additives being some of them. Glucose, salt, emulsifiers, organic solvents, gluten, microbial transglutaminase, and nanoparticles are extensively and increasingly used by the food industry, claim the manufacturers, to improve the qualities of food. However, all of the aforementioned additives increase intestinal permeability by breaching the integrity of tight junction paracellular transfer. In fact, tight junction dysfunction is common in multiple autoimmune diseases and the central part played by the tight junction in autoimmune diseases pathogenesis is extensively described. It is hypothesized that commonly used industrial food additives abrogate human epithelial barrier function, thus, increasing intestinal permeability through the opened tight junction, resulting in entry of foreign immunogenic antigens and activation of the autoimmune cascade. Future research on food additives exposure-intestinal permeability-autoimmunity interplay will enhance our knowledge of the common mechanisms associated with autoimmune progression.

  14. The Nectin-4/Afadin Protein Complex and Intercellular Membrane Pores Contribute to Rapid Spread of Measles Virus in Primary Human Airway Epithelia

    PubMed Central

    Singh, Brajesh K.; Hornick, Andrew L.; Krishnamurthy, Sateesh; Locke, Anna C.; Mendoza, Crystal A.; Mateo, Mathieu; Miller-Hunt, Catherine L.; Cattaneo, Roberto

    2015-01-01

    ABSTRACT The discovery that measles virus (MV) uses the adherens junction protein nectin-4 as its epithelial receptor provides a new vantage point from which to characterize its rapid spread in the airway epithelium. We show here that in well-differentiated primary cultures of airway epithelial cells from human donors (HAE), MV infectious centers form rapidly and become larger than those of other respiratory pathogens: human respiratory syncytial virus, parainfluenza virus 5, and Sendai virus. While visible syncytia do not form after MV infection of HAE, the cytoplasm of an infected cell suddenly flows into an adjacent cell, as visualized through wild-type MV-expressed cytoplasmic green fluorescent protein (GFP). High-resolution video microscopy documents that GFP flows through openings that form on the lateral surfaces between columnar epithelial cells. To assess the relevance of the protein afadin, which connects nectin-4 to the actin cytoskeleton, we knocked down its mRNA. This resulted in more-limited infectious-center formation. We also generated a nectin-4 mutant without the afadin-binding site in its cytoplasmic tail. This mutant was less effective than wild-type human nectin-4 at promoting MV infection in primary cultures of porcine airway epithelia. Thus, in airway epithelial cells, MV spread requires the nectin-4/afadin complex and is based on cytoplasm transfer between columnar cells. Since the viral membrane fusion apparatus may open the passages that allow cytoplasm transfer, we refer to them as intercellular membrane pores. Virus-induced intercellular pores may contribute to extremely efficient measles contagion by promoting the rapid spread of the virus through the upper respiratory epithelium. IMPORTANCE Measles virus (MV), while targeted for eradication, still causes about 120,000 deaths per year worldwide. The recent reemergence of measles in insufficiently vaccinated populations in Europe and North America reminds us that measles is extremely

  15. Signatures of topological Josephson junctions

    NASA Astrophysics Data System (ADS)

    Peng, Yang; Pientka, Falko; Berg, Erez; Oreg, Yuval; von Oppen, Felix

    2016-08-01

    Quasiparticle poisoning and diabatic transitions may significantly narrow the window for the experimental observation of the 4 π -periodic dc Josephson effect predicted for topological Josephson junctions. Here, we show that switching-current measurements provide accessible and robust signatures for topological superconductivity which persist in the presence of quasiparticle poisoning processes. Such measurements provide access to the phase-dependent subgap spectrum and Josephson currents of the topological junction when incorporating it into an asymmetric SQUID together with a conventional Josephson junction with large critical current. We also argue that pump-probe experiments with multiple current pulses can be used to measure the quasiparticle poisoning rates of the topological junction. The proposed signatures are particularly robust, even in the presence of Zeeman fields and spin-orbit coupling, when focusing on short Josephson junctions. Finally, we also consider microwave excitations of short topological Josephson junctions which may complement switching-current measurements.

  16. MicroRNAs regulate tight junction proteins and modulate epithelial/endothelial barrier functions

    PubMed Central

    Cichon, Christoph; Sabharwal, Harshana; Rüter, Christian; Schmidt, M Alexander

    2014-01-01

    Tightly controlled epithelial and endothelial barriers are a prerequisite for life as these barriers separate multicellular organisms from their environment and serve as first lines of defense. Barriers between neighboring epithelial cells are formed by multiple intercellular junctions including the ‘apical junctional complex—AJC’ with tight junctions (TJ), adherens junctions (AJ), and desmosomes. TJ consist of tetraspan transmembrane proteins like occludin, various claudins that directly control paracellular permeability, and the ‘Junctional Adhesion Molecules’ (JAMs). For establishing tight barriers TJ are essential but at the same time have to allow also selective permeability. For this, TJ need to be tightly regulated and controlled. This is organized by a variety of adaptor molecules, i.e., protein kinases, phosphatases and GTPases, which in turn are regulated and fine-tuned involving microRNAs (miRNAs). In this review we summarize available data on the role and targeting of miRNAs in the maintenance of epithelial and/or endothelial barriers. PMID:25610754

  17. Calcium-dependent dynamics of cadherin interactions at cell–cell junctions

    PubMed Central

    Kim, Sally A.; Tai, Chin-Yin; Mok, Lee-Peng; Mosser, Eric A.; Schuman, Erin M.

    2011-01-01

    Cadherins play a key role in the dynamics of cell–cell contact formation and remodeling of junctions and tissues. Cadherin–cadherin interactions are gated by extracellular Ca2+, which serves to rigidify the cadherin extracellular domains and promote trans junctional interactions. Here we describe the direct visualization and quantification of spatiotemporal dynamics of N-cadherin interactions across intercellular junctions in living cells using a genetically encodable FRET reporter system. Direct measurements of transjunctional cadherin interactions revealed a sudden, but partial, loss of homophilic interactions (τ = 1.17 ± 0.06 s−1) upon chelation of extracellular Ca2+. A cadherin mutant with reduced adhesive activity (W2A) exhibited a faster, more substantial loss of homophilic interactions (τ = 0.86 ± 0.02 s−1), suggesting two types of native cadherin interactions—one that is rapidly modulated by changes in extracellular Ca2+ and another with relatively stable adhesive activity that is Ca2+ independent. The Ca2+-sensitive dynamics of cadherin interactions were transmitted to the cell interior where β-catenin translocated to N-cadherin at the junction in both cells. These data indicate that cadherins can rapidly convey dynamic information about the extracellular environment to both cells that comprise a junction. PMID:21613566

  18. Gap junctions between CA3 pyramidal cells contribute to network synchronization in neonatal hippocampus.

    PubMed

    Molchanova, Svetlana M; Huupponen, Johanna; Lauri, Sari E; Taira, Tomi

    2016-08-01

    Direct electrical coupling between neurons through gap junctions is prominent during development, when synaptic connectivity is scarce, providing the additional intercellular connectivity. However, functional studies of gap junctions are hampered by the unspecificity of pharmacological tools available. Here we have investigated gap-junctional coupling between CA3 pyramidal cells in neonatal hippocampus and its contribution to early network activity. Four different gap junction inhibitors, including the general blocker carbenoxolone, decreased the frequency of network activity bursts in CA3 area of hippocampus of P3-6 rats, suggesting the involvement of electrical connections in the generation of spontaneous network activity. In CA3 pyramidal cells, spikelets evoked by local stimulation of stratum oriens, were inhibited by carbenoxolone, but not by inhibitors of glutamatergic and GABAergic synaptic transmission, signifying the presence of electrical connectivity through axo-axonic gap junctions. Carbenoxolone also decreased the success rate of firing antidromic action potentials in response to stimulation, and changed the pattern of spontaneous action potential firing of CA3 pyramidal cells. Altogether, these data suggest that electrical coupling of CA3 pyramidal cells contribute to the generation of the early network events in neonatal hippocampus by modulating their firing pattern and synchronization.

  19. An induced junction photovoltaic cell

    NASA Technical Reports Server (NTRS)

    Call, R. L.

    1974-01-01

    Silicon solar cells operating with induced junctions rather than diffused junctions have been fabricated and tested. Induced junctions were created by forming an inversion layer near the surface of the silicon by supplying a sheet of positive charge above the surface. Measurements of the response of the inversion layer cell to light of different wavelengths indicated it to be more sensitive to the shorter wavelengths of the sun's spectrum than conventional cells. The greater sensitivity occurs because of the shallow junction and the strong electric field at the surface.

  20. GUARD RING SEMICONDUCTOR JUNCTION

    DOEpatents

    Goulding, F.S.; Hansen, W.L.

    1963-12-01

    A semiconductor diode having a very low noise characteristic when used under reverse bias is described. Surface leakage currents, which in conventional diodes greatly contribute to noise, are prevented from mixing with the desired signal currents. A p-n junction is formed with a thin layer of heavily doped semiconductor material disposed on a lightly doped, physically thick base material. An annular groove cuts through the thin layer and into the base for a short distance, dividing the thin layer into a peripheral guard ring that encircles the central region. Noise signal currents are shunted through the guard ring, leaving the central region free from such currents. (AEC)

  1. YBCO Josephson Junction Arrays

    DTIC Science & Technology

    1993-07-14

    Also, CaRuO 3 is chemically compatible with YBa2Cu30 7 and its conductivity does not appear to be strongly dependent on doping or oxygen concentration...barrier conductivity is quite high. The first YBa2Cu30 7 layer and the SrTiO3 layer are deposited first and then patterned with ion milling (to help form...the edge junction will dominate any leakage through the SrTiO3 , thus the integrity of that dielectric will not be a concern here. The integrity of the

  2. Contribution of Host Intracellular Transport Machineries to Intercellular Movement of Turnip Mosaic Virus

    PubMed Central

    Nelson, Richard S.; Zheng, Huanquan; Laliberté, Jean-François

    2013-01-01

    The contribution of different host cell transport systems in the intercellular movement of turnip mosaic virus (TuMV) was investigated. To discriminate between primary infections and secondary infections associated with the virus intercellular movement, a gene cassette expressing GFP-HDEL was inserted adjacent to a TuMV infectious cassette expressing 6K2:mCherry, both within the T-DNA borders of the binary vector pCambia. In this system, both gene cassettes were delivered to the same cell by a single binary vector and primary infection foci emitted green and red fluorescence while secondarily infected cells emitted only red fluorescence. Intercellular movement was measured at 72 hours post infiltration and was estimated to proceed at an average rate of one cell being infected every three hours over an observation period of 17 hours. To determine if the secretory pathway were important for TuMV intercellular movement, chemical and protein inhibitors that blocked both early and late secretory pathways were used. Treatment with Brefeldin A or Concanamycin A or expression of ARF1 or RAB-E1d dominant negative mutants, all of which inhibit pre- or post-Golgi transport, reduced intercellular movement by the virus. These treatments, however, did not inhibit virus replication in primary infected cells. Pharmacological interference assays using Tyrphostin A23 or Wortmannin showed that endocytosis was not important for TuMV intercellular movement. Lack of co-localization by endocytosed FM4-64 and Ara7 (AtRabF2b) with TuMV-induced 6K2-tagged vesicles further supported this conclusion. Microfilament depolymerizing drugs and silencing expression of myosin XI-2 gene, but not myosin VIII genes, also inhibited TuMV intercellular movement. Expression of dominant negative myosin mutants confirmed the role played by myosin XI-2 as well as by myosin XI-K in TuMV intercellular movement. Using this dual gene cassette expression system and transport inhibitors, components of the secretory

  3. Rescue of Notch signaling in cells incapable of GDP-L-fucose synthesis by gap junction transfer of GDP-L-fucose in Drosophila.

    PubMed

    Ayukawa, Tomonori; Matsumoto, Kenjiroo; Ishikawa, Hiroyuki O; Ishio, Akira; Yamakawa, Tomoko; Aoyama, Naoki; Suzuki, Takuya; Matsuno, Kenji

    2012-09-18

    Notch (N) is a transmembrane receptor that mediates cell-cell interactions to determine many cell-fate decisions. N contains EGF-like repeats, many of which have an O-fucose glycan modification that regulates N-ligand binding. This modification requires GDP-L-fucose as a donor of fucose. The GDP-L-fucose biosynthetic pathways are well understood, including the de novo pathway, which depends on GDP-mannose 4,6 dehydratase (Gmd) and GDP-4-keto-6-deoxy-D-mannose 3,5-epimerase/4-reductase (Gmer). However, the potential for intercellularly supplied GDP-L-fucose and the molecular basis of such transportation have not been explored in depth. To address these points, we studied the genetic effects of mutating Gmd and Gmer on fucose modifications in Drosophila. We found that these mutants functioned cell-nonautonomously, and that GDP-L-fucose was supplied intercellularly through gap junctions composed of Innexin-2. GDP-L-fucose was not supplied through body fluids from different isolated organs, indicating that the intercellular distribution of GDP-L-fucose is restricted within a given organ. Moreover, the gap junction-mediated supply of GDP-L-fucose was sufficient to support the fucosylation of N-glycans and the O-fucosylation of the N EGF-like repeats. Our results indicate that intercellular delivery is a metabolic pathway for nucleotide sugars in live animals under certain circumstances.

  4. Tight Junction Defects in Atopic Dermatitis

    PubMed Central

    De Benedetto, Anna; Rafaels, Nicholas M.; McGirt, Laura Y.; Ivanov, Andrei I.; Georas, Steve N.; Cheadle, Chris; Berger, Alan E.; Zhang, Kunzhong; Vidyasagar, Sadasivan; Yoshida, Takeshi; Boguniewicz, Mark; Hata, Tissa; Schneider, Lynda C.; Hanifin, Jon M.; Gallo, Richard L.; Novak, Natalija; Weidinger, Stephan; Beaty, Terri H.; Leung, Donald Y.; Barnes, Kathleen C.; Beck, Lisa A.

    2010-01-01

    Background Atopic dermatitis (AD) is characterized by dry skin and a hyperreactive immune response to allergens, two cardinal features that are caused in part by epidermal barrier defects. Tight junctions (TJ) reside immediately below the stratum corneum and regulate the selective permeability of the paracellular pathway. Objective We evaluated the expression/function of the TJ protein, claudin-1 in epithelium from AD and nonatopic (NA) subjects and screened two American populations for SNPs in CLDN1. Methods Expression profiles of nonlesional epithelium from extrinsic AD, NA and psoriasis subjects were generated using Illumina’s BeadChips. Dysregulated intercellular proteins were validated by tissue staining and qPCR. Bioelectric properties of epithelium were measured in Ussing chambers. Functional relevance of claudin-1 was assessed using a knockdown approach in primary human keratinocytes (PHK). Twenty seven haplotype-tagging SNPs in CLDN1 were screened in two independent AD populations. Results We observed strikingly reduced expression of the TJ proteins claudin-1 and -23 only in AD, which were validated at the mRNA and protein levels. Claudin-1 expression inversely correlated with Th2 biomarkers. We observed a remarkable impairment of the bioelectric barrier function in AD epidermis. In vitro, we confirmed that silencing claudin-1 expression in human keratinocytes diminishes TJ function while enhancing keratinocyte proliferation. Finally, CLDN1 haplotype-tagging single nucleotide polymorphisms revealed associations with AD in two North American populations. Conclusion Taken together, these data suggest that an impaired epidermal TJ is a novel feature of skin barrier dysfunction and immune dysregulation observed in AD, and that CLDN1 may be a new susceptibility gene in this disease. PMID:21163515

  5. Intercellular ultrafast Ca2+ wave in vascular smooth muscle cells: numerical and experimental study

    NASA Astrophysics Data System (ADS)

    Quijano, J. C.; Raynaud, F.; Nguyen, D.; Piacentini, N.; Meister, J. J.

    2016-08-01

    Vascular smooth muscle cells exhibit intercellular Ca2+ waves in response to local mechanical or KCl stimulation. Recently, a new type of intercellular Ca2+ wave was observed in vitro in a linear arrangement of smooth muscle cells. The intercellular wave was denominated ultrafast Ca2+ wave and it was suggested to be the result of the interplay between membrane potential and Ca2+ dynamics which depended on influx of extracellular Ca2+, cell membrane depolarization and its intercel- lular propagation. In the present study we measured experimentally the conduction velocity of the membrane depolarization and performed simulations of the ultrafast Ca2+ wave along coupled smooth muscle cells. Numerical results reproduced a wide spectrum of experimental observations, including Ca2+ wave velocity, electrotonic membrane depolarization along the network, effects of inhibitors and independence of the Ca2+ wave speed on the intracellular stores. The numerical data also provided new physiological insights suggesting ranges of crucial model parameters that may be altered experimentally and that could significantly affect wave kinetics allowing the modulation of the wave characteristics experimentally. Numerical and experimental results supported the hypothesis that the propagation of membrane depolarization acts as an intercellular messenger mediating intercellular ultrafast Ca2+ waves in smooth muscle cells.

  6. Low dose gamma irradiation enhances defined signaling components of intercellular reactive oxygen-mediated apoptosis induction

    NASA Astrophysics Data System (ADS)

    Bauer, G.

    2011-01-01

    Transformed cells are selectively removed by intercellular ROS-mediated induction of apoptosis. Signaling is based on the HOCl and the NO/peroxynitrite pathway (major pathways) and the nitryl chloride and the metal-catalyzed Haber-Weiss pathway (minor pathways). During tumor progression, resistance against intercellular induction of apoptosis is acquired through expression of membrane-associated catalase. Low dose radiation of nontransformed cells has been shown to enhance intercellular induction of apoptosis. The present study was performed to define the signaling components which are modulated by low dose gamma irradiation. Low dose radiation induced the release of peroxidase from nontransformed, transformed and tumor cells. Extracellular superoxide anion generation was strongly enhanced in the case of transformed cells and tumor cells, but not in nontransformed cells. Enhancement of peroxidase release and superoxide anion generation either increased intercellular induction of apoptosis of transformed cells, or caused a partial protection under specific signaling conditions. In tumor cells, low dose radiation enhanced the production of major signaling components, but this had no effect on apoptosis induction, due to the strong resistance mechanism of tumor cells. Our data specify the nature of low dose radiation-induced effects on specific signaling components of intercellular induction of apoptosis at defined stages of multistep carcinogenesis.

  7. Intercellular ultrafast Ca2+ wave in vascular smooth muscle cells: numerical and experimental study

    PubMed Central

    Quijano, J. C.; Raynaud, F.; Nguyen, D.; Piacentini, N.; Meister, J. J.

    2016-01-01

    Vascular smooth muscle cells exhibit intercellular Ca2+ waves in response to local mechanical or KCl stimulation. Recently, a new type of intercellular Ca2+ wave was observed in vitro in a linear arrangement of smooth muscle cells. The intercellular wave was denominated ultrafast Ca2+ wave and it was suggested to be the result of the interplay between membrane potential and Ca2+ dynamics which depended on influx of extracellular Ca2+, cell membrane depolarization and its intercel- lular propagation. In the present study we measured experimentally the conduction velocity of the membrane depolarization and performed simulations of the ultrafast Ca2+ wave along coupled smooth muscle cells. Numerical results reproduced a wide spectrum of experimental observations, including Ca2+ wave velocity, electrotonic membrane depolarization along the network, effects of inhibitors and independence of the Ca2+ wave speed on the intracellular stores. The numerical data also provided new physiological insights suggesting ranges of crucial model parameters that may be altered experimentally and that could significantly affect wave kinetics allowing the modulation of the wave characteristics experimentally. Numerical and experimental results supported the hypothesis that the propagation of membrane depolarization acts as an intercellular messenger mediating intercellular ultrafast Ca2+ waves in smooth muscle cells. PMID:27507785

  8. A new strategy to measure intercellular adhesion forces in mature cell-cell contacts

    PubMed Central

    Sancho, Ana; Vandersmissen, Ine; Craps, Sander; Luttun, Aernout; Groll, Jürgen

    2017-01-01

    Intercellular adhesion plays a major role in tissue development and homeostasis. Yet, technologies to measure mature cell-cell contacts are not available. We introduce a methodology based on fluidic probe force microscopy to assess cell-cell adhesion forces after formation of mature intercellular contacts in cell monolayers. With this method we quantify that L929 fibroblasts exhibit negligible cell-cell adhesion in monolayers whereas human endothelial cells from the umbilical artery (HUAECs) exert strong intercellular adhesion forces per cell. We use a new in vitro model based on the overexpression of Muscle Segment Homeobox 1 (MSX1) to induce Endothelial-to-Mesenchymal Transition (EndMT), a process involved in cardiovascular development and disease. We reveal how intercellular adhesion forces in monolayer decrease significantly at an early stage of EndMT and we show that cells undergo stiffening and flattening at this stage. This new biomechanical insight complements and expands the established standard biomolecular analyses. Our study thus introduces a novel tool for the assessment of mature intercellular adhesion forces in a physiological setting that will be of relevance to biological processes in developmental biology, tissue regeneration and diseases like cancer and fibrosis. PMID:28393890

  9. Novel preparation of intercellular lipid models of the stratum corneum containing stereoactive ceramide.

    PubMed

    Watanabe, Hiroshi; Obata, Yasuko; Onuki, Yoshinori; Ishida, Kenya; Takayama, Kozo

    2010-03-01

    The microstructure formed by intercellular lipids in the stratum corneum is important for the barrier function of the skin. However, the correlation between lipid composition and microstructure has not yet been clarified. To elucidate the microstructure of intercellular lipids in the stratum corneum, an intercellular lipid model was prepared from ceramide 5 (CER5), cholesterol (CHOL), and palmitic acid (PA), considering the nonuniformity of the lipid components of the stratum corneum. A response surface method incorporating thin-plate spline interpolation (RSM-S) was employed to prepare the CER5/CHOL/PA lipid bilayers. Fluorescence anisotropy of the CER5/CHOL/PA bilayers showed four distinct clusters based on Kohonen's self-organizing maps (SOM). At the centroid formulation of those clusters, the microstructures of CER5/CHOL/PA bilayers were determined using synchrotron X-ray scattering. Three kinds of lamellar structures and two kinds of lateral packing-namely, hexagonal and orthorhombic-were formed. The microstructure of the CER5/CHOL/PA bilayers was likely to be intrinsic to the intercellular lipids in the stratum corneum. In conclusion, the CER5/CHOL/PA bilayers prepared based on RSM-S and SOM were useful as models of the intercellular lipids in the stratum corneum.

  10. A Gap Junction Protein, Inx2, Modulates Calcium Flux to Specify Border Cell Fate during Drosophila oogenesis

    PubMed Central

    Ghosh, Ritabrata; Deshpande, Girish

    2017-01-01

    Intercellular communication mediated by gap junction (GJ) proteins is indispensable during embryogenesis, tissue regeneration and wound healing. Here we report functional analysis of a gap junction protein, Innexin 2 (Inx2), in cell type specification during Drosophila oogenesis. Our data reveal a novel involvement of Inx2 in the specification of Border Cells (BCs), a migratory cell type, whose identity is determined by the cell autonomous STAT activity. We show that Inx2 influences BC fate specification by modulating STAT activity via Domeless receptor endocytosis. Furthermore, detailed experimental analysis has uncovered that Inx2 also regulates a calcium flux that transmits across the follicle cells. We propose that Inx2 mediated calcium flux in the follicle cells stimulates endocytosis by altering Dynamin (Shibire) distribution which is in turn critical for careful calibration of STAT activation and, thus for BC specification. Together our data provide unprecedented molecular insights into how gap junction proteins can regulate cell-type specification. PMID:28114410

  11. Functional heterologous gap junctions in Fundulus ovarian follicles maintain meiotic arrest and permit hydration during oocyte maturation.

    PubMed

    Cerdá, J L; Petrino, T R; Wallace, R A

    1993-11-01

    The physiological significance of heterologous gap junctions between granulosa cells and the oocyte was investigated in late vitellogenic ovarian follicles of the teleost Fundulus heteroclitus. Lucifer Yellow injected into the oocyte readily passed to the overlying granulosa cells, demonstrating effective dye-coupling. Passage of the fluorescent dye, and hence intercellular communication, was inhibited both by the tumor-promoting phorbol ester phorbol 12-myristate 13-acetate (PMA) and by 1-octanol, known uncouplers of gap junctions in a variety of invertebrate and vertebrate cell types. Octanol alone also initiated resumption of meiosis in follicle-enclosed oocytes, indicating that granulosa cells normally maintain meiotic arrest, as apparently occurs in mammalian and amphibian follicles. Both PMA and octanol also consistently inhibited the hydration process that normally accompanies meiotic maturation. These results support a previously suggested hypothesis that K+, which is the primary osmotic effector for oocyte hydration, is translocated via gap junction from granulosa cells to the maturing oocyte.

  12. Mefloquine gap junction blockade and risk of pregnancy loss.

    PubMed

    Nevin, Remington Lee

    2012-09-01

    Obstetric use of the antimalarial drug mefloquine has historically been discouraged during the first trimester and immediately before conception owing to concerns of potential fetal harm. With the rise of resistance to the antimalarial drug sulfadoxine-pyrimethamine (SP), mefloquine is now being considered as a replacement for SP for universal antenatal administration to women from malaria-endemic regions. Recent recommendations have also suggested that mefloquine may be used cautiously among pregnant travelers who cannot otherwise avoid visiting these areas. Mefloquine has been demonstrated to cause blockade of gap junction protein alpha 1 (GJA1) gap junction intercellular communication (GJIC), and recent evidence suggests that GJA1 GJIC is critical to successful embryonic implantation and early placental development. During routine use, mefloquine accumulates in organ and peripheral tissue, crosses the blood-placental barrier, and may plausibly accumulate in developing decidua and trophoblast at concentrations sufficient to interfere with GJA1 GJIC and, thus, cause deleterious effects on fetal outcomes. This conclusion is supported by epidemiological evidence that demonstrates use of the drug during early development is associated with an increased risk of miscarriage and stillbirth. Confirmatory studies are pending, but the available experimental and epidemiological evidence support renewed adherence, where feasible, to existing mefloquine package insert guidance that women avoid the drug during the periconceptional period.

  13. Cell-to-cell coupling in engineered pairs of rat ventricular cardiomyocytes: relation between Cx43 immunofluorescence and intercellular electrical conductance

    PubMed Central

    McCain, Megan L.; Desplantez, Thomas; Geisse, Nicholas A.; Rothen-Rutishauser, Barbara; Oberer, Helene; Parker, Kevin Kit

    2012-01-01

    Gap junctions are composed of connexin (Cx) proteins, which mediate intercellular communication. Cx43 is the dominant Cx in ventricular myocardium, and Cx45 is present in trace amounts. Cx43 immunosignal has been associated with cell-to-cell coupling and electrical propagation, but no studies have directly correlated Cx43 immunosignal to electrical cell-to-cell conductance, gj, in ventricular cardiomyocyte pairs. To assess the correlation between Cx43 immunosignal and gj, we developed a method to determine both parameters from the same cell pair. Neonatal rat ventricular cardiomyocytes were seeded on micropatterned islands of fibronectin. This allowed formation of cell pairs with reproducible shapes and facilitated tracking of cell pair locations. Moreover, cell spreading was limited by the fibronectin pattern, which allowed us to increase cell height by reducing the surface area of the pattern. Whole cell dual voltage clamp was used to record gj of cell pairs after 3–5 days in culture. Fixation of cell pairs before removal of patch electrodes enabled preservation of cell morphology and offline identification of patched pairs. Subsequently, pairs were immunostained, and the volume of junctional Cx43 was quantified using confocal microscopy, image deconvolution, and three-dimensional reconstruction. Our results show a linear correlation between gj and Cx43 immunosignal within a range of 8–50 nS. PMID:22081700

  14. Spontaneous calcium signals induced by gap junctions in a network model of astrocytes

    NASA Astrophysics Data System (ADS)

    Kazantsev, V. B.

    2009-01-01

    The dynamics of a network model of astrocytes coupled by gap junctions is investigated. Calcium dynamics of the single cell is described by the biophysical model comprising the set of three nonlinear differential equations. Intercellular dynamics is provided by the diffusion of inositol 1,4,5-trisphosphate (IP3) through gap junctions between neighboring astrocytes. It is found that the diffusion induces the appearance of spontaneous activity patterns in the network. Stability of the network steady state is analyzed. It is proved that the increase of the diffusion coefficient above a certain critical value yields the generation of low-amplitude subthreshold oscillatory signals in a certain frequency range. It is shown that such spontaneous oscillations can facilitate calcium pulse generation and provide a certain time scale in astrocyte signaling.

  15. Regulation of gap junction channels by infectious agents and inflammation in the CNS

    PubMed Central

    Castellano, Paul; Eugenin, Eliseo A.

    2014-01-01

    Gap junctions (GJs) are conglomerates of intercellular channels that connect the cytoplasm of two or more cells, and facilitate the transfer of ions and small molecules, including second messengers, resulting in metabolic and electrical coordination. In general, loss of gap junctional communication (GJC) has been associated with cellular damage and inflammation resulting in compromise of physiological functions. Recently, it has become evident that GJ channels also play a critical role in the pathogenesis of infectious diseases and associated inflammation. Several pathogens use the transfer of intracellular signals through GJ channels to spread infection and toxic signals that amplify inflammation to neighboring cells. Thus, identification of the mechanisms by which several infectious agents alter GJC could result in new potential therapeutic approaches to reduce inflammation and their pathogenesis. PMID:24847208

  16. CD24 regulated gene expression and distribution of tight junction proteins is associated with altered barrier function in oral epithelial monolayers

    PubMed Central

    Ye, Ping; Nadkarni, Mangala A; Simonian, Mary; Hunter, Neil

    2009-01-01

    Background Control of intercellular penetration of microbial products is critical for the barrier function of oral epithelia. We demonstrated that CD24 is selectively and strongly expressed in the cells of the epithelial attachment to the tooth and the epithelial lining of the diseased periodontal pocket and studies in vitro showed that CD24 regulated expression of the epithelial intercellular adhesion protein E-cadherin. Results In the present study, the barrier function of oral epithelial cell monolayers to low molecular weight dextran was assayed as a model for the normal physiological function of the epithelial attachment to limit ingress of microbial products from oral microbial biofilms. Paracellular transfer of low molecular weight dextran across monolayers of oral epithelial cells was specifically decreased following incubation with anti-CD24 peptide antibody whereas passage of dextran across the monolayer was increased following silencing of mRNA for CD24. Changes in barrier function were related to the selective regulation of the genes encoding zonula occludens-1, zonula occludens-2 and occludin, proteins implicated in tight junctions. More particularly, enhanced barrier function was related to relocation of these proteins to the cell periphery, compatible with tight junctions. Conclusion CD24 has the constitutive function of maintaining expression of selected genes encoding tight junction components associated with a marginal barrier function of epithelial monolayers. Activation by binding of an external ligand to CD24 enhances this expression but is also effective in re-deployment of tight junction proteins that is aligned with enhanced intercellular barrier function. These results establish the potential of CD24 to act as a potent regulator of the intercellular barrier function of epithelia in response to local microbial ecology. PMID:19138432

  17. Electron Transport through Porphyrin Molecular Junctions

    NASA Astrophysics Data System (ADS)

    Zhou, Qi

    The goal of this work is to study the properties that would affect the electron transport through a porphyrin molecular junction. This work contributes to the field of electron transport in molecular junctions in the following 3 aspects. First of all, by carrying out experiments comparing the conductance of the iron (III) porphyrin (protected) and the free base porphyrin (protected), it is confirmed that the molecular energy level broadening and shifting occurs for porphyrin molecules when coupled with the metal electrodes, and this level broadening and shifting plays an important role in the electron transport through molecular junctions. Secondly, by carrying out an in-situ deprotection of the acetyl-protected free base porphyrin molecules, it is found out that the presence of acetyl groups reduces the conductance. Thirdly, by incorporating the Matrix-assisted laser desorption/ionization (MALDI) spectrum and the in-situ deprotection prior to formation of molecular junctions, it allows a more precise understanding of the molecules involved in the formation of molecular junctions, and therefore allows an accurate analysis of the conductance histogram. The molecules are prepared by self-assembly and the junctions are formed using a Scanning Tunneling Microscopy (STM) molecular break junction technique. The porphyrin molecules are characterized by MALDI in solution before self-assembly to a gold/mica substrate. The self-assembled monolayers (SAMs) of porphyrins on gold are characterized by Ultraviolet-visible (UV-Vis) reflection spectroscopy to confirm that the molecules are attached to the substrate. The SAMs are then characterized by Angle-Resolved X-ray photoelectron spectroscopy (ARXPS) to determine the thickness and the average molecular orientation of the molecular layer. The electron transport is measured by conductance-displacement (G-S) experiments under a given bias (-0.4V). The conductance value of a single molecule is identified by a statistical analysis

  18. Development and fabrication of a solar cell junction processing system

    NASA Technical Reports Server (NTRS)

    Bunker, S.

    1981-01-01

    A solar cell junction processing system was developed and fabricated. A pulsed electron beam for the four inch wafers is being assembled and tested, wafers were successfully pulsed, and solar cells fabricated. Assembly of the transport locks is completed. The transport was operated successfully but not with sufficient reproducibility. An experiment test facility to examine potential scaleup problems associated with the proposed ion implanter design was constructed and operated. Cells were implanted and found to have efficiency identical to the normal Spire implant process.

  19. Developmental gene regulation by an ancient intercellular communication system in social amoebae.

    PubMed

    Asghar, Asma; Groth, Marco; Siol, Oliver; Gaube, Friedemann; Enzensperger, Christoph; Glöckner, Gernot; Winckler, Thomas

    2012-01-01

    The social amoebae (Dictyostelia) use quorum sensing-like communication systems to coordinate the periodic transition from uni- to multicellularity. The monophyletic descent of the Dictyostelia provides a unique opportunity to study the origin and adaptive evolution of such intercellular communication systems. We determined that the ability of aggregation-competent cells to respond to the intercellular messenger glorin occurred in the most ancient taxa of the Dictyostelia. We show using Illumina sequencing technology that glorin mediates rapid changes in gene expression at the transition from vegetative growth to aggregation. We conclude that peptide-based communication is the most ancient form of intercellular signaling in the evolution of multicellularity in the social amoebae, but has been repeatedly replaced by other communication systems during the monophyletic evolution of the social amoebae. Glorin communication has parallels with quorum sensing in that the molecule diffuses into the field, stimulates gene expression in receptive cells and coordinates a population-wide response.

  20. Intercellular Coupling of the Cell Cycle and Circadian Clock in Adult Stem Cell Culture.

    PubMed

    Matsu-Ura, Toru; Dovzhenok, Andrey; Aihara, Eitaro; Rood, Jill; Le, Hung; Ren, Yan; Rosselot, Andrew E; Zhang, Tongli; Lee, Choogon; Obrietan, Karl; Montrose, Marshall H; Lim, Sookkyung; Moore, Sean R; Hong, Christian I

    2016-12-01

    Circadian clock-gated cell division cycles are observed from cyanobacteria to mammals via intracellular molecular connections between these two oscillators. Here we demonstrate WNT-mediated intercellular coupling between the cell cycle and circadian clock in 3D murine intestinal organoids (enteroids). The circadian clock gates a population of cells with heterogeneous cell-cycle times that emerge as 12-hr synchronized cell division cycles. Remarkably, we observe reduced-amplitude oscillations of circadian rhythms in intestinal stem cells and progenitor cells, indicating an intercellular signal arising from differentiated cells governing circadian clock-dependent synchronized cell division cycles. Stochastic simulations and experimental validations reveal Paneth cell-secreted WNT as the key intercellular coupling component linking the circadian clock and cell cycle in enteroids.

  1. Asymmetric homotypic interactions of the atypical cadherin flamingo mediate intercellular polarity signaling.

    PubMed

    Chen, Wei-Shen; Antic, Dragana; Matis, Maja; Logan, Catriona Y; Povelones, Michael; Anderson, Graham A; Nusse, Roel; Axelrod, Jeffrey D

    2008-06-13

    Acquisition of planar cell polarity (PCP) in epithelia involves intercellular communication, during which cells align their polarity with that of their neighbors. The transmembrane proteins Frizzled (Fz) and Van Gogh (Vang) are essential components of the intercellular communication mechanism, as loss of either strongly perturbs the polarity of neighboring cells. How Fz and Vang communicate polarity information between neighboring cells is poorly understood. The atypical cadherin, Flamingo (Fmi), is implicated in this process, yet whether Fmi acts permissively as a scaffold or instructively as a signal is unclear. Here, we provide evidence that Fmi functions instructively to mediate Fz-Vang intercellular signal relay, recruiting Fz and Vang to opposite sides of cell boundaries. We propose that two functional forms of Fmi, one of which is induced by and physically interacts with Fz, bind each other to create cadherin homodimers that signal bidirectionally and asymmetrically, instructing unequal responses in adjacent cell membranes to establish molecular asymmetry.

  2. Bacteriophage assembly.

    PubMed

    Aksyuk, Anastasia A; Rossmann, Michael G

    2011-03-01

    Bacteriophages have been a model system to study assembly processes for over half a century. Formation of infectious phage particles involves specific protein-protein and protein-nucleic acid interactions, as well as large conformational changes of assembly precursors. The sequence and molecular mechanisms of phage assembly have been elucidated by a variety of methods. Differences and similarities of assembly processes in several different groups of bacteriophages are discussed in this review. The general principles of phage assembly are applicable to many macromolecular complexes.

  3. Azobenzenes as light-controlled molecular electronic switches in nanoscale metal-molecule-metal junctions.

    PubMed

    Mativetsky, Jeffrey M; Pace, Giuseppina; Elbing, Mark; Rampi, Maria A; Mayor, Marcel; Samorì, Paolo

    2008-07-23

    Conductance switching associated with the photoisomerization of azobenzene-based (Azo) molecules was observed in nanoscopic metal-molecule-metal junctions. The junctions were formed by using a conducting atomic force microscope (C-AFM) approach, where a metallic AFM tip was used to electrically contact a gold-supported Azo self-assembled monolayer. The measured 30-fold increase in conductance is consistent with the expected decrease in tunneling barrier length resulting from the conformational change of the Azo molecule.

  4. Ion bipolar junction transistors

    PubMed Central

    Tybrandt, Klas; Larsson, Karin C.; Richter-Dahlfors, Agneta; Berggren, Magnus

    2010-01-01

    Dynamic control of chemical microenvironments is essential for continued development in numerous fields of life sciences. Such control could be achieved with active chemical circuits for delivery of ions and biomolecules. As the basis for such circuitry, we report a solid-state ion bipolar junction transistor (IBJT) based on conducting polymers and thin films of anion- and cation-selective membranes. The IBJT is the ionic analogue to the conventional semiconductor BJT and is manufactured using standard microfabrication techniques. Transistor characteristics along with a model describing the principle of operation, in which an anionic base current amplifies a cationic collector current, are presented. By employing the IBJT as a bioelectronic circuit element for delivery of the neurotransmitter acetylcholine, its efficacy in modulating neuronal cell signaling is demonstrated. PMID:20479274

  5. Ion bipolar junction transistors.

    PubMed

    Tybrandt, Klas; Larsson, Karin C; Richter-Dahlfors, Agneta; Berggren, Magnus

    2010-06-01

    Dynamic control of chemical microenvironments is essential for continued development in numerous fields of life sciences. Such control could be achieved with active chemical circuits for delivery of ions and biomolecules. As the basis for such circuitry, we report a solid-state ion bipolar junction transistor (IBJT) based on conducting polymers and thin films of anion- and cation-selective membranes. The IBJT is the ionic analogue to the conventional semiconductor BJT and is manufactured using standard microfabrication techniques. Transistor characteristics along with a model describing the principle of operation, in which an anionic base current amplifies a cationic collector current, are presented. By employing the IBJT as a bioelectronic circuit element for delivery of the neurotransmitter acetylcholine, its efficacy in modulating neuronal cell signaling is demonstrated.

  6. Chemical control over the energy-level alignment in a two-terminal junction.

    PubMed

    Yuan, Li; Franco, Carlos; Crivillers, Núria; Mas-Torrent, Marta; Cao, Liang; Sangeeth, C S Suchand; Rovira, Concepció; Veciana, Jaume; Nijhuis, Christian A

    2016-07-26

    The energy-level alignment of molecular transistors can be controlled by external gating to move molecular orbitals with respect to the Fermi levels of the source and drain electrodes. Two-terminal molecular tunnelling junctions, however, lack a gate electrode and suffer from Fermi-level pinning, making it difficult to control the energy-level alignment of the system. Here we report an enhancement of 2 orders of magnitude of the tunnelling current in a two-terminal junction via chemical molecular orbital control, changing chemically the molecular component between a stable radical and its non-radical form without altering the supramolecular structure of the junction. Our findings demonstrate that the energy-level alignment in self-assembled monolayer-based junctions can be regulated by purely chemical modifications, which seems an attractive alternative to control the electrical properties of two-terminal junctions.

  7. Chemical control over the energy-level alignment in a two-terminal junction

    PubMed Central

    Yuan, Li; Franco, Carlos; Crivillers, Núria; Mas-Torrent, Marta; Cao, Liang; Sangeeth, C. S. Suchand; Rovira, Concepció; Veciana, Jaume; Nijhuis, Christian A.

    2016-01-01

    The energy-level alignment of molecular transistors can be controlled by external gating to move molecular orbitals with respect to the Fermi levels of the source and drain electrodes. Two-terminal molecular tunnelling junctions, however, lack a gate electrode and suffer from Fermi-level pinning, making it difficult to control the energy-level alignment of the system. Here we report an enhancement of 2 orders of magnitude of the tunnelling current in a two-terminal junction via chemical molecular orbital control, changing chemically the molecular component between a stable radical and its non-radical form without altering the supramolecular structure of the junction. Our findings demonstrate that the energy-level alignment in self-assembled monolayer-based junctions can be regulated by purely chemical modifications, which seems an attractive alternative to control the electrical properties of two-terminal junctions. PMID:27456200

  8. Regulation of neuronal axon specification by glia-neuron gap junctions in C. elegans

    PubMed Central

    Meng, Lingfeng; Zhang, Albert; Jin, Yishi; Yan, Dong

    2016-01-01

    Axon specification is a critical step in neuronal development, and the function of glial cells in this process is not fully understood. Here, we show that C. elegans GLR glial cells regulate axon specification of their nearby GABAergic RME neurons through GLR-RME gap junctions. Disruption of GLR-RME gap junctions causes misaccumulation of axonal markers in non-axonal neurites of RME neurons and converts microtubules in those neurites to form an axon-like assembly. We further uncover that GLR-RME gap junctions regulate RME axon specification through activation of the CDK-5 pathway in a calcium-dependent manner, involving a calpain clp-4. Therefore, our study reveals the function of glia-neuron gap junctions in neuronal axon specification and shows that calcium originated from glial cells can regulate neuronal intracellular pathways through gap junctions. DOI: http://dx.doi.org/10.7554/eLife.19510.001 PMID:27767956

  9. Junctional proteins of the blood-brain barrier: New insights into function and dysfunction

    PubMed Central

    Stamatovic, Svetlana M; Johnson, Allison M; Keep, Richard F; Andjelkovic, Anuska V

    2016-01-01

    abstract The blood-brain barrier (BBB) is a highly complex and dynamic barrier. It is formed by an interdependent network of brain capillary endothelial cells, endowed with barrier properties, and perivascular cells (astrocytes and pericytes) responsible for inducing and maintaining those properties. One of the primary properties of the BBB is a strict regulation of paracellular permeability due to the presence of junctional complexes (tight, adherens and gap junctions) between the endothelial cells. Alterations in junction assembly and function significantly affect BBB properties, particularly barrier permeability. However, such alterations are also involved in remodeling the brain endothelial cell surface and regulating brain endothelial cell phenotype. This review summarizes the characteristics of brain endothelial tight, adherens and gap junctions and highlights structural and functional alterations in junctional proteins that may contribute to BBB dysfunction. PMID:27141427

  10. Roles of gap junctions and hemichannels in bone cell functions and in signal transmission of mechanical stress

    PubMed Central

    Jiang, Jean Xin; Siller-Jackson, Arlene Janel; Burra, Sirisha

    2007-01-01

    Gap junctions formed by connexins (Cx) play an important role in transmitting signals between bone cells such as osteoblasts and osteoclasts, cells responsible for bone formation and bone remodeling, respectively. Gap junction intercellular communication (GJIC) has been demonstrated to mediate the process of osteoblast differentiation and bone formation. Furthermore, GJIC propagates Ca2+ signaling, conveys anabolic effects of hormones and growth factors, and regulates gene transcription of osteoblast differentiation markers. GJIC is also implicated to regulate osteoclast formation, survival and apoptosis. Compared with other bone cells, the most abundant type are osteocytes, which express large amounts of connexins. Mechanosensing osteocytes connect and form gap junctions with themselves and other cells only through the tips of their dendritic processes, a relatively small percent of the total cell surface area compared to other cells. Recent studies show that in addition to gap junctions, osteoblasts and osteocytes express functional hemichannels, the un-opposed halves of gap junction channels. Hemichannels are localized at the cell surface and function independently of gap junctions. Hemichannels in osteocytes mediate the immediate release of prostaglandins in response to mechanical stress. The major challenges remaining in the field are how the functions of these two types of channels are coordinated in bone cells and what the asserted, distinct effects of these channels are on bone formation and remodeling processes, and on conveying signals elicited by mechanical loading. PMID:17127393

  11. Roles of gap junctions and hemichannels in bone cell functions and in signal transmission of mechanical stress.

    PubMed

    Jiang, Jean Xin; Siller-Jackson, Arlene Janel; Burra, Sirisha

    2007-01-01

    Gap junctions formed by connexins (Cx) play an important role in transmitting signals between bone cells such as osteoblasts and osteoclasts, cells responsible for bone formation and bone remodeling, respectively. Gap junction intercellular communication (GJIC) has been demonstrated to mediate the process of osteoblast differentiation and bone formation. Furthermore, GJIC propagates Ca2+ signaling, conveys anabolic effects of hormones and growth factors, and regulates gene transcription of osteoblast differentiation markers. GJIC is also implicated to regulate osteoclast formation, survival and apoptosis. Compared with other bone cells, the most abundant type are osteocytes, which express large amounts of connexins. Mechanosensing osteocytes connect and form gap junctions with themselves and other cells only through the tips of their dendritic processes, a relatively small percent of the total cell surface area compared to other cells. Recent studies show that in addition to gap junctions, osteoblasts and osteocytes express functional hemichannels, the un-opposed halves of gap junction channels. Hemichannels are localized at the cell surface and function independently of gap junctions. Hemichannels in osteocytes mediate the immediate release of prostaglandins in response to mechanical stress. The major challenges remaining in the field are how the functions of these two types of channels are coordinated in bone cells and what the asserted, distinct effects of these channels are on bone formation and remodeling processes, and on conveying signals elicited by mechanical loading.

  12. Rapid remodeling of tight junctions during paracellular diapedesis in a human model of the blood-brain barrier.

    PubMed

    Winger, Ryan C; Koblinski, Jennifer E; Kanda, Takashi; Ransohoff, Richard M; Muller, William A

    2014-09-01

    Leukocyte transendothelial migration (TEM; diapedesis) is a critical event in immune surveillance and inflammation. Most TEM occurs at endothelial cell borders (paracellular). However, there is indirect evidence to suggest that, at the tight junctions of the blood-brain barrier (BBB), leukocytes migrate directly through the endothelial cell body (transcellular). Why leukocytes migrate through the endothelial cell body rather than the cell borders is unknown. To test the hypothesis that the tightness of endothelial cell junctions influences the pathway of diapedesis, we developed an in vitro model of the BBB that possessed 10-fold higher electrical resistance than standard culture conditions and strongly expressed the BBB tight junction proteins claudin-5 and claudin-3. We found that paracellular TEM was still the predominant pathway (≥98%) and TEM was dependent on PECAM-1 and CD99. We show that endothelial tight junctions expressing claudin-5 are dynamic and undergo rapid remodeling during TEM. Membrane from the endothelial lateral border recycling compartment is mobilized to the exact site of tight junction remodeling. This preserves the endothelial barrier by sealing the intercellular gaps with membrane and engaging the migrating leukocyte with unligated adhesion molecules (PECAM-1 and CD99) as it crosses the cell border. These findings provide new insights into leukocyte-endothelial interactions at the BBB and suggest that tight junctions are more dynamic than previously appreciated.

  13. The Wnt and BMP Families of Signaling Morphogens at the Vertebrate Neuromuscular Junction

    PubMed Central

    Henríquez, Juan P.; Krull, Catherine E.; Osses, Nelson

    2011-01-01

    The neuromuscular junction has been extensively employed in order to identify crucial determinants of synaptogenesis. At the vertebrate neuromuscular synapse, extracellular matrix and signaling proteins play stimulatory and inhibitory roles on the assembly of functional synapses. Studies in invertebrate species have revealed crucial functions of early morphogens during the assembly and maturation of the neuromuscular junction. Here, we discuss growing evidence addressing the function of Wnt and Bone morphogenetic protein (BMP) signaling pathways at the vertebrate neuromuscular synapse. We focus on the emerging role of Wnt proteins as positive and negative regulators of postsynaptic differentiation. We also address the possible involvement of BMP pathways on motor neuron behavior for the assembly and/or regeneration of the neuromuscular junction. PMID:22272112

  14. Mic13 Is Essential for Formation of Crista Junctions in Mammalian Cells.

    PubMed

    Anand, Ruchika; Strecker, Valentina; Urbach, Jennifer; Wittig, Ilka; Reichert, Andreas S

    2016-01-01

    Mitochondrial cristae are connected to the inner boundary membrane via crista junctions which are implicated in the regulation of oxidative phosphorylation, apoptosis, and import of lipids and proteins. The MICOS complex determines formation of crista junctions. We performed complexome profiling and identified Mic13, also termed Qil1, as a subunit of the MICOS complex. We show that MIC13 is an inner membrane protein physically interacting with MIC60, a central subunit of the MICOS complex. Using the CRISPR/Cas method we generated the first cell line deleted for MIC13. These knockout cells show a complete loss of crista junctions demonstrating that MIC13 is strictly required for the formation of crista junctions. MIC13 is required for the assembly of MIC10, MIC26, and MIC27 into the MICOS complex. However, it is not needed for the formation of the MIC60/MIC19/MIC25 subcomplex suggesting that the latter is not sufficient for crista junction formation. MIC13 is also dispensable for assembly of respiratory chain complexes and for maintaining mitochondrial network morphology. Still, lack of MIC13 resulted in a moderate reduction of mitochondrial respiration. In summary, we show that MIC13 has a fundamental role in crista junction formation and that assembly of respiratory chain supercomplexes is independent of mitochondrial cristae shape.

  15. Mic13 Is Essential for Formation of Crista Junctions in Mammalian Cells

    PubMed Central

    Anand, Ruchika; Strecker, Valentina; Urbach, Jennifer; Wittig, Ilka; Reichert, Andreas S.

    2016-01-01

    Mitochondrial cristae are connected to the inner boundary membrane via crista junctions which are implicated in the regulation of oxidative phosphorylation, apoptosis, and import of lipids and proteins. The MICOS complex determines formation of crista junctions. We performed complexome profiling and identified Mic13, also termed Qil1, as a subunit of the MICOS complex. We show that MIC13 is an inner membrane protein physically interacting with MIC60, a central subunit of the MICOS complex. Using the CRISPR/Cas method we generated the first cell line deleted for MIC13. These knockout cells show a complete loss of crista junctions demonstrating that MIC13 is strictly required for the formation of crista junctions. MIC13 is required for the assembly of MIC10, MIC26, and MIC27 into the MICOS complex. However, it is not needed for the formation of the MIC60/MIC19/MIC25 subcomplex suggesting that the latter is not sufficient for crista junction formation. MIC13 is also dispensable for assembly of respiratory chain complexes and for maintaining mitochondrial network morphology. Still, lack of MIC13 resulted in a moderate reduction of mitochondrial respiration. In summary, we show that MIC13 has a fundamental role in crista junction formation and that assembly of respiratory chain supercomplexes is independent of mitochondrial cristae shape. PMID:27479602

  16. Construction of van der Waals magnetic tunnel junction using ferromagnetic layered dichalcogenide

    NASA Astrophysics Data System (ADS)

    Arai, Miho; Moriya, Rai; Yabuki, Naoto; Masubuchi, Satoru; Ueno, Keiji; Machida, Tomoki

    2015-09-01

    We investigate the micromechanical exfoliation and van der Waals (vdW) assembly of ferromagnetic layered dichalcogenide Fe0.25TaS2. The vdW interlayer coupling at the Fe-intercalated plane of Fe0.25TaS2 allows exfoliation of flakes. A vdW junction between the cleaved crystal surfaces is constructed by dry transfer method. We observe tunnel magnetoresistance in the resulting junction under an external magnetic field applied perpendicular to the plane, demonstrating spin-polarized tunneling between the ferromagnetic layered material and the vdW junction.

  17. Evaluation of the formation of a junctional DNA nanostructure through annealing curve analysis.

    PubMed

    Shin, Seung Won; Park, Kyung Soo; Um, Soong Ho

    2015-02-20

    During the self-assembly of different numbers of oligonucleotides comprising junctional DNA nanostructures, a change in environmental variables (e.g., temperature or salt concentration) has a substantial influence on the final products. Further, distinctive annealing temperatures of oligonucleotides are observed depending on the state of hybridization. Here, we present an evaluation of the annealing characteristics of oligonucleotides for the formation of a simple junctional DNA nanostructure using an annealing curve analysis. This method may be useful for analyzing the formation of complex junctional DNA nanostructures.

  18. Construction of van der Waals magnetic tunnel junction using ferromagnetic layered dichalcogenide

    SciTech Connect

    Arai, Miho; Moriya, Rai Yabuki, Naoto; Masubuchi, Satoru; Ueno, Keiji; Machida, Tomoki

    2015-09-07

    We investigate the micromechanical exfoliation and van der Waals (vdW) assembly of ferromagnetic layered dichalcogenide Fe{sub 0.25}TaS{sub 2}. The vdW interlayer coupling at the Fe-intercalated plane of Fe{sub 0.25}TaS{sub 2} allows exfoliation of flakes. A vdW junction between the cleaved crystal surfaces is constructed by dry transfer method. We observe tunnel magnetoresistance in the resulting junction under an external magnetic field applied perpendicular to the plane, demonstrating spin-polarized tunneling between the ferromagnetic layered material and the vdW junction.

  19. Molecular cloning and functional expression of human connexin37, an endothelial cell gap junction protein.

    PubMed Central

    Reed, K E; Westphale, E M; Larson, D M; Wang, H Z; Veenstra, R D; Beyer, E C

    1993-01-01

    Gap junctions allow direct intercellular coupling between many cells including those in the blood vessel wall. They are formed by a group of related proteins called connexins, containing conserved transmembrane and extracellular domains, but unique cytoplasmic regions that may confer connexin-specific physiological properties. We used polymerase chain reaction amplification and cDNA library screening to clone DNA encoding a human gap junction protein, connexin37 (Cx37). The derived human Cx37 polypeptide contains 333 amino acids, with a predicted molecular mass of 37,238 D. RNA blots demonstrate that Cx37 is expressed in multiple organs and tissues (including heart, uterus, ovary, and blood vessel endothelium) and in primary cultures of vascular endothelial cells. Cx37 mRNA is coexpressed with connexin43 at similar levels in some endothelial cells, but at much lower levels in others. To demonstrate that Cx37 could form functional channels, we stably transfected communication-deficient Neuro2A cells with the Cx37 cDNA. The induced intercellular channels were studied by the double whole cell patch clamp technique. These channels were reversibly inhibited by the uncoupling agent, heptanol (2 mM). The expressed Cx37 channels exhibited multiple conductance levels and showed a pronounced voltage dependence. These electrophysiological characteristics are similar to, but distinct from, those of previously characterized connexins. Images PMID:7680674

  20. A Computational Approach to Detect Gap Junction Plaques and Associate Them with Cells in Fluorescent Images

    PubMed Central

    Goldberg, Joshua S.; Vadakkan, Tegy J.; Hirschi, Karen K.

    2013-01-01

    Intercellular signaling is a fundamental requirement for complex biological system function and survival. Communication between adjoining cells is largely achieved via gap junction channels made up of multiple subunits of connexin proteins, each with unique selectivity and regulatory properties. Intercellular communication via gap junction channels facilitates transmission of an array of cellular signals, including ions, macromolecules, and metabolites that coordinate physiological processes throughout tissues and entire organisms. Although current methods used to quantify connexin expression rely on number or area density measurements in a field of view, they lack cellular assignment, distance measurement capabilities (both within the cell and to extracellular structures), and complete automation. We devised an automated computational approach built on a contour expansion algorithm platform that allows connexin protein detection and assignment to specific cells within complex tissues. In addition, parallel implementation of the contour expansion algorithm allows for high-throughput analysis as the complexity of the biological sample increases. This method does not depend specifically on connexin identification and can be applied more widely to the analysis of numerous immunocytochemical markers as well as to identify particles within tissues such as nanoparticles, gene delivery vehicles, or even cellular fragments such as exosomes or microparticles. PMID:23324867

  1. Thermopower measurements in molecular junctions.

    PubMed

    Rincón-García, Laura; Evangeli, Charalambos; Rubio-Bollinger, Gabino; Agraït, Nicolás

    2016-08-07

    The measurement of thermopower in molecular junctions offers complementary information to conductance measurements and is becoming essential for the understanding of transport processes at the nanoscale. In this review, we discuss the recent advances in the study of the thermoelectric properties of molecular junctions. After presenting the theoretical background for thermoelectricity at the nanoscale, we review the experimental techniques for measuring the thermopower in these systems and discuss the main results. Finally, we consider the challenges in the application of molecular junctions in viable thermoelectric devices.

  2. Joint assembly

    NASA Technical Reports Server (NTRS)

    Wilson, Andrew (Inventor); Punnoose, Andrew (Inventor); Strausser, Katherine (Inventor); Parikh, Neil (Inventor)

    2010-01-01

    A joint assembly is provided which includes a drive assembly and a swivel mechanism. The drive assembly features a motor operatively associated with a plurality of drive shafts for driving auxiliary elements, and a plurality of swivel shafts for pivoting the drive assembly. The swivel mechanism engages the swivel shafts and has a fixable element that may be attached to a foundation. The swivel mechanism is adapted to cooperate with the swivel shafts to pivot the drive assembly with at least two degrees of freedom relative to the foundation. The joint assembly allows for all components to remain encased in a tight, compact, and sealed package, making it ideal for space, exploratory, and commercial applications.

  3. Lightweight IMM PV Flexible Blanket Assembly

    NASA Technical Reports Server (NTRS)

    Spence, Brian

    2015-01-01

    Deployable Space Systems (DSS) has developed an inverted metamorphic multijunction (IMM) photovoltaic (PV) integrated modular blanket assembly (IMBA) that can be rolled or z-folded. This IMM PV IMBA technology enables a revolutionary flexible PV blanket assembly that provides high specific power, exceptional stowed packaging efficiency, and high-voltage operation capability. DSS's technology also accommodates standard third-generation triple junction (ZTJ) PV device technologies to provide significantly improved performance over the current state of the art. This SBIR project demonstrated prototype, flight-like IMM PV IMBA panel assemblies specifically developed, designed, and optimized for NASA's high-voltage solar array missions.

  4. Physiological role of gap-junctional hemichannels. Extracellular calcium-dependent isosmotic volume regulation.

    PubMed

    Quist, A P; Rhee, S K; Lin, H; Lal, R

    2000-03-06

    Hemichannels in the overlapping regions of apposing cells plasma membranes join to form gap junctions and provide an intercellular communication pathway. Hemichannels are also present in the nonjunctional regions of individual cells and their activity is gated by several agents, including calcium. However, their physiological roles are unknown. Using techniques of atomic force microscopy (AFM), fluorescent dye uptake assay, and laser confocal immunofluorescence imaging, we have examined the extracellular calcium-dependent modulation of cell volume. In response to a change in the extracellular physiological calcium concentration (1.8 to junctional proteins (connexins). Volume change did not occur in cells that were not expressing connexins. However, after the transient or stable transfection of connexin43, volume change did occur. The volume increase was accompanied by cytochalasin D-sensitive higher cell stiffness, which helped maintain cell integrity. These cellular physical changes were prevented by gap-junctional blockers, oleamide and beta-glycyrrhetinic acid, or were reversed by returning extracellular calcium to the normal level. We conclude that nongap-junctional hemichannels regulate cell volume in response to the change in extracellular physiological calcium in an otherwise isosmotic situation.

  5. Reversible inhibition of gap junctional communication by tamoxifen in cultured cardiac myocytes.

    PubMed

    Verrecchia, F; Hervé, J

    1997-05-01

    Gap junction channels provide a cell-to-cell conduction pathway for direct exchange of ions and small molecules. The intercellular diffusion of a fluorescent dye, quantified in cardiac myocytes from neonatal rats by monitoring the fluorescence recovery after photobleaching, was found to be interrupted after short-term exposure (15 min) to tamoxifen, an anti-oestrogen drug often used in the treatment of human breast cancer. This diffusional uncoupling was dose dependent, occurred in the concentration range 3-25 microM and reversed after tamoxifen withdrawal. Some possible mechanisms of junctional channel closure have been examined. The cytosolic calcium concentration, examined using the fluorescent indicator Indo-1, did not vary during the short-term action of tamoxifen. A second anti-oestrogen agent (clomiphene) was able to impair gap junctional communication, whereas a third (nafoxidine) had no effect. Protein-kinase-C-inhibitor properties of tamoxifen did not seem to be involved in its uncoupling action. The characteristics of tamoxifen's action (i.e. channel inhibition delay, active concentration range, reversibility, etc.) were very similar to the previously observed effects of several other lipophilic compounds (e. g. 17beta-oestradiol, etc.) on junctional channels, and to recently reported effects of tamoxifen on voltage-gated calcium currents.

  6. Intercellular Extensions Are Induced by the Alphavirus Structural Proteins and Mediate Virus Transmission

    PubMed Central

    Martinez, Maria Guadalupe

    2016-01-01

    Alphaviruses are highly organized enveloped RNA viruses with an internal nucleocapsid surrounded by a membrane containing the E2 and E1 transmembrane proteins. Alphavirus budding takes place at the plasma membrane and requires the interaction of the cytoplasmic domain of E2 with the capsid protein. Here we used WT alphaviruses and Sindbis virus in which E2 was fused to a fluorescent protein to characterize virus exit from host cells. Our results show that alphavirus infection induced striking modifications of the host cell cytoskeleton and resulted in the formation of stable intercellular extensions that emanated exclusively from the infected cell. The intercellular extensions were long (> 10 μM), contained actin and tubulin, and formed flattened contacts with neighboring cells, but did not mediate membrane or cytoplasmic continuity between cells. Receptor down-regulation studies indicated that formation of stable extensions did not require the virus receptor, and that extensions promoted cell-to-cell virus transmission to receptor-depleted cells. Virus mutant experiments demonstrated that formation of extensions required the E2-capsid interaction but not active particle budding, while intercellular transmission of infection required the production of fusion-active virus particles. Protein expression studies showed that even in the absence of virus infection, the viral structural proteins alone induced intercellular extensions, and that these extensions were preferentially targeted to non-expressing cells. Together, our results identify a mechanism for alphavirus cell-to-cell transmission and define the key viral protein interactions that it requires. PMID:27977778

  7. Extracellular ultrathin fibers sensitive to intracellular reactive oxygen species: Formation of intercellular membrane bridges

    SciTech Connect

    Jung, Se-Hui; Park, Jin-Young; Joo, Jung-Hoon; Kim, Young-Myeong; Ha, Kwon-Soo

    2011-07-15

    Membrane bridges are key cellular structures involved in intercellular communication; however, dynamics for their formation are not well understood. We demonstrated the formation and regulation of novel extracellular ultrathin fibers in NIH3T3 cells using confocal and atomic force microscopy. At adjacent regions of neighboring cells, phorbol 12-myristate 13-acetate (PMA) and glucose oxidase induced ultrathin fiber formation, which was prevented by Trolox, a reactive oxygen species (ROS) scavenger. The height of ROS-sensitive ultrathin fibers ranged from 2 to 4 nm. PMA-induced formation of ultrathin fibers was inhibited by cytochalasin D, but not by Taxol or colchicine, indicating that ultrathin fibers mainly comprise microfilaments. PMA-induced ultrathin fibers underwent dynamic structural changes, resulting in formation of intercellular membrane bridges. Thus, these fibers are formed by a mechanism(s) involving ROS and involved in formation of intercellular membrane bridges. Furthermore, ultrastructural imaging of ultrathin fibers may contribute to understanding the diverse mechanisms of cell-to-cell communication and the intercellular transfer of biomolecules, including proteins and cell organelles.

  8. EXPOSURE OF CULTURED MYOCYTES TO ZINC RESULTS IN ALTERED BEAT RATE AND INTERCELLULAR COMMUNICATION.

    EPA Science Inventory

    Exposure of cultured myocytes to zinc results in altered beat rate and intercellular communication

    Graff, Donald W, Devlin, Robert B, Brackhan, Joseph A, Muller-Borer, Barbara J, Bowman, Jill S, Cascio, Wayne E.

    Exposure to ambient air pollution particulate matter (...

  9. Starting points in plant-bacteria nitrogen-fixing symbioses: intercellular invasion of the roots.

    PubMed

    Ibáñez, Fernando; Wall, Luis; Fabra, Adriana

    2016-10-18

    Agricultural practices contribute to climate change by releasing greenhouse gases such as nitrous oxide that are mainly derived from nitrogen fertilizers. Therefore, understanding biological nitrogen fixation in farming systems is beneficial to agriculture and environmental preservation. In this context, a better grasp of nitrogen-fixing systems and nitrogen-fixing bacteria-plant associations will contribute to the optimization of these biological processes. Legumes and actinorhizal plants can engage in a symbiotic interaction with nitrogen-fixing rhizobia or actinomycetes, resulting in the formation of specialized root nodules. The legume-rhizobia interaction is mediated by a complex molecular signal exchange, where recognition of different bacterial determinants activates the nodulation program in the plant. To invade plants roots, bacteria follow different routes, which are determined by the host plant. Entrance via root hairs is probably the best understood. Alternatively, entry via intercellular invasion has been observed in many legumes. Although there are common features shared by intercellular infection mechanisms, differences are observed in the site of root invasion and bacterial spread on the cortex reaching and infecting a susceptible cell to form a nodule. This review focuses on intercellular bacterial invasion of roots observed in the Fabaceae and considers, within an evolutionary context, the different variants, distribution and molecular determinants involved. Intercellular invasion of actinorhizal plants and Parasponia is also discussed.

  10. Significance of intercellular spaces (windows) in effusion fluid cytology: a study of 46 samples.

    PubMed

    Murugan, Paari; Siddaraju, Neelaiah; Habeebullah, Syed; Basu, Debdatta

    2008-09-01

    The presence and nature of intercellular windows were studied on 46 body cavity fluid samples chosen on the basis of an unequivocal diagnosis on May-Grünwald-Giemsa (MGG)/Papanicolaou-stained smears and cell blocks. Of these, 24 cases had adenocarcinoma (AC) and seven had reactive mesothelium (RM) with 15 having distinct populations of both. Mucicarmine and PAS stains were used wherever indicated. The specificity of windows for predicting reactive mesothelium was evaluated. Intercellular windows were found in all cases of reactive mesothelium coinciding with the presence of the fuzzy peripheral microvillous borders. Surprisingly, as many as 17/39 (44%) of the adenocarcinomas also exhibited this feature, of which 13 had a distinctly visible evenly distributed ciliated cell membrane. In addition, 30/39 (77%) cases of AC exhibited a "window-like" appearance caused by cytoplasmic vacuolation. None of the adenocarcinoma clusters with true window formation showed positivity for the mucin stains, whereas all the clusters with pseudowindows caused by vacuolation were stained. Thus the specificity of intercellular windows for RM was merely 56%, though the sensitivity was 100%. On the other hand, the absence of windows was 100% specific for adenocarcinoma. Intercellular windows, though a feature of reactive mesothelial cell populations, can also be found in cases of ciliated adenocarcinomas and may not have a significant predictive value.

  11. Miraculin, a taste-modifying protein is secreted into intercellular spaces in plant cells.

    PubMed

    Hirai, Tadayoshi; Sato, Mayuko; Toyooka, Kiminari; Sun, Hyeon-Jin; Yano, Megumu; Ezura, Hiroshi

    2010-02-15

    A taste-modifying protein, miraculin, is highly accumulated in ripe fruit of miracle fruit (Richadella dulcifica) and the content can reach up to 10% of the total soluble protein in these fruits. Although speculated for decades that miraculin is secreted into intercellular spaces in miracle fruit, no evidence exists of its cellular localization. To study the cellular localization of miraculin in plant cells, using miracle fruit and transgenic tomato that constitutively express miraculin, immunoelectron microscopy, imaging GFP fusion proteins, and immunological detection of secreted proteins in culture medium of transgenic tomato were carried out. Immunoelectron microscopy showed the specific accumulation of miraculin in the intercellular layers of both miracle fruit and transgenic tomato. Imaging GFP fusion protein demonstrated that the miraculin-GFP fusion protein was accumulated in the intercellular spaces of tomato epidermal cells. Immunological detection of secreted proteins in culture medium of transgenic tomato indicated that miraculin was secreted from the roots of transgenic tomato expressing miraculin. This study firstly showed the evidences of the intercellular localization of miraculin, and provided a new insight of biological roles of miraculin in plants.

  12. Diabetes Increases Cryoinjury Size with Associated Effects on Cx43 Gap Junction Function and Phosphorylation in the Mouse Heart

    PubMed Central

    Palatinus, Joseph A.; Gourdie, Robert G.

    2016-01-01

    Diabetic patients develop larger myocardial infarctions and have an increased risk of death following a heart attack. The poor response to myocardial injury in the diabetic heart is likely related to the many metabolic derangements from diabetes that create a poor substrate in general for wound healing, response to injury and infection. Studies in rodents have implicated a role for the gap junction protein connexin 43 (Cx43) in regulating the injury response in diabetic skin wounds. In this study, we sought to determine whether diabetes alters Cx43 molecular interactions or intracellular communication in the cryoinjured STZ type I diabetic mouse heart. We found that epicardial cryoinjury size is increased in diabetic mice and this increase is prevented by preinjury insulin administration. Consistent with these findings, we found that intercellular coupling via gap junctions is decreased after insulin administration in diabetic and nondiabetic mice. This decrease in coupling is associated with a concomitant increase in phosphorylation of Cx43 at serine 368, a residue known to decrease channel conductance. Taken together, our results suggest that insulin regulates both gap junction-mediated intercellular communication and injury propagation in the mouse heart. PMID:27034963

  13. Connexin40, a component of gap junctions in vascular endothelium, is restricted in its ability to interact with other connexins.

    PubMed Central

    Bruzzone, R; Haefliger, J A; Gimlich, R L; Paul, D L

    1993-01-01

    The cellular distribution of connexin40 (Cx40), a newly cloned gap junction structural protein, was examined by immunofluorescence microscopy using two different specific anti-peptide antibodies. Cx40 was detected in the endothelium of muscular as well as elastic arteries in a punctate pattern consistent with the known distribution of gap junctions. However, it was not detected in other cells of the vascular wall. By contrast, Cx43, another connexin present in the cardiovascular system, was not detected in endothelial cells of muscular arteries but was abundant in the myocardium and aortic smooth muscle. We have tested the ability of these connexins to interact functionally. Cx40 was functionally expressed in pairs of Xenopus oocytes and induced the formation of intercellular channels with unique voltage dependence. Unexpectedly, communication did not occur when oocytes expressing Cx40 were paired with those expressing Cx43, although each could interact with a different connexin, Cx37, to form gap junction channels in paired oocytes. These findings indicate that establishment of intercellular communication can be spatially regulated by the selective expression of different connexins and suggest a mechanism that may operate to control the extent of communication between cells. Images PMID:8382974

  14. Thermal conductance of superlattice junctions

    SciTech Connect

    Lu, Simon; McGaughey, Alan J. H.

    2015-05-15

    We use molecular dynamics simulations and the lattice-based scattering boundary method to compute the thermal conductance of finite-length Lennard-Jones superlattice junctions confined by bulk crystalline leads. The superlattice junction thermal conductance depends on the properties of the leads. For junctions with a superlattice period of four atomic monolayers at temperatures between 5 and 20 K, those with mass-mismatched leads have a greater thermal conductance than those with mass-matched leads. We attribute this lead effect to interference between and the ballistic transport of emergent junction vibrational modes. The lead effect diminishes when the temperature is increased, when the superlattice period is increased, and when interfacial disorder is introduced, but is reversed in the harmonic limit.

  15. Josephson junction Q-spoiler

    DOEpatents

    Clarke, J.; Hilbert, C.; Hahn, E.L.; Sleator, T.

    1986-03-25

    An automatic Q-spoiler comprising at least one Josephson tunnel junction connected in an LC circuit for flow of resonant current therethrough. When in use in a system for detecting the magnetic resonance of a gyromagnetic particle system, a high energy pulse of high frequency energy irradiating the particle system will cause the critical current through the Josephson tunnel junctions to be exceeded, causing the tunnel junctions to act as resistors and thereby damp the ringing of the high-Q detection circuit after the pulse. When the current has damped to below the critical current, the Josephson tunnel junctions revert to their zero-resistance state, restoring the Q of the detection circuit and enabling the low energy magnetic resonance signals to be detected.

  16. Josephson junction Q-spoiler

    DOEpatents

    Clarke, John; Hilbert, Claude; Hahn, Erwin L.; Sleator, Tycho

    1988-01-01

    An automatic Q-spoiler comprising at least one Josephson tunnel junction connected in an LC circuit for flow of resonant current therethrough. When in use in a system for detecting the magnetic resonance of a gyromagnetic particle system, a high energy pulse of high frequency energy irradiating the particle system will cause the critical current through the Josephson tunnel junctions to be exceeded, causing the tunnel junctions to act as resistors and thereby damp the ringing of the high-Q detection circuit after the pulse. When the current has damped to below the critical current, the Josephson tunnel junctions revert to their zero-resistance state, restoring the Q of the detection circuit and enabling the low energy magnetic resonance signals to be detected.

  17. Control of Junction Flow

    NASA Astrophysics Data System (ADS)

    Su, T.-C.; Bingham, C.; Kellier, L.

    2001-11-01

    Control for horseshoe vortices resulting from boundary layer separation in front of a structure has long been sought without satisfactory results. Tests were carried out in a water channel with the objective of seeking such a control. The water channel has a test section of .6m wide, .4m deep and 8m long, with an adjustable mean flow speed of up to .5m/s. Flow visualization technique was used to elucidate the flow process. To control the horseshoe vortex a long airfoil of 1cm chord was placed horizontally near the ground upstream of a 10cm thin square plate. It was found that the original horseshoe vortex moved toward and circulated around the airfoil. The junction flow immediately upstream of the obstacle was noticeably steady and free of disturbance. The process was insensitive to the streamwise location of the airfoil, horseshoe's vortical structure, stream speed and acceleration, upstream vortical influx, and magnitude/sign of airfoil's angle of attack. Experimental results with obliquely mounted square cylinder were similar, which demonstrated that controls were effective for all angles of attack.

  18. Charge Transport Processes in Molecular Junctions

    NASA Astrophysics Data System (ADS)

    Smith, Christopher Eugene

    Molecular electronics (ME) has evolved into a rich area of exploration that combines the fields of chemistry, materials, electronic engineering and computational modeling to explore the physics behind electronic conduction at the molecular level. Through studying charge transport properties of single molecules and nanoscale molecular materials the field has gained the potential to bring about new avenues for the miniaturization of electrical components where quantum phenomena are utilized to achieve solid state molecular device functionality. Molecular junctions are platforms that enable these studies and consist of a single molecule or a small group of molecules directly connected to electrodes. The work presented in this thesis has built upon the current understanding of the mechanisms of charge transport in ordered junctions using self-assembled monolayer (SAM) molecular thin films. Donor and acceptor compounds were synthesized and incorporated into SAMs grown on metal substrates then the transport properties were measured with conducting probe atomic force microscopy (CP-AFM). In addition to experimentally measured current-voltage (I-V) curves, the transport properties were addressed computationally and modeled theoretically. The key objectives of this project were to 1) investigate the impact of molecular structure on hole and electron charge transport, 2) understand the nature of the charge carriers and their structure-transport properties through long (<4 nm) conjugated molecular wires, and 3) quantitatively extract interfacial properties characteristic to macroscopic junctions, such as energy level alignment and molecule-contact electronic coupling from experimental I-V curves. Here, we lay ground work for creating a more complete picture of charge transport in macroscopically ordered molecular junctions of controlled architecture, length and charge carrier. The polaronic nature of hopping transport has been predicted in long, conjugated molecular wires

  19. Stimulating the Release of Exosomes Increases the Intercellular Transfer of Prions.

    PubMed

    Guo, Belinda B; Bellingham, Shayne A; Hill, Andrew F

    2016-03-04

    Exosomes are small extracellular vesicles released by cells and play important roles in intercellular communication and pathogen transfer. Exosomes have been implicated in several neurodegenerative diseases, including prion disease and Alzheimer disease. Prion disease arises upon misfolding of the normal cellular prion protein, PrP(C), into the disease-associated isoform, PrP(Sc). The disease has a unique transmissible etiology, and exosomes represent a novel and efficient method for prion transmission. The precise mechanism by which prions are transmitted from cell to cell remains to be fully elucidated, although three hypotheses have been proposed: direct cell-cell contact, tunneling nanotubes, and exosomes. Given the reported presence of exosomes in biological fluids and in the lipid and nucleic acid contents of exosomes, these vesicles represent an ideal mechanism for encapsulating prions and potential cofactors to facilitate prion transmission. This study investigates the relationship between exosome release and intercellular prion dissemination. Stimulation of exosome release through treatment with an ionophore, monensin, revealed a corresponding increase in intercellular transfer of prion infectivity. Conversely, inhibition of exosome release using GW4869 to target the neutral sphingomyelinase pathway induced a decrease in intercellular prion transmission. Further examination of the effect of monensin on PrP conversion revealed that monensin also alters the conformational stability of PrP(C), leading to increased generation of proteinase K-resistant prion protein. The findings presented here provide support for a positive relationship between exosome release and intercellular transfer of prion infectivity, highlighting an integral role for exosomes in facilitating the unique transmissible nature of prions.

  20. Electronic thermometry in tunable tunnel junction

    DOEpatents

    Maksymovych, Petro

    2016-03-15

    A tunable tunnel junction thermometry circuit includes a variable width tunnel junction between a test object and a probe. The junction width is varied and a change in thermovoltage across the junction with respect to the change in distance across the junction is determined. Also, a change in biased current with respect to a change in distance across the junction is determined. A temperature gradient across the junction is determined based on a mathematical relationship between the temperature gradient, the change in thermovoltage with respect to distance and the change in biased current with respect to distance. Thermovoltage may be measured by nullifying a thermoelectric tunneling current with an applied voltage supply level. A piezoelectric actuator may modulate the probe, and thus the junction width, to vary thermovoltage and biased current across the junction. Lock-in amplifiers measure the derivatives of the thermovoltage and biased current modulated by varying junction width.

  1. Cooperativity-based modeling of heterotypic DNA nanostructure assembly.

    PubMed

    Shapiro, Anastasia; Hozeh, Avital; Girshevitz, Olga; Abu-Horowitz, Almogit; Bachelet, Ido

    2015-07-27

    DNA origami is a robust method for the fabrication of nanoscale 2D and 3D objects with complex features and geometries. The process of DNA origami folding has been recently studied, however quantitative understanding of it is still elusive. Here, we describe a systematic quantification of the assembly process of DNA nanostructures, focusing on the heterotypic DNA junction-in which arms are unequal-as their basic building block. Using bulk fluorescence studies we tracked this process and identified multiple levels of cooperativity from the arms in a single junction to neighboring junctions in a large DNA origami object, demonstrating that cooperativity is a central underlying mechanism in the process of DNA nanostructure assembly. We show that the assembly of junctions in which the arms are consecutively ordered is more efficient than junctions with randomly-ordered components, with the latter showing assembly through several alternative trajectories as a potential mechanism explaining the lower efficiency. This highlights consecutiveness as a new design consideration that could be implemented in DNA nanotechnology CAD tools to produce more efficient and high-yield designs. Altogether, our experimental findings allowed us to devise a quantitative, cooperativity-based heuristic model for the assembly of DNA nanostructures, which is highly consistent with experimental observations.

  2. Constructing higher order DNA origami arrays using DNA junctions of anti-parallel/parallel double crossovers

    NASA Astrophysics Data System (ADS)

    Ma, Zhipeng; Park, Seongsu; Yamashita, Naoki; Kawai, Kentaro; Hirai, Yoshikazu; Tsuchiya, Toshiyuki; Tabata, Osamu

    2016-06-01

    DNA origami provides a versatile method for the construction of nanostructures with defined shape, size and other properties; such nanostructures may enable a hierarchical assembly of large scale architecture for the placement of other nanomaterials with atomic precision. However, the effective use of these higher order structures as functional components depends on knowledge of their assembly behavior and mechanical properties. This paper demonstrates construction of higher order DNA origami arrays with controlled orientations based on the formation of two types of DNA junctions: anti-parallel and parallel double crossovers. A two-step assembly process, in which preformed rectangular DNA origami monomer structures themselves undergo further self-assembly to form numerically unlimited arrays, was investigated to reveal the influences of assembly parameters. AFM observations showed that when parallel double crossover DNA junctions are used, the assembly of DNA origami arrays occurs with fewer monomers than for structures formed using anti-parallel double crossovers, given the same assembly parameters, indicating that the configuration of parallel double crossovers is not energetically preferred. However, the direct measurement by AFM force-controlled mapping shows that both DNA junctions of anti-parallel and parallel double crossovers have homogeneous mechanical stability with any part of DNA origami.

  3. Octagonal Defects at Carbon Nanotube Junctions

    PubMed Central

    Jaskólski, W.; Pelc, M.; Chico, Leonor; Ayuela, A.

    2013-01-01

    We investigate knee-shaped junctions of semiconductor zigzag carbon nanotubes. Two dissimilar octagons appear at such junctions; one of them can reconstruct into a pair of pentagons. The junction with two octagons presents two degenerate localized states at Fermi energy (EF). The reconstructed junction has only one state near EF, indicating that these localized states are related to the octagonal defects. The inclusion of Coulomb interaction splits the localized states in the junction with two octagons, yielding an antiferromagnetic system. PMID:24089604

  4. Modulation of human cell responses to space radiation by gap-junction communication

    NASA Astrophysics Data System (ADS)

    Autsavapromporn, Narongchai; de Toledo, Sonia M.; Buonanno, Manuela; Yang, Zhi; Harris, Andrew; Jay-Gerin, Jean-Paul; Azzam, Edouard

    Understanding the biological effects of space radiation and their underlying mechanism is critical to estimating the health risk associated with human exploration of space. A coordinated interaction of multiple cellular processes is likely involved in the sensing and processing of stressful effects induced by different types of space radiation. Here, we focused on the role of gap-junction intercellular communication (GJIC) in responses of human cells exposed to 1 GeV/n protons or 56 Fe-ions. We compared the results with data obtained in human cells exposed, in parallel, to γ-rays or α-particles. As expected, a higher level of cell killing and DNA damage, per unit dose, was induced in confluent, density-inhibited cells (98% in G0 /G1 ) exposed to α-particles or energetic 56 Fe-ions than γ-rays or protons. Strikingly, greatly attenuated effects occurred when sub-confluent cultures, synchronized in G0 /G1 ,were exposed to 56 Fe-ions. These data suggest that direct intercellular communication is involved in the effects of high linear energy transfer (LET) 56 Fe-ions. To examine the role of gap-junctions in propagating stressful effect, confluent cultures were exposed to 56 Fe-ions or α-particles and incubated for various time periods at 37° C in the presence or absence of the gap-junction inhibitor α-glycyrrhetinic acid (AGA). No repair of potentially lethal radiation damage occurred in cells incubated in the absence of AGA. In contrast, inhibition of functional GJIC significantly enhanced clonogenic survival of irradiated cells. To test the role of junctional channel permeability in the observed effects, we used human adenocarcinoma (HeLa) cells in which specific connexins (Cx) can be expressed in the absence of endogenous connexins. Whereas HeLa cells with selective inducible expression of Cx26 gap-junctions promoted radiation toxic effects, expression of Cx32 junctional channels in HeLa cells promoted pro-survival effects. Experiments are in progress to

  5. Intercellular ice propagation: experimental evidence for ice growth through membrane pores.

    PubMed

    Acker, J P; Elliott, J A; McGann, L E

    2001-09-01

    Propagation of intracellular ice between cells significantly increases the prevalence of intracellular ice in confluent monolayers and tissues. It has been proposed that gap junctions facilitate ice propagation between cells. This study develops an equation for capillary freezing-point depression to determine the effect of temperature on the equilibrium radius of an ice crystal sufficiently small to grow through gap junctions. Convection cryomicroscopy and video image analysis were used to examine the incidence and pattern of intracellular ice formation (IIF) in the confluent monolayers of cell lines that do (MDCK) and do not (V-79W) form gap junctions. The effect of gap junctions on intracellular ice propagation was strongly temperature-dependent. For cells with gap junctions, IIF occurred in a directed wave-like pattern in 100% of the cells below -3 degrees C. At temperatures above -3 degrees C, there was a marked drop in the incidence of IIF, with isolated individual cells initially freezing randomly throughout the sample. This random pattern of IIF was also observed in the V-79W monolayers and in MDCK monolayers treated to prevent gap junction formation. The significant change in the low temperature behavior of confluent MDCK monolayers at -3 degrees C is likely the result of the inhibition of gap junction-facilitated ice propagation, and supports the theory that gap junctions facilitate ice nucleation between cells.

  6. Regulation of gap junction function and Connexin 43 expression by cytochrome P450 oxidoreductase (CYPOR)

    SciTech Connect

    Polusani, Srikanth R.; Kar, Rekha; Riquelme, Manuel A.; Masters, Bettie Sue; Panda, Satya P.

    2011-08-05

    Highlights: {yields} Humans with severe forms of cytochrome P450 oxidoreductase (CYPOR) mutations show bone defects as observed in Antley-Bixler Syndrome. {yields} First report showing knockdown of CYPOR in osteoblasts decreased Connexin 43 (Cx43) protein levels. Cx43 is known to play an important role in bone modeling. {yields} Knockdown of CYPOR decreased Gap Junctional Intercellular Communication and hemichannel activity. {yields} Knockdown of CYPOR decreased Cx43 in mouse primary calvarial osteoblasts. {yields} Decreased Cx43 expression was observed at the transcriptional level. -- Abstract: Cytochrome P450 oxidoreductase (CYPOR) is a microsomal electron-transferring enzyme containing both FAD and FMN as co-factors, which provides the reducing equivalents to various redox partners, such as cytochromes P450 (CYPs), heme oxygenase (HO), cytochrome b{sub 5} and squalene monooxygenase. Human patients with severe forms of CYPOR mutation show bone defects such as cranio- and humeroradial synostoses and long bone fractures, known as Antley-Bixler-like Syndrome (ABS). To elucidate the role of CYPOR in bone, we knocked-down CYPOR in multiple osteoblast cell lines using RNAi technology. In this study, knock-down of CYPOR decreased the expression of Connexin 43 (Cx43), known to play a critical role in bone formation, modeling, and remodeling. Knock-down of CYPOR also decreased Gap Junction Intercellular Communication (GJIC) and hemichannel activity. Promoter luciferase assays revealed that the decrease in expression of Cx43 in CYPOR knock-down cells was due to transcriptional repression. Primary osteoblasts isolated from bone specific Por knock-down mice calvariae confirmed the findings in the cell lines. Taken together, our study provides novel insights into the regulation of gap junction function by CYPOR and suggests that Cx43 may play an important role(s) in CYPOR-mediated bone defects seen in patients.

  7. ATP counteracts the rundown of gap junctional channels of rat ventricular myocytes by promoting protein phosphorylation.

    PubMed

    Verrecchia, F; Duthe, F; Duval, S; Duchatelle, I; Sarrouilhe, D; Herve, J C

    1999-04-15

    1. The degree of cell-to-cell coupling between ventricular myocytes of neonatal rats appeared well preserved when studied in the perforated version of the patch clamp technique or, in double whole-cell conditions, when ATP was present in the patch pipette solution. In contrast, when ATP was omitted, the amplitude of junctional current rapidly declined (rundown). 2. To examine the mechanism(s) of ATP action, an 'internal perfusion technique' was adapted to dual patch clamp conditions, and reintroduction of ATP partially reversed the rundown of junctional channels. 3. Cell-to-cell communication was not preserved by a non-hydrolysable ATP analogue (5'-adenylimidodiphosphate, AMP-PNP), indicating that the effect most probably did not involve direct interaction of ATP with the channel-forming proteins. 4. An ATP analogue supporting protein phosphorylation but not active transport processes (adenosine 5'-O-(3-thiotriphosphate), ATPgammaS) maintained normal intercellular communication, suggesting that the effect was due to kinase activity rather than to altered intracellular Ca2+. 5. A broad spectrum inhibitor of endogenous serine/threonine protein kinases (H7) reversibly reduced the intercellular coupling. A non-specific exogenous protein phosphatase (alkaline phosphatase) mimicked the effects of ATP deprivation. The non-specific inhibition of endogenous protein phosphatases resulted in the preservation of substantial cell-to-cell communication in ATP-free conditions. 6. The activity of gap junctional channels appears to require both the presence of ATP and protein kinase activity to counteract the tonic activity of endogenous phosphatase(s).

  8. Influenza A Virus Uses Intercellular Connections To Spread to Neighboring Cells

    PubMed Central

    Roberts, Kari L.; Manicassamy, Balaji

    2014-01-01

    ABSTRACT In the extracellular environment, cell-free virions seek out naive host cells over long distances and between organisms. This is the primary mechanism of spread for most viruses. Here we provide evidence for an alternative pathway previously undescribed for orthomyxoviruses, whereby the spread of influenza A virus (IAV) infectious cores to neighboring cells can occur within intercellular connections. The formation of these connections requires actin dynamics and is enhanced by viral infection. Connected cells have contiguous membranes, and the core infectious viral machinery (RNP and polymerase) was present inside the intercellular connections. A live-cell movie of green fluorescent protein (GFP)-tagged NS1 of IAV shows viral protein moving from one cell to another through an intercellular connection. The movement of tagged protein was saltatory but overall traveled only in one direction. Infectious virus cores can move from one cell to another without budding and release of cell-free virions, as evidenced by the finding that whereas a neuraminidase inhibitor alone did not inhibit the development of IAV microplaques, the presence of a neuraminidase inhibitor together with drugs inhibiting actin dynamics or the microtubule stabilizer paclitaxel (originally named taxol) precluded microplaque formation. Similar results were also observed with parainfluenza virus 5 (PIV5), a paramyxovirus, when neutralizing antibody was used to block spread by cell-free virions. Intercellular spread of infectious core particles was unaffected or enhanced in the presence of nocodazole for IAV but inhibited for PIV5. The intercellular connections have a core of filamentous actin, which hints toward transport of virus particles through the use of a myosin motor. IMPORTANCE Here we describe a new method by which influenza A virus (IAV) spreads from cell to cell: IAV uses intracellular connections. The formation of these connections requires actin dynamics and is enhanced by viral

  9. Endocytosis of Epithelial Apical Junctional Proteins by a Clathrin-mediated Pathway into a Unique Storage Compartment

    PubMed Central

    Ivanov, Andrei I.; Nusrat, Asma; Parkos, Charles A.

    2004-01-01

    The adherens junction (AJ) and tight junction (TJ) are key regulators of epithelial polarity and barrier function. Loss of epithelial phenotype is accompanied by endocytosis of AJs and TJs via unknown mechanisms. Using a model of calcium depletion, we defined the pathway of internalization of AJ and TJ proteins (E-cadherin, p120 and β-catenins, occludin, JAM-1, claudins 1 and 4, and ZO-1) in T84 epithelial cells. Proteinase protection assay and immunocytochemistry revealed orchestrated internalization of AJs and TJs into a subapical cytoplasmic compartment. Disruption of caveolae/lipid rafts did not prevent endocytosis, nor did caveolin-1 colocalize with internalized junctional proteins. Furthermore, AJ and TJ proteins did not colocalize with the macropinocytosis marker dextran. Inhibitors of clathrin-mediated endocytosis blocked internalization of AJs and TJs, and junctional proteins colocalized with clathrin and α-adaptin. AJ and TJ proteins were observed to enter early endosomes followed by movement to organelles that stained with syntaxin-4 but not with markers of late and recycling endosomes, lysosomes, or Golgi. These results indicate that endocytosis of junctional proteins is a clathrin-mediated process leading into a unique storage compartment. Such mechanisms may mediate the disruption of intercellular contacts during normal tissue remodeling and in pathology. PMID:14528017

  10. Expression of functional gap junctions and regulation by fluid flow in osteocyte-like MLO-Y4 cells.

    PubMed

    Cheng, B; Zhao, S; Luo, J; Sprague, E; Bonewald, L F; Jiang, J X

    2001-02-01

    Osteocytes are thought to be mechanosensory cells that respond to mechanical stress by sending signals to other bone cells to initiate bone remodeling. An osteocyte-like cell line MLO-Y4 provides a model system to examine whether gap junctions participate in the regulation of osteocyte function and signaling by mechanical stress. In this study, we show that MLO-Y4 cells are coupled and that gap junction channels mediate this coupling. Biochemical analyses show that connexin 43 (Cx43) is a major gap junction protein expressed in MLO-Y4 cells and approximately 5% of Cx43 protein is phosphorylated. MLO-Y4 cells were exposed to mechanical stress using a parallel plate flow chamber to model bone fluid flow shear stress. Fluid flow increased significantly the length of the dendritic processes, a morphological characteristic of osteocytes. A redistribution of the gap junction protein, Cx43 also was observed from a location circling the nucleus to punctate spots in the cytoplasm and in the dendritic processes. "Scrape-loading" dye transfer analyses showed that fluid flow increased intercellular coupling and increased the number of cells coupled immediately after fluid flow treatment, in direct proportion to shear stress magnitude. Although intercellular coupling continued to increase, stimulation of Cx43 protein expression during the poststress period was found to be biphasic. Cx43 protein was elevated 30 minutes after application of stress but decreased at 24 h poststress. Pulsating fluid flow had a similar stimulatory effect as steady fluid flow on gap junctions. However, this stimulatory effect in osteocyte-like cells was not observed in osteoblast-like 2T3 cells. Together, these results show that fluid flow has stimulatory effects on osteocyte-like MLO-Y4 cells with early effects on cellular morphology, opening of gap junctions, and redistribution of Cx43 protein and delayed effects on Cx43 protein expression. The high expression of Cx43 and its location in the

  11. Effects of mechanical strain on the function of Gap junctions in osteocytes are mediated through the prostaglandin EP2 receptor.

    PubMed

    Cherian, Priscilla P; Cheng, Benxu; Gu, Sumin; Sprague, Eugene; Bonewald, Lynda F; Jiang, Jean X

    2003-10-31

    Osteocytes embedded in the matrix of bone are thought to be mechanosensory cells that translate mechanical strain into biochemical signals that regulate bone modeling and remodeling. We have shown previously that fluid flow shear stress dramatically induces prostaglandin release and COX-2 mRNA expression in osteocyte-like MLO-Y4 cells, and that prostaglandin E2 (PGE2) released by these cells functions in an autocrine manner to regulate gap junction function and connexin 43 (Cx43) expression. Here we show that fluid flow regulates gap junctions through the PGE2 receptor EP2 activation of cAMP-dependent protein kinase A (PKA) signaling. The expression of the EP2 receptor, but not the subtypes EP1,EP3, and EP4, increased in response to fluid flow. Application of PGE2 or conditioned medium from fluid flow-treated cells to non-stressed MLO-Y4 cells increased expression of the EP2 receptor. The EP2 receptor antagonist, AH6809, suppressed the stimulatory effects of PGE2 and fluid flow-conditioned medium on the expression of the EP2 receptor, on Cx43 protein expression, and on gap junction-mediated intercellular coupling. In contrast, the EP2 receptor agonist butaprost, not the E1/E3 receptor agonist sulprostone, stimulated the expression of Cx43 and gap junction function. Fluid flow conditioned medium and PGE2 stimulated cAMP production and PKA activity suggesting that PGE2 released by mechanically stimulated cells is responsible for the activation of cAMP and PKA. The adenylate cyclase activators, forskolin and 8-bromo-cAMP, enhanced intercellular connectivity, the number of functional gap junctions, and Cx43 protein expression, whereas the PKA inhibitor, H89, inhibited the stimulatory effect of PGE2 on gap junctions. These studies suggest that the EP2 receptor mediates the effects of autocrine PGE2 on the osteocyte gap junction in response to fluid flow-induced shear stress. These data support the hypothesis that the EP2 receptor, cAMP, and PKA are critical components

  12. Automated Array Assembly, Phase 2

    NASA Technical Reports Server (NTRS)

    Carbajal, B. G.

    1979-01-01

    The Automated Array Assembly Task, Phase 2 of the Low Cost Silicon Solar Array Project is a process development task. The contract provides for the fabrication of modules from large area tandem junction cells (TJC). During this quarter, effort was focused on the design of a large area, approximately 36 sq cm, TJC and process verification runs. The large area TJC design was optimized for minimum I squared R power losses. In the TJM activity, the cell-module interfaces were defined, module substrates were formed and heat treated and clad metal interconnect strips were fabricated.

  13. Crew Assembly

    NASA Video Gallery

    Train to improve your dexterity and hand-eye coordination by assembling a puzzle.The Train Like an Astronaut project uses the excitement of exploration to challenge students to set goals, practice ...

  14. CROSS-DISCIPLINARY PHYSICS AND RELATED AREAS OF SCIENCE AND TECHNOLOGY: Modelling of intercellular synchronization in the Drosophila circadian clock

    NASA Astrophysics Data System (ADS)

    Wang, Jun-Wei; Chen, Ai-Min; Zhang, Jia-Jun; Yuan, Zhan-Jiang; Zhou, Tian-Shou

    2009-03-01

    In circadian rhythm generation, intercellular signaling factors are shown to play a crucial role in both sustaining intrinsic cellular rhythmicity and acquiring collective behaviours across a population of circadian neurons. However, the physical mechanism behind their role remains to be fully understood. In this paper, we propose an indirectly coupled multicellular model for the synchronization of Drosophila circadian oscillators combining both intracellular and intercellular dynamics. By simulating different experimental conditions, we find that such an indirect coupling way can synchronize both heterogeneous self-sustained circadian neurons and heterogeneous mutational damped circadian neurons. Moreover, they can also be entrained to ambient light-dark (LD) cycles depending on intercellular signaling.

  15. A phospholipid kinase regulates actin organization and intercellular bridge formation during germline cytokinesis.

    PubMed

    Brill, J A; Hime, G R; Scharer-Schuksz, M; Fuller, M T

    2000-09-01

    The endgame of cytokinesis can follow one of two pathways depending on developmental context: resolution into separate cells or formation of a stable intercellular bridge. Here we show that the four wheel drive (fwd) gene of Drosophila melanogaster is required for intercellular bridge formation during cytokinesis in male meiosis. In fwd mutant males, contractile rings form and constrict in dividing spermatocytes, but cleavage furrows are unstable and daughter cells fuse together, producing multinucleate spermatids. fwd is shown to encode a phosphatidylinositol 4-kinase (PI 4-kinase), a member of a family of proteins that perform the first step in the synthesis of the key regulatory membrane phospholipid PIP2. Wild-type activity of the fwd PI 4-kinase is required for tyrosine phosphorylation in the cleavage furrow and for normal organization of actin filaments in the constricting contractile ring. Our results suggest a critical role for PI 4-kinases and phosphatidylinositol derivatives during the final stages of cytokinesis.

  16. Exosomes: vehicles of intercellular signaling, biomarkers, and vectors of cell therapy.

    PubMed

    Kourembanas, Stella

    2015-01-01

    Mesenchymal stem cells (MSCs), whose mechanism of action is predominantly paracrine, are being widely tested for the treatment of a variety of human diseases. No one factor has been proven sufficient to mediate the therapeutic effects of MSCs. However, exosomes--membrane vesicles secreted by many cells, including MSCs--are appealing candidates as vectors of their efficacy. Exosomes can transport and deliver a large cargo of proteins, lipids, and nucleic acids and can modify cell and organ function. In addition to their key role as vehicles of intercellular communication, exosomes are increasingly recognized as biomarkers and prognosticators of disease. Moreover, they have the potential to be used as vehicles of gene and drug delivery for clinical application. This article reviews the biogenesis of exosomes, their molecular composition, and their role as messengers of intercellular communication, focusing on their potential as therapeutic vectors for stem cell therapy.

  17. Effects of dinitrotoluenes on morphological cell transformation and intercellular communication in Syrian hamster embryo cells.

    PubMed

    Holen, I; Mikalsen, S O; Sanner, T

    1990-01-01

    The effects of four isomers of dinitrotoluene (DNT) and technical DNT (a mixture of DNT isomers and other compounds, with 2,4-DNT as the major constituent) were studied in two short-term in vitro assays. None of the isomers or technical DNT induced an increase in morphological transformation of Syrian hamster embryo (SHE) cells. Four DNT metabolites (2,4-diaminotoluene, 2-amino-4-nitrotoluene, 2-amino-6-nitrotoluene, and 2,4-dinitobenzoic acid), representing different stages in reduction or oxidation of DNT isomers, were also negative for induction of morphological transformation. The DNT isomers were tested in an intercellular communication assay based on dye transfer. 2,4-DNT, 2,6-DNT, and technical DNT inhibited intercellular communication in the SHE cell line BPNi at toxic concentrations. This may be reminiscent of in vivo data showing promoting activity of these compound. 2,3-DNT and 3,4-DNT did not inhibit communication.

  18. An intercellular heme trafficking protein delivers maternal heme to the embryo during development in C. elegans

    PubMed Central

    Chen, Caiyong; Samuel, Tamika K.; Sinclair, Jason; Dailey, Harry A.; Hamza, Iqbal

    2011-01-01

    Summary Heme is a cytotoxic, hydrophobic tetrapyrrole that crosses multiple biological membranes for incorporation into proteins critical for numerous biological processes. Thus, a prima facie argument can be made that heme trafficking within the aqueous cellular milieu must be mediated by specific intra- and intercellular pathways. Embryonic development in Caenorhabditis elegans, a heme auxotroph, is inextricably dependent on maternal heme acquisition. Here we show that HRG-3 is required to deliver maternal heme to oocytes for zygotic development. HRG-3 binds heme and is exclusively secreted by the intestine during heme insufficiency into the interstitial fluid for transport of maternal heme to extra-intestinal cells. HRG-3 deficiency results either in death during embryogenesis or in developmental arrest immediately post hatching – phenotypes that are fully suppressed by maternal but not zygotic hrg-3 expression. Our results establish an unprecedented role for HRG-3 as an intercellular heme chaperone in zygotic development and maternal-embryonic nutrition in C. elegans. PMID:21620137

  19. Detection of cancerous kidney tissue areas by means of infrared spectroscopy of intercellular fluid

    NASA Astrophysics Data System (ADS)

    Urboniene, V.; Jankevicius, F.; Zelvys, A.; Steiner, G.; Sablinskas, V.

    2014-03-01

    In this work the infrared absorption spectra of intercellular fluid of normal and tumor kidney tissue were recorded and analyzed. The samples were prepared by stamping freshly resected tissue onto a CaF2 substrate. FT-IR spectra obtained from intracellular fluid of tumor tissue exhibit stronger absorption bands in the spectral region from 1000-1200 cm-1 and around 1750 cm-1 than those obtained from normal tissue. It is likely the spectra of extracellular matrix of kidney tumor tissue with large increases in the intensities of these bands represent a higher concentration of fatty acids and glycerol. Amide I and amide II bands are stronger in spectra of normal tissue indicating a higher level of proteins. The results demonstrate that FT-IR spectroscopy of intercellular fluids is a novel approach for a quick diagnosis during surgical resection, which can improve the therapy of kidney tumors.

  20. Effect of cyto/chemokine degradation in effective intercellular communication distances

    NASA Astrophysics Data System (ADS)

    Gupta, V. K.

    2017-02-01

    Many complex biological processes such as cell differentiation, cell proliferation, and cell motility are governed by cell signaling. This mode of intercellular communication is of paramount importance for tissue function and ultimately for entire organism. In intercellular communication cells secrete signaling molecules such as cyto/chemokines which diffuse through the surrounding medium and eventually bind to receptors on other cells whereby the signal transduction is completed. An accurate estimation of the effective communication distances and the time scale on which signaling takes place are important for the interpretation of cell and organ physiology and ultimately in the effective and efficient chemotactically driven tissue engineering. The present study uses a solitary cell model incorporating degradation of secreted molecules to estimate the effective communication distances and the time scale on which signaling takes place. We demonstrate through our model that in presence of degradation the effective communication distances are significantly reduced.

  1. Seal assembly

    DOEpatents

    Johnson, Roger Neal; Longfritz, William David

    2001-01-01

    A seal assembly that seals a gap formed by a groove comprises a seal body, a biasing element, and a connection that connects the seal body to the biasing element to form the seal assembly. The seal assembly further comprises a concave-shaped center section and convex-shaped contact portions at each end of the seal body. The biasing element is formed from an elastic material and comprises a convex-shaped center section and concave-shaped biasing zones that are opposed to the convex-shaped contact portions. The biasing element is adapted to be compressed to change a width of the seal assembly from a first width to a second width that is smaller than the first width. In the compressed state, the seal assembly can be disposed in the groove. After release of the compressing force, the seal assembly expands. The contact portions will move toward a surface of the groove and the biasing zones will move into contact with another surface of the groove. The biasing zones will bias the contact portions of the seal body against the surface of the groove.

  2. The Occurrence of Intercellular Bridges in Groups of Cells Exhibiting Synchronous Differentiation

    PubMed Central

    Fawcett, Don W.; Ito, Susumu; Slautterback, David

    1959-01-01

    A previous electron microscopic study of the cat testis revealed that spermatids derived from the same spermatogonium are joined together by intercellular bridges. The present paper records the observation of similar connections between spermatocytes and between spermatids in Hydra, fruit-fly, opossum, pigeon, rat, hamster, guinea pig, rabbit, monkey, and man. In view of these findings, it is considered likely that a syncytial relationship within groups of developing male germ cells is of general occurrence and is probably responsible for their synchronous differentiation. When clusters of spermatids, freshly isolated from the germinal epithelium are observed by phase contrast microscopy, the constrictions between the cellular units of the syncytium disappear and the whole group coalesces into a spherical multinucleate mass. The significance of this observation in relation to the occurrence of abnormal spermatozoa in semen and the prevalence of multinucleate giant cells in pathological testes is discussed. In the ectoderm of Hydra, the clusters of cnidoblasts that arise from proliferation of interstitial cells are also connected by intercellular bridges. The development of nematocysts within these groups of conjoined cells is precisely synchronized. Both in the testis of vertebrates and the ectoderm of Hydra, a syncytium results from incomplete cytokinesis in the proliferation of relatively undifferentiated cells. The intercellular bridges between daughter cells are formed when the cleavage furrow encounters the spindle remnant and is arrested by it. The subsequent dissolution of the spindle filaments establishes free communication between the cells. The discovery of intercellular bridges in the two unrelated tissues discussed here suggests that a similar syncytial relationship may be found elsewhere in nature where groups of cells of common origin differentiate synchronously. PMID:13664686

  3. Noncontact estimation of intercellular breaking force using a femtosecond laser impulse quantified by atomic force microscopy.

    PubMed

    Hosokawa, Yoichiroh; Hagiyama, Man; Iino, Takanori; Murakami, Yoshinori; Ito, Akihiko

    2011-02-01

    When a femtosecond laser pulse (fsLP) is focused through an objective lens into a culture medium, an impulsive force (fsLP-IF) is generated that propagates from the laser focal point (O(f)) in a micron-sized space. This force can detach individual adherent cells without causing considerable cell damage. In this study, an fsLP-IF was reflected in the vibratory movement of an atomic force microscopy (AFM) cantilever. Based on the magnitude of the vibration and the geometrical relationship between O(f) and the cantilever, the fsLP-IF generated at O(f) was calculated as a unit of impulse [N-s]. This impulsive force broke adhesion molecule-mediated intercellular interactions in a manner that depended on the adhesion strength that was estimated by the cell aggregation assay. The force also broke the interactions between streptavidin-coated microspheres and a biotin-coated substrate with a measurement error of approximately 7%. These results suggest that fsLP-IF can be used to break intermolecular and intercellular interactions and estimate the adhesion strength. The fsLP-IF was used to break intercellular contacts in two biologically relevant cultures: a coculture of leukocytes seeded over on an endothelial cell monolayer, and a polarized monolayer culture of epithelial cells. The impulses needed to break leukocyte-endothelial and interepithelial interactions, which were calculated based on the geometrical relationship between O(f) and the adhesive interface, were on the order of 10(-13) and 10(-12) N-s, respectively. When the total impulse at O(f) is well-defined, fsLP-IF can be used to estimate the force required to break intercellular adhesions in a noncontact manner under biologically relevant conditions.

  4. Role of auxin during intercellular infection of Discaria trinervis by Frankia

    PubMed Central

    Imanishi, Leandro; Perrine-Walker, Francine M.; Ndour, Adama; Vayssières, Alice; Conejero, Genevieve; Lucas, Mikaël; Champion, Antony; Laplaze, Laurent; Wall, Luis; Svistoonoff, Sergio

    2014-01-01

    Nitrogen-fixing nodules induced by Frankia in the actinorhizal plant Discaria trinervis result from a primitive intercellular root invasion pathway that does not involve root hair deformation and infection threads. Here, we analyzed the role of auxin in this intercellular infection pathway at the molecular level and compared it with our previous work in the intracellular infected actinorhizal plant Casuarina glauca. Immunolocalisation experiments showed that auxin accumulated in Frankia-infected cells in both systems. We then characterized the expression of auxin transporters in D. trinervis nodules. No activation of the heterologous CgAUX1 promoter was detected in infected cells in D. trinervis. These results were confirmed with the endogenous D. trinervis gene, DtAUX1. However, DtAUX1 was expressed in the nodule meristem. Consistently, transgenic D. trinervis plants containing the auxin response marker DR5:VENUS showed expression of the reporter gene in the meristem. Immunolocalisation experiments using an antibody against the auxin efflux carrier PIN1, revealed the presence of this transporter in the plasma membrane of infected cells. Finally, we used in silico cellular models to analyse auxin fluxes in D. trinervis nodules. Our results point to the existence of divergent roles of auxin in intercellularly- and intracellularly-infected actinorhizal plants, an ancestral infection pathways leading to root nodule symbioses. PMID:25191330

  5. Extracellular ATP and other nucleotides-ubiquitous triggers of intercellular messenger release.

    PubMed

    Zimmermann, Herbert

    2016-03-01

    Extracellular nucleotides, and ATP in particular, are cellular signal substances involved in the control of numerous (patho)physiological mechanisms. They provoke nucleotide receptor-mediated mechanisms in select target cells. But nucleotides can considerably expand their range of action. They function as primary messengers in intercellular communication by stimulating the release of other extracellular messenger substances. These in turn activate additional cellular mechanisms through their own receptors. While this applies also to other extracellular messengers, its omnipresence in the vertebrate organism is an outstanding feature of nucleotide signaling. Intercellular messenger substances released by nucleotides include neurotransmitters, hormones, growth factors, a considerable variety of other proteins including enzymes, numerous cytokines, lipid mediators, nitric oxide, and reactive oxygen species. Moreover, nucleotides activate or co-activate growth factor receptors. In the case of hormone release, the initially paracrine or autocrine nucleotide-mediated signal spreads through to the entire organism. The examples highlighted in this commentary suggest that acting as ubiquitous triggers of intercellular messenger release is one of the major functional roles of extracellular nucleotides. While initiation of messenger release by nucleotides has been unraveled in many contexts, it may have been overlooked in others. It can be anticipated that additional nucleotide-driven messenger functions will be uncovered with relevance for both understanding physiology and development of therapy.

  6. Inhibition of salivary fluid secretion by occlusion of the intercellular space.

    PubMed

    Nakahari, T; Morimatsu, S; Imai, Y

    1998-08-01

    The effects of HCO3- on fluid secretory rate, cell volume and tissue structure were studied in perfused submandibular salivary glands under hyposmotic conditions (240 mosM). Fluid secretion was elicited by acetylcholine (ACh) in a hyposmotic HCO3(-)-free solution. Upon switching the perfusate from the HCO3(-)-free to the HCO3(-)-containing solution during ACh stimulation, the fluid secretory rate exhibited a small transient increase followed by a sharp decrease. ACh stimulation evoked rapid cell shrinkage in the absence of HCO3-, but upon switching from the HCO3(-)-free to the HCO3(-)-containing solution during ACh stimulation, the acinar cells exhibited increases in volume. In laser confocal microscopic examination of isolated acini, fluorescence of lucifer yellow was detected in the lateral intercellular spaces during ACh stimulation in the absence of HCO3-, but not in the presence of HCO3-. Electron microscopic examination revealed similar findings. We propose that the occlusion of the lateral intercellular space induced by cell swelling may result in an increase in the resistance to fluid flow in the space and consequently a decrease in the fluid secretion rate. Maintenance of an open lateral intercellular space may be an important requirement for fluid secretion. These observations suggest that a paracellular pathway may play a significant role in salivary fluid secretion.

  7. Fluorescent nanodiamond as a probe for the intercellular transport of proteins in vivo.

    PubMed

    Kuo, Yung; Hsu, Tsung-Yuan; Wu, Yi-Chun; Chang, Huan-Cheng

    2013-11-01

    This study investigates the intercellular transport of yolk lipoproteins in Caenorhabditis elegans by using fluorescence lifetime imaging microscopy (FLIM) and fluorescent nanodiamonds (FNDs) as photostable labels and tracers. The yolk lipoproteins in the nematode are similar to human serum low-density lipoproteins (LDLs), serving as an intercellular transporter of fat molecules and cholesterol. To study this fundamentally important process, FNDs were first coated with yolk lipoprotein complexes (YLCs) and then microinjected into the intestinal cells of the living organism. Real-time imaging over a time period of more than 50 min with FLIM revealed the process of YLC-FND secretion from the intestine to the pseudocoelomic space, followed by transporting into oocytes and subsequent accumulation in the multi-cellular embryos derived from the oocytes. Colocalization studies of the rme-2 adult hermaphrodites expressing green fluorescent protein (GFP)-tagged YLCs confirmed that the injected YLC-FNDs were taken up by oocytes through endocytosis mediated by the LDL receptor, RME-2, functioning as an YLC receptor. Our results demonstrate that FND is useful as a biomolecular nanocarrier without significantly altering the functionality of the cargos for intercellular transport, cell-specific targeting, and long-term imaging applications in vivo.

  8. Nanodiamond-Mediated Intercellular Transport of Proteins through Membrane Tunneling Nanotubes.

    PubMed

    Epperla, Chandra Prakash; Mohan, Nitin; Chang, Che-Wei; Chen, Chia-Chun; Chang, Huan-Cheng

    2015-12-02

    Recently discovered tunneling nanotubes (TNTs) are capable of creating intercellular communication pathways through which transport of proteins and other cytoplasmic components occurs. Intercellular transport is related to many diseases and nanotubes are potentially useful as drug-delivery channels for cancer therapy. Here, we apply fluorescent nanodiamond (FND) as a photostable tracker, as well as a protein carrier, to illustrate the transport events in TNTs of human cells. Proteins, including bovine serum albumin and green fluorescent protein, are first coated on 100-nm FNDs by physical adsorption and then single-particle tracking of the bioconjugates in the transient membrane connections is carried out by fluorescence microscopy. Stop-and-go and to-and-fro motions mediated by molecular motors are found for the active transport of protein-loaded FNDs trapped in the endosomal vehicles of human embryonic kidney cells (HEK293T). Quantitative analysis of the heterotypical transport between HEK293T and SH-SY5Y neuroblastoma cells by flow cytometry confirm the formation of open-ended nanotubes between them, despite that their TNTs differ in structural components. Our results demonstrate the promising applications of this novel carbon-based nanomaterial for intercellular delivery of biomolecular cargo down to the single-particle level.

  9. Structure of artificial and natural VE-cadherin-based adherens junctions.

    PubMed

    Taveau, Jean-Christophe; Dubois, Mathilde; Le Bihan, Olivier; Trépout, Sylvain; Almagro, Sébastien; Hewat, Elizabeth; Durmort, Claire; Heyraud, Stéphanie; Gulino-Debrac, Danielle; Lambert, Olivier

    2008-04-01

    In vascular endothelium, adherens junctions between endothelial cells are composed of VE-cadherin (vascular endothelial cadherin), an adhesive receptor that is crucial for the proper assembly of vascular structures and the maintenance of vascular integrity. As a classical cadherin, VE-cadherin links endothelial cells together by homophilic interactions mediated by its extracellular part and associates intracellularly with the actin cytoskeleton via catenins. Although, from structural crystallographic data, a dimeric structure arranged in a trans orientation has emerged as a potential mechanism of cell-cell adhesion, the cadherin organization within adherens junctions remains controversial. Concerning VE-cadherin, its extracellular part possesses the capacity to self-associate in solution as hexamers consisting of three antiparallel cadherin dimers. VE-cadherin-based adherens junctions were reconstituted in vitro by assembly of a VE-cadherin EC (extracellular repeat) 1-EC4 hexamer at the surfaces of liposomes. The artificial adherens junctions revealed by cryoelectron microscopy appear as a two-dimensional self-assembly of hexameric structures. This cadherin organization is reminiscent of that found in native desmosomal junctions. Further structural studies performed on native VE-cadherin junctions would provide a better understanding of the cadherin organization within adherens junctions. Homophilic interactions between cadherins are strengthened intracellularly by connection to the actin cytoskeleton. Recently, we have discovered that annexin 2, an actin-binding protein connects the VE-cadherin-catenin complex to the actin cytoskeleton. This novel link is labile and promotes the endothelial cell switch from a quiescent to an angiogenic state.

  10. Conducting polyaniline nanowire electrode junction

    NASA Astrophysics Data System (ADS)

    Gaikwad, Sumedh; Bodkhe, Gajanan; Deshmukh, Megha; Patil, Harshada; Rushi, Arti; Shirsat, Mahendra D.; Koinkar, Pankaj; Kim, Yun-Hae; Mulchandani, Ashok

    2015-03-01

    In this paper, a synthesis of conducting polyaniline nanowires electrode junction (CPNEJ) has been reported. Conducting polyaniline nanowires electrode junction on Si/SiO2 substrate (having 3 μm gap between two gold microelectrodes) is prepared. Polyaniline nanowires with diameter (ca. 140 nm to 160 nm) were synthesized by one step electrochemical polymerization using galvanostatic (constant current) technique to bridge this gap. The surface morphology of CPNEJ was studied by scanning electron microscope (SEM). The synthesized CPNEJ is an excellent platform for biosensor applications.

  11. Atomically thin p-n junctions with van der Waals heterointerfaces.

    PubMed

    Lee, Chul-Ho; Lee, Gwan-Hyoung; van der Zande, Arend M; Chen, Wenchao; Li, Yilei; Han, Minyong; Cui, Xu; Arefe, Ghidewon; Nuckolls, Colin; Heinz, Tony F; Guo, Jing; Hone, James; Kim, Philip

    2014-09-01

    Semiconductor p-n junctions are essential building blocks for electronic and optoelectronic devices. In conventional p-n junctions, regions depleted of free charge carriers form on either side of the junction, generating built-in potentials associated with uncompensated dopant atoms. Carrier transport across the junction occurs by diffusion and drift processes influenced by the spatial extent of this depletion region. With the advent of atomically thin van der Waals materials and their heterostructures, it is now possible to realize a p-n junction at the ultimate thickness limit. Van der Waals junctions composed of p- and n-type semiconductors--each just one unit cell thick--are predicted to exhibit completely different charge transport characteristics than bulk heterojunctions. Here, we report the characterization of the electronic and optoelectronic properties of atomically thin p-n heterojunctions fabricated using van der Waals assembly of transition-metal dichalcogenides. We observe gate-tunable diode-like current rectification and a photovoltaic response across the p-n interface. We find that the tunnelling-assisted interlayer recombination of the majority carriers is responsible for the tunability of the electronic and optoelectronic processes. Sandwiching an atomic p-n junction between graphene layers enhances the collection of the photoexcited carriers. The atomically scaled van der Waals p-n heterostructures presented here constitute the ultimate functional unit for nanoscale electronic and optoelectronic devices.

  12. Functional chromaffin cell plasticity in response to stress: focus on nicotinic, gap junction, and voltage-gated Ca2+ channels

    PubMed Central

    Guérineau, Nathalie C.; Desarménien, Michel G.; Carabelli, Valentina; Carbone, Emilio

    2012-01-01

    An increase in circulating catecholamines constitutes one of the mechanisms whereby human body responds to stress. In response to chronic stressful situations, the adrenal medullary tissue exhibits crucial morphological and functional changes that are consistent with an improvement of chromaffin cell stimulus-secretion coupling efficiency. Stimulus-secretion coupling encompasses multiple intracellular (chromaffin cell excitability, Ca2+ signaling, exocytosis, endocytosis) and intercellular pathways (splanchnic nerve-mediated synaptic transmission, paracrine and endocrine communication, gap junctional coupling), each of them being potentially subjected to functional remodeling upon stress. This review focuses on three chromaffin cell incontrovertible actors, the cholinergic nicotinic receptors and the voltage-dependent T-type Ca2+ channels that are directly involved in Ca2+-dependent events controlling catecholamine secretion and electrical activity, and the gap junctional communication involved in the modulation of catecholamine secretion. We show here that these three actors react differently to various stressors, sometimes independently, sometimes in concert or in opposition. PMID:22252244

  13. Characterization of Tight Junction Proteins in Cultured Human Urothelial Cells

    PubMed Central

    Rickard, Alice; Dorokhov, Nikolay; Ryerse, Jan; Klumpp, David J.; McHowat, Jane

    2010-01-01

    Tight junctions (TJs) are essential for normal function of epithelia, restricting paracellular diffusion and contributing to the maintainance of cell surface polarity. Superficial cells of the urothelium develop TJs, the basis for the paracellular permeability barrier of the bladder against diffusion of urinary solutes. Focusing on the superficial cell layer of stratified cell cultures of an immortalized human ureteral cell line, TEU-2 cells, we have examined the presence of TJ and TJ-associated proteins. TEU-2 cells were treated with calcium chloride and fetal bovine serum culture conditions used to induce stratification that resembles the normal transitional epithelial phenotype. Cultures were examined for TJ and TJ-associated proteins by confocal immuno-fluorescence microscopy and evaluated for TJ mRNA by reverse transcriptase-polymerase chain reaction (RT- PCR). TEU-2 cultures exhibited immunoreactivity at intercellular margins for claudins 1, 4, 5, 7, 14 and 16 whereas claudins 2, 8 and 12 were intracellular. RT-PCR corroborated the presence of these claudins at the mRNA level. The TJ-associated proteins occludin, JAM-1, and zonula occludens (ZO-1, ZO-2 and ZO-3) were localized at cell margins. We have found that numerous TJs and TJ-associated proteins are expressed in stratified TEU-2 cultures. Further, we propose TEU-2s provide a useful ureteral model for future studies on the involvement of TJs proteins in the normal and pathological physiology of the human urinary system. PMID:18553212

  14. Simple Electronic Analog of a Josephson Junction.

    ERIC Educational Resources Information Center

    Henry, R. W.; And Others

    1981-01-01

    Demonstrates that an electronic Josephson junction analog constructed from three integrated circuits plus an external reference oscillator can exhibit many of the circuit phenomena of a real Josephson junction. Includes computer and other applications of the analog. (Author/SK)

  15. Low dose/low fluence ionizing radiation-induced biological effects: The role of intercellular communication and oxidative metabolism

    NASA Astrophysics Data System (ADS)

    Azzam, Edouard

    Mechanistic investigations have been considered critical to understanding the health risks of exposure to ionizing radiation. To gain greater insight in the biological effects of exposure to low dose/low fluence space radiations with different linear energy transfer (LET) properties, we examined short and long-term biological responses to energetic protons and high charge (Z) and high energy (E) ions (HZE particles) in human cells maintained in culture and in targeted and non-targeted tissues of irradiated rodents. Particular focus of the studies has been on mod-ulation of gene expression, proliferative capacity, induction of DNA damage and perturbations in oxidative metabolism. Exposure to mean doses of 1000 MeV/nucleon iron ions, by which a small to moderate proportion of cells in an exposed population is targeted through the nucleus by an HZE particle, induced stressful effects in the irradiated and non-irradiated cells in the population. Direct intercellular communication via gap-junctions was a primary mediator of the propagation of stressful effects from irradiated to non-irradiated cells. Compromised prolif-erative capacity, elevated level of DNA damage and oxidative stress evaluated by measurements of protein carbonylation, lipid peroxidation and activity of metabolic enzymes persisted in the progeny of irradiated and non-irradiated cells. In contrast, progeny of cells exposed to high or low doses from 150-1000 MeV protons retained the ability to form colonies and harbored similar levels of micronuclei, a surrogate form of DNA damage, as control, which correlated with normal reactive oxygen species (ROS) levels. Importantly, a significant increase in the spontaneous neoplastic transformation frequency was observed in progeny of bystander mouse embryo fibroblasts (MEFs) co-cultured with MEFs irradiated with energetic iron ions but not protons. Of particular significance, stressful effects were detected in non-targeted tissues of rats that received partial

  16. Connexin 43 mediated gap junctional communication enhances breast tumor cell diapedesis in culture

    PubMed Central

    Pollmann, Mary-Ann; Shao, Qing; Laird, Dale W; Sandig, Martin

    2005-01-01

    Introduction Metastasis involves the emigration of tumor cells through the vascular endothelium, a process also known as diapedesis. The molecular mechanisms regulating tumor cell diapedesis are poorly understood, but may involve heterocellular gap junctional intercellular communication (GJIC) between tumor cells and endothelial cells. Method To test this hypothesis we expressed connexin 43 (Cx43) in GJIC-deficient mammary epithelial tumor cells (HBL100) and examined their ability to form gap junctions, establish heterocellular GJIC and migrate through monolayers of human microvascular endothelial cells (HMVEC) grown on matrigel-coated coverslips. Results HBL100 cells expressing Cx43 formed functional heterocellular gap junctions with HMVEC monolayers within 30 minutes. In addition, immunocytochemistry revealed Cx43 localized to contact sites between Cx43 expressing tumor cells and endothelial cells. Quantitative analysis of diapedesis revealed a two-fold increase in diapedesis of Cx43 expressing cells compared to empty vector control cells. The expression of a functionally inactive Cx43 chimeric protein in HBL100 cells failed to increase migration efficiency, suggesting that the observed up-regulation of diapedesis in Cx43 expressing cells required heterocellular GJIC. This finding is further supported by the observation that blocking homocellular and heterocellular GJIC with carbenoxolone in co-cultures also reduced diapedesis of Cx43 expressing HBL100 tumor cells. Conclusion Collectively, our results suggest that heterocellular GJIC between breast tumor cells and endothelial cells may be an important regulatory step during metastasis. PMID:15987459

  17. Influence of the molecular structure of steroids on their ability to interrupt gap junctional communication.

    PubMed

    Hervé, J C; Pluciennik, F; Verrecchia, F; Bastide, B; Delage, B; Joffre, M; Délèze, J

    1996-02-01

    17 beta-estradiol propionate was found to reduce the gap junctional communication in a concentration range similar to that of testosterone propionate, in primary cultures of rat Sertoli cells and cardiac myocytes. Uncoupling was reversible on washing out and occurred without concomitant rise in the intracellular calcium concentration. Esterification was prerequisite for the activity of extracellularly applied steroid compounds (for example, testosterone was ineffective even at external concentrations up to 100 microM, whereas its intracellular application at 1 microM totally interrupted intercellular communication), but their uncoupling efficiency did not depend on the nature of the ester chain nor on its position on the steroid nucleus. The derivatives of two other androgen hormones (derivatives of the androstane nucleus) were also efficient as junctional uncouplers. Among five steroid molecules belonging to the pregnane family, only one (pregnanediol diacetate) interrupted the junctional communication. Neither cholic acid nor cholesteryl acetate or ouabain showed this effect. Altogether, no correlation with the presence or position of double bonds nor with the trans- or cis-fusion of the A and B rings could be recognized. These results suggest that this reversible, nondeleterious uncoupling effect of steroids is independent of the shape of the molecules and is more probably related to their size and liposolubility, that condition their insertion into the lipid bilayer. Their incorporation into the membrane could disturb the activity of the membrane proteins by a physical mechanism.

  18. Reversible interruption of gap junctional communication by testosterone propionate in cultured Sertoli cells and cardiac myocytes.

    PubMed

    Pluciennik, F; Verrecchia, F; Bastide, B; Hervé, J C; Joffre, M; Délèze, J

    1996-02-01

    A direct cell-to-cell exchange of ions and molecules occurs through specialized membrane channels built by the interaction of two half channels, termed connexons, contributed by each of the two adjacent cells. The electrical and diffusional couplings have been investigated by monitoring respectively the cell-to-cell conductance and the fluorescence recovery after photobleaching, in Sertoli and cardiac cells of young rat. In both cell types, a rapid impairment of the intercellular coupling has been observed in the presence of testosterone propionate. This interruption of the cell-to-cell communication through gap junction channels was dose-dependent, observed in the concentration range 1 to 25 microM and was progressively reversed after withdrawing the testosterone ester. Pretreatment with cyproterone acetate, an antiandrogen which blocks the nuclear testosterone receptor by binding, did not prevent the uncoupling action of the androgen ester. This observation, together with the rapid time course of the uncoupling and recoupling, and the rather high effective concentration (micromolar) of the steroid compound, suggests a nongenomic mechanism of action. The uncoupling concentrations were very similar to those of other steroid compounds known to interrupt gap junctional communication. The uncoupling could result from a direct interaction of the steroid with the proteolipidic structure of the membrane, that might alter the conformation of the gap junction channels and their functional state.

  19. GAP: A computer program for gene assembly

    SciTech Connect

    Eisnstein, J.R.; Uberbacher, E.C.; Guan, X.; Mural, R.J.; Mann, R.C.

    1991-09-01

    A computer program, GAP (Gene Assembly Program), has been written to assemble and score hypothetical genes, given a DNA sequence containing the gene, and the outputs of several other programs which analyze the sequence. These programs include the codign-recognition and splice-junction-recognition modules developed in this laboratory. GAP is a prototype of a planned system in which it will be integrated with an expert system and rule base. Initial tests of GAP have been carried out with four sequences, the exons of which have been determined by biochemcial methods. The highest-scoring hypothetical genes for each of the four sequences had percent correct splice junctions ranging from 50 to 100% (average 81%) and percent correct bases ranging from 92 to 100% (average 96%). 9 refs., 1 tab.

  20. The Yolla Bolly junction revisited

    SciTech Connect

    Blake, M.C.; Jayko, A.S. ); Jones, D.L. . Dept. of Geology and Geophysics); Engebretson, D.C. . Dept. of Geology)

    1993-04-01

    West of Red Bluff, California, rocks of the northern Coast Ranges, Klamath-Sierra Nevada, and Great Valley provinces come together at what has been called the Yolla Bolly junction. Mapping of the Red Bluff and Willows 1:100,000 quadrangles has greatly clarified the enigmatic features of this complex area. Terranes of the Klamath Mountains and their Cretaceous sedimentary cover have been thrust northwestward over the Elder Creek terrane and Franciscan rocks, north of the left-lateral Cold Fork fault zone. The Condrey Mountain window (Franciscan Pickett Peak terrane) provides a measure of the magnitude of this thrusting (ca 90 km). South of the Cold Fork fault zone, the Franciscan and Elder Creek terranes were driven southeastward as tectonic wedges onto Sierran-Klamath basement. Timing of this scissor-tectonics is not constrained near the junction, but further north in southwest Oregon, Lower Eocene strata were deformed by overthrusting of the Klamath block whereas Upper Eocene strata overlap the thrust, indicating that thrusting occurred between about 52 and 60 Ma. Plate reconstructions for this time interval indicate the close proximity of the Kula-Farallon-North America triple junction and that old (ca 100 m.y.) Farallon lithosphere was being subducted north of the junction whereas to the south, very young (ca 10 m.y.) Kula plate was presumably obducted onto North America.

  1. Hinge assembly

    DOEpatents

    Vandergriff, D.H.

    1999-08-31

    A hinge assembly is disclosed having a first leaf, a second leaf and linking member. The first leaf has a contact surface. The second leaf has a first contact surface and a second contact surface. The linking member pivotally connects to the first leaf and to the second leaf. The hinge assembly is capable of moving from a closed position to an open position. In the closed position, the contact surface of the first leaf merges with the first contact surface of the second leaf. In the open position, the contact surface of the first leaf merges with the second contact surface of the second leaf. The hinge assembly can include a seal on the contact surface of the first leaf. 8 figs.

  2. Hinge assembly

    DOEpatents

    Vandergriff, David Houston

    1999-01-01

    A hinge assembly having a first leaf, a second leaf and linking member. The first leaf has a contact surface. The second leaf has a first contact surface and a second contact surface. The linking member pivotally connects to the first leaf and to the second leaf. The hinge assembly is capable of moving from a closed position to an open position. In the closed position, the contact surface of the first leaf merges with the first contact surface of the second leaf. In the open position, the contact surface of the first leaf merges with the second contact surface of the second leaf. The hinge assembly can include a seal on the contact surface of the first leaf.

  3. Cooperativity-based modeling of heterotypic DNA nanostructure assembly

    PubMed Central

    Shapiro, Anastasia; Hozeh, Avital; Girshevitz, Olga; Abu-Horowitz, Almogit; Bachelet, Ido

    2015-01-01

    DNA origami is a robust method for the fabrication of nanoscale 2D and 3D objects with complex features and geometries. The process of DNA origami folding has been recently studied, however quantitative understanding of it is still elusive. Here, we describe a systematic quantification of the assembly process of DNA nanostructures, focusing on the heterotypic DNA junction—in which arms are unequal—as their basic building block. Using bulk fluorescence studies we tracked this process and identified multiple levels of cooperativity from the arms in a single junction to neighboring junctions in a large DNA origami object, demonstrating that cooperativity is a central underlying mechanism in the process of DNA nanostructure assembly. We show that the assembly of junctions in which the arms are consecutively ordered is more efficient than junctions with randomly-ordered components, with the latter showing assembly through several alternative trajectories as a potential mechanism explaining the lower efficiency. This highlights consecutiveness as a new design consideration that could be implemented in DNA nanotechnology CAD tools to produce more efficient and high-yield designs. Altogether, our experimental findings allowed us to devise a quantitative, cooperativity-based heuristic model for the assembly of DNA nanostructures, which is highly consistent with experimental observations. PMID:26071955

  4. [Introduction to the structure and functions of junction communications or gap junctions].

    PubMed

    Rousset, B

    1996-01-01

    Cell-to-cell communication through gap junctions (GJ) represents a direct route of exchange of informations between neighboring cells within tissues and organs. GJ are formed from the assembly of a large number of channels that differ from the other known channels because they connect the cytoplasm of adjacent cells. The GJ channel is built from two parts: the connexons. A connexon inserted into the plasma membrane of a cell interacts with another connexon belonging to an adjacent cell. Connexons are composed of proteins with four transmembrane domains that are named connexins (Cx). Six Cx form a connexon. Cx belong to a protein family with 13 known members at present. Each Cx is defined by its molecular mass in kDa (ex: Cx32, Cx43...). A given cell type expresses one or several Cx. The cell to cell transfer of molecules through GJ channels exhibit a size selectivity; only molecules with a molecular mass lower than 1000 Da such as ions and second messengers freely pass through GJ. Depending on the Cx they are made of, GJ seem to differ somewhat in their permeability properties. Cell-to-cell communication via GJ is a regulated process. GJ channels can be either open or closed. GJ mediated cell-to-cell communication or junctional coupling can be detected and quantified by visualization of the cell to cell transfer of a fluorescent probe (such as Lucifer Yellow...) previously introduced in a single cell by microinjection. The presence of GJ channels can also be identified by recording the passage of an electric current between contiguous cells. GJ are involved in numerous fundamental biological processes from the embryonic development to the homeostasis in adult tissues and organs. GJ coordinate cell activities and sometimes synchronize cell behaviour. This is the case for the propagation of the excitation wave in the cardiac muscle and smooth muscle. GJ mediate metabolic cooperation between cells; they represent a way of supply of nutrients for tissues that are

  5. 27 CFR 9.164 - River Junction.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 27 Alcohol, Tobacco Products and Firearms 1 2014-04-01 2014-04-01 false River Junction. 9.164... River Junction. (a) Name. The name of the viticultural area described in this section is “River Junction.” (b) Approved maps. The appropriate maps for determining the boundaries of the River...

  6. 27 CFR 9.164 - River Junction.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 27 Alcohol, Tobacco Products and Firearms 1 2011-04-01 2011-04-01 false River Junction. 9.164... River Junction. (a) Name. The name of the viticultural area described in this section is “River Junction.” (b) Approved maps. The appropriate maps for determining the boundaries of the River...

  7. Sabot assembly

    DOEpatents

    Bzorgi, Fariborz

    2016-11-08

    A sabot assembly includes a projectile and a housing dimensioned and configured for receiving the projectile. An air pressure cavity having a cavity diameter is disposed between a front end and a rear end of the housing. Air intake nozzles are in fluid communication with the air pressure cavity and each has a nozzle diameter less than the cavity diameter. In operation, air flows through the plurality of air intake nozzles and into the air pressure cavity upon firing of the projectile from a gun barrel to pressurize the air pressure cavity for assisting in separation of the housing from the projectile upon the sabot assembly exiting the gun barrel.

  8. Latch assembly

    DOEpatents

    Frederickson, J.R.; Harper, W.H.; Perez, R.

    1984-08-17

    A latch assembly for releasably securing an article in the form of a canister within a container housing. The assembly includes a cam pivotally mounted on the housing wall and biased into the housing interior. The cam is urged into a disabled position by the canister as it enters the housing and a latch release plate maintains the cam disabled when the canister is properly seated in the housing. Upon displacement of the release plate, the cam snaps into latching engagement against the canister for securing the same within the housing. 2 figs.

  9. Nitrogenase assembly

    PubMed Central

    Hu, Yilin; Ribbe, Markus W.

    2013-01-01

    Nitrogenase contains two unique metalloclusters: the P-cluster and the M-cluster. The assembly processes of P- and M-clusters are arguably the most complicated processes in bioinorganic chemistry. There is considerable interest in decoding the biosynthetic mechanisms of the P- and M-clusters, because these clusters are not only biologically important, but also chemically unprecedented. Understanding the assembly mechanisms of these unique metalloclusters is crucial for understanding the structure-function relationship of nitrogenase. Here, we review the recent advances in this research area, with an emphasis on our work that provide important insights into the biosynthetic pathways of these high-nuclearity metal centers. PMID:23232096

  10. Latch assembly

    DOEpatents

    Frederickson, James R.; Harper, William H.; Perez, Raymond

    1986-01-01

    A latch assembly for releasably securing an article in the form of a canister within a container housing. The assembly includes a cam pivotally mounted on the housing wall and biased into the housing interior. The cam is urged into a disabled position by the canister as it enters the housing and a latch release plate maintains the cam disabled when the canister is properly seated in the housing. Upon displacement of the release plate, the cam snaps into latching engagement against the canister for securing the same within the housing.

  11. Adipocytes in both brown and white adipose tissue of adult mice are functionally connected via gap junctions: implications for Chagas disease

    PubMed Central

    Burke, Shoshana; Nagajyothi, Fnu; Thi, Mia M.; Hanani, Menachem; Scherer, Philipp E.; Tanowitz, Herbert B.; Spray, David C.

    2015-01-01

    Adipose tissue serves as a host reservoir for the protozoan Trypanosoma cruzi, the causative organism in Chagas disease. Gap junctions interconnect cells of most tissues, serving to synchronize cell activities including secretion in glandular tissue, and we have previously demonstrated that gap junctions are altered in various tissues and cells infected with T. cruzi. Herein, we examined the gap junction protein connexin 43 (Cx43) expression in infected adipose tissues. Adipose tissue is the largest endocrine organ of the body and is also involved in other physiological functions. In mammals, it is primarily composed of white adipocytes. Although gap junctions are a prominent feature of brown adipocytes, they have not been explored extensively in white adipocytes, especially in the setting of infection. Thus, we examined functional coupling in both white and brown adipocytes in mice. Injection of electrical current or the dye Lucifer Yellow into adipocytes within fat tissue spread to adjacent cells, which was reduced by treatment with agents known to block gap junctions. Moreover, Cx43 was detected in both brown and white fat tissue. At thirty and ninety days post-infection, Cx43 was downregulated in brown adipocytes and upregulated in white adipocytes. Gap junction-mediated intercellular communication likely contributes to hormone secretion and other functions in white adipose tissue and to nonshivering thermogenesis in brown fat, and modulation of the coupling by T. cruzi infection is expected to impact these functions. PMID:25150689

  12. Adipocytes in both brown and white adipose tissue of adult mice are functionally connected via gap junctions: implications for Chagas disease.

    PubMed

    Burke, Shoshana; Nagajyothi, Fnu; Thi, Mia M; Hanani, Menachem; Scherer, Philipp E; Tanowitz, Herbert B; Spray, David C

    2014-11-01

    Adipose tissue serves as a host reservoir for the protozoan Trypanosoma cruzi, the causative organism in Chagas disease. Gap junctions interconnect cells of most tissues, serving to synchronize cell activities including secretion in glandular tissue, and we have previously demonstrated that gap junctions are altered in various tissues and cells infected with T. cruzi. Herein, we examined the gap junction protein connexin 43 (Cx43) expression in infected adipose tissues. Adipose tissue is the largest endocrine organ of the body and is also involved in other physiological functions. In mammals, it is primarily composed of white adipocytes. Although gap junctions are a prominent feature of brown adipocytes, they have not been explored extensively in white adipocytes, especially in the setting of infection. Thus, we examined functional coupling in both white and brown adipocytes in mice. Injection of electrical current or the dye Lucifer Yellow into adipocytes within fat tissue spread to adjacent cells, which was reduced by treatment with agents known to block gap junctions. Moreover, Cx43 was detected in both brown and white fat tissue. At thirty and ninety days post-infection, Cx43 was downregulated in brown adipocytes and upregulated in white adipocytes. Gap junction-mediated intercellular communication likely contributes to hormone secretion and other functions in white adipose tissue and to nonshivering thermogenesis in brown fat, and modulation of the coupling by T. cruzi infection is expected to impact these functions.

  13. The uncoupling effect of diacylglycerol on gap junctional communication of mammalian heart cells is independent of protein kinase C.

    PubMed

    Bastide, B; Hervé, J C; Délèze, J

    1994-10-01

    Possible regulatory effects on cell-to-cell communication of a synthetic diacylglycerol, an activator of protein kinase C (PKC), were examined in pairs of synchronously beating ventricular myocytes of neonatal rats in primary culture. Junctional communication was estimated by measuring either the rate constant of dye diffusion, with the fluorescence recovery after photobleaching technique, or the cell-to-cell electrical conductance with a double whole-cell voltage clamp. The addition of a freshly prepared emulsion of 1-oleoyl-2-acetyl-sn-glycerol (OAG, 100 micrograms/ml), either in the bath or in the solution filling the patch pipet, was seen to interrupt intercellular communication within approximately 8 to 10 min. This effect is neither mimicked by stimulation of PKC by a phorbol ester, nor prevented by PKC inhibitors, making it unlikely that, in these cells, PKC activation could induce intercellular uncoupling. During OAG exposures, the intracellular calcium concentration was very modestly increased (by a factor 1.5 to 2), which does not suffice to account for uncoupling. OAG might trigger interruption of cell-to-cell communication by a mechanism analogous to that of other lipophilic molecules (such as aliphatic alcohols or long chain unsaturated fatty acids) which interfere with gap junctions.

  14. Furnace assembly

    DOEpatents

    Panayotou, Nicholas F.; Green, Donald R.; Price, Larry S.

    1985-01-01

    A method of and apparatus for heating test specimens to desired elevated temperatures for irradiation by a high energy neutron source. A furnace assembly is provided for heating two separate groups of specimens to substantially different, elevated, isothermal temperatures in a high vacuum environment while positioning the two specimen groups symmetrically at equivalent neutron irradiating positions.

  15. Furnace assembly

    DOEpatents

    Panayotou, N.F.; Green, D.R.; Price, L.S.

    A method of and apparatus for heating test specimens to desired elevated temperatures for irradiation by a high energy neutron source. A furnace assembly is provided for heating two separate groups of specimens to substantially different, elevated, isothermal temperatures in a high vacuum environment while positioning the two specimen groups symmetrically at equivalent neutron irradiating positions.

  16. Modulation of N-cadherin junctions and their role as epicenters of differentiation-specific actin regulation in the developing lens.

    PubMed

    Leonard, Michelle; Zhang, Liping; Zhai, Ni; Cader, Ahmad; Chan, Yim; Nowak, Roberta B; Fowler, Velia M; Menko, A Sue

    2011-01-15

    Extensive elongation of lens fiber cells is a central feature of lens morphogenesis. Our study investigates the role of N-cadherin junctions in this process in vivo. We investigate both the molecular players involved in N-cadherin junctional maturation and the subsequent function of these junctions as epicenters for the assembly of an actin cytoskeleton that drives morphogenesis. We present the first evidence of nascent cadherin junctions in vivo, and show that they are a prominent feature along lateral interfaces of undifferentiated lens epithelial cells. Maturation of these N-cadherin junctions, required for lens cell differentiation, preceded organization of a cortical actin cytoskeleton along the cells' lateral borders, but was linked to recruitment of α-catenin and dephosphorylation of N-cadherin-linked β-catenin. Biochemical analysis revealed differentiation-specific recruitment of actin regulators cortactin and Arp3 to maturing N-cadherin junctions of differentiating cells, linking N-cadherin junctional maturation with actin cytoskeletal assembly during fiber cell elongation. Blocking formation of mature N-cadherin junctions led to reduced association of α-catenin with N-cadherin, prevented organization of actin along lateral borders of differentiating lens fiber cells and blocked their elongation. These studies provide a molecular link between N-cadherin junctions and the organization of an actin cytoskeleton that governs lens fiber cell morphogenesis in vivo.

  17. Tight junction proteins: from barrier to tumorigenesis.

    PubMed

    Runkle, E Aaron; Mu, David

    2013-08-28

    The tight junction is a multi-protein complex and is the apical most junctional complex in certain epithelial and endothelial cells. A great deal of attention has been devoted to the understanding of these proteins in contributing to the barrier function - that is, regulating the paracellular flux or permeability between adjacent cells. However, tight junction proteins are now recognized as having functions beyond the barrier. The focus of this review is to discuss the barrier function of the tight junction and to summarize the literature with a focus on the role of tight junction proteins in proliferation, transformation, and metastasis.

  18. Tight Junction Proteins: From Barrier to Tumorigenesis

    PubMed Central

    Runkle, E. Aaron; Mu, David

    2013-01-01

    The tight junction is a multi-protein complex and is the apical most junctional complex in certain epithelial and endothelial cells. A great deal of attention has been devoted to the understanding of these proteins in contributing to the barrier function - that is, regulating the paracellular flux or permeability between adjacent cells. However, tight junction proteins are now recognized as having functions beyond the barrier. The focus of this review is to discuss the barrier function of the tight junction and to summarize the literature with a focus on the role of tight junction proteins in proliferation, transformation, and metastasis. PMID:23743355

  19. Tight Junction Proteins in Human Schwann Cell Autotypic Junctions

    PubMed Central

    Alanne, Maria H.; Pummi, Kati; Heape, Anthony M.; Grènman, Reidar; Peltonen, Juha; Peltonen, Sirkku

    2009-01-01

    Tight junctions (TJs) form physical barriers in various tissues and regulate paracellular transport of ions, water, and molecules. Myelinating Schwann cells form highly organized structures, including compact myelin, nodes of Ranvier, paranodal regions, Schmidt-Lanterman incisures, periaxonal cytoplasmic collars, and mesaxons. Autotypic TJs are formed in non-compacted myelin compartments between adjacent membrane lamellae of the same Schwann cell. Using indirect immunofluorescence and RT-PCR, we analyzed the expression of adherens junction (E-cadherin) and TJ [claudins, zonula occludens (ZO)-1, occludin] components in human peripheral nerve endoneurium, showing clear differences with published rodent profiles. Adult nerve paranodal regions contained E-cadherin, claudin-1, claudin-2, and ZO-1. Schmidt-Lanterman incisures contained E-cadherin, claudin-1, claudin-2, claudin-3, claudin-5, ZO-1, and occludin. Mesaxons contained E-cadherin, claudin-1, claudin-2, claudin-3, ZO-1, and occludin. None of the proteins studied were associated with nodal inter-Schwann cell junctions. Fetal nerve expression of claudin-1, claudin-3, ZO-1, and occludin was predominantly punctate, with a mesaxonal labeling pattern, but paranodal (ZO-1, claudin-3) and Schmidt-Lanterman incisure (claudins-1 and -3) expression profiles typical of compact myelin were visible by gestational week 37. The clear differences observed between human and published rodent nerve profiles emphasize the importance of human studies when translating the results of animal models to human diseases. (J Histochem Cytochem 57:523–529, 2009) PMID:19153196

  20. Glucose Transporters are Abundant in Cells with "Occluding" Junctions at the Blood-Eye Barriers

    NASA Astrophysics Data System (ADS)

    Harik, Sami I.; Kalaria, Rajesh N.; Whitney, Paul M.; Andersson, Lars; Lundahl, Per; Ledbetter, Steven R.; Perry, George

    1990-06-01

    We studied the distribution of the "erythroid/brain" glucose transporter protein in the human and rat eye by immunocytochemistry with monoclonal and polyclonal antibodies to the C terminus of the human erythrocyte glucose transporter. We found intense immunocytochemical staining in the endothelium of microvessels of the retina, optic nerve, and iris but not in microvessels of the choroid, ciliary body, sclera, and other retro-orbital tissues. In addition, we found marked immunocytochemical staining of retinal pigment epithelium, ciliary body epithelium, and posterior epithelium of the iris. The common feature of all those endothelial and epithelial cells that stained intensely for the glucose transporter is the presence of "occluding" intercellular junctions, which constitute the anatomical bases of the blood-eye barriers. We propose that a high density of the glucose transporter is a biochemical concomitant of epithelial and endothelial cells with barrier characteristics, at least in tissues that have a high metabolic requirement for glucose.

  1. Connexin Type and Fluorescent Protein Fusion Tag Determine Structural Stability of Gap Junction Plaques*

    PubMed Central

    Stout, Randy F.; Snapp, Erik Lee; Spray, David C.

    2015-01-01

    Gap junctions (GJs) are made up of plaques of laterally clustered intercellular channels and the membranes in which the channels are embedded. Arrangement of channels within a plaque determines subcellular distribution of connexin binding partners and sites of intercellular signaling. Here, we report the discovery that some connexin types form plaque structures with strikingly different degrees of fluidity in the arrangement of the GJ channel subcomponents of the GJ plaque. We uncovered this property of GJs by applying fluorescence recovery after photobleaching to GJs formed from connexins fused with fluorescent protein tags. We found that connexin 26 (Cx26) and Cx30 GJs readily diffuse within the plaque structures, whereas Cx43 GJs remain persistently immobile for more than 2 min after bleaching. The cytoplasmic C terminus of Cx43 was required for stability of Cx43 plaque arrangement. We provide evidence that these qualitative differences in GJ arrangement stability reflect endogenous characteristics, with the caveat that the sizes of the GJs examined were necessarily large for these measurements. We also uncovered an unrecognized effect of non-monomerized fluorescent protein on the dynamically arranged GJs and the organization of plaques composed of multiple connexin types. Together, these findings redefine our understanding of the GJ plaque structure and should be considered in future studies using fluorescent protein tags to probe dynamics of highly ordered protein complexes. PMID:26265468

  2. LINGO-1, a transmembrane signaling protein, inhibits oligodendrocyte differentiation and myelination through intercellular self-interactions.

    PubMed

    Jepson, Scott; Vought, Bryan; Gross, Christian H; Gan, Lu; Austen, Douglas; Frantz, J Daniel; Zwahlen, Jacque; Lowe, Derek; Markland, William; Krauss, Raul

    2012-06-22

    Overcoming remyelination failure is a major goal of new therapies for demyelinating diseases like multiple sclerosis. LINGO-1, a key negative regulator of myelination, is a transmembrane signaling protein expressed in both neurons and oligodendrocytes. In neurons, LINGO-1 is an integral component of the Nogo receptor complex, which inhibits axonal growth via RhoA. Because the only ligand-binding subunit of this complex, the Nogo receptor, is absent in oligodendrocytes, the extracellular signals that inhibit myelination through a LINGO-1-mediated mechanism are unknown. Here we show that LINGO-1 inhibits oligodendrocyte terminal differentiation through intercellular interactions and is capable of a self-association in trans. Consistent with previous reports, overexpression of full-length LINGO-1 inhibited differentiation of oligodendrocyte precursor cells (OPCs). Unexpectedly, treatment with a soluble recombinant LINGO-1 ectodomain also had an inhibitory effect on OPCs and decreased myelinated axonal segments in cocultures with neurons from dorsal root ganglia. We demonstrated LINGO-1-mediated inhibition of OPCs through intercellular signaling by using a surface-bound LINGO-1 construct expressed ectopically in astrocytes. Further investigation showed that the soluble LINGO-1 ectodomain can interact with itself in trans by binding to CHO cells expressing full-length LINGO-1. Finally, we observed that soluble LINGO-1 could activate RhoA in OPCs. We propose that LINGO-1 acts as both a ligand and a receptor and that the mechanism by which it negatively regulates OPC differentiation and myelination is mediated by a homophilic intercellular interaction. Disruption of this protein-protein interaction could lead to a decrease of LINGO-1 inhibition and an increase in myelination.

  3. Follicle-Stimulating Hormone Increases Gap Junctional Communication Between Somatic and Germ-Line Follicular Compartments During Murine Oogenesis.

    PubMed

    El-Hayek, Stephany; Clarke, Hugh J

    2015-08-01

    Germ cells develop in intimate contact and communication with somatic cells of the gonad. In female mammals, oocyte development depends crucially on gap junctions that couple it to the surrounding somatic granulosa cells of the follicle, yet the mechanisms that regulate this essential intercellular communication remain incompletely understood. Follicle-stimulating hormone (FSH) drives the terminal stage of follicular development. We found that FSH increases the steady-state levels of mRNAs encoding the principal connexins that constitute gap junctions and cadherins that mediate cell attachment. This increase occurs both in granulosa cells, which express the FSH-receptor, and in oocytes, which do not. FSH also increased the number of transzonal projections that provide the sites of granulosa cell-oocyte contact. Consistent with increased connexin expression, FSH increased gap junctional communication between granulosa cells and between the oocyte and granulosa cells, and it accelerated oocyte development. These results demonstrate that FSH regulates communication between the female germ cell and its somatic microenvironment. We propose that FSH-regulated gap junctional communication ensures that differentiation processes occurring in distinct cellular compartments within the follicle are precisely coordinated to ensure production of a fertilizable egg.

  4. Effect of Mefloquine, a Gap Junction Blocker, on Circadian Period2 Gene Oscillation in the Mouse Suprachiasmatic Nucleus Ex Vivo

    PubMed Central

    Koo, Jinmi; Choe, Han Kyoung; Kim, Hee-Dae; Chun, Sung Kook; Son, Gi Hoon

    2015-01-01

    Background In mammals, the master circadian pacemaker is localized in an area of the ventral hypothalamus known as the suprachiasmatic nucleus (SCN). Previous studies have shown that pacemaker neurons in the SCN are highly coupled to one another, and this coupling is crucial for intrinsic self-sustainability of the SCN central clock, which is distinguished from peripheral oscillators. One plausible mechanism underlying the intercellular communication may involve direct electrical connections mediated by gap junctions. Methods We examined the effect of mefloquine, a neuronal gap junction blocker, on circadian Period 2 (Per2) gene oscillation in SCN slice cultures prepared from Per2::luciferase (PER2::LUC) knock-in mice using a real-time bioluminescence measurement system. Results Administration of mefloquine causes instability in the pulse period and a slight reduction of amplitude in cyclic PER2::LUC expression. Blockade of gap junctions uncouples PER2::LUC-expressing cells, in terms of phase transition, which weakens synchrony among individual cellular rhythms. Conclusion These findings suggest that neuronal gap junctions play an important role in synchronizing the central pacemaker neurons and contribute to the distinct self-sustainability of the SCN master clock. PMID:25491783

  5. Comparative analysis of theophylline and cholera toxin in rat colon reveals an induction of sealing tight junction proteins.

    PubMed

    Markov, Alexander G; Falchuk, Evgeny L; Kruglova, Natalia M; Rybalchenko, Oksana V; Fromm, Michael; Amasheh, Salah

    2014-11-01

    Claudin tight junction proteins have been identified to primarily determine intestinal epithelial barrier properties. While functional contribution of single claudins has been characterized in detail, information on the interplay with secretory mechanisms in native intestinal epithelium is scarce. Therefore, effects of cholera toxin and theophylline on rat colon were analyzed, including detection of sealing claudins. Tissue specimens were stripped off submucosal tissue layers and mounted in Ussing chambers, and short-circuit current (ISC) and transepithelial resistance (TER) were recorded. In parallel, expression and localization of claudins was analyzed and histological studies were performed employing hematoxylin-eosin staining and light and electron microscopy. Theophylline induced a strong increase of ISC in colon tissue specimens. In parallel, a decrease of TER was observed. In contrast, cholera toxin did not induce a significant increase of ISC, whereas an increase of TER was detected after 120 min. Western blots of membrane fractions revealed an increase of claudin-3 and -4 after incubation with cholera toxin, and theophylline induced an increase of claudin-4. In accordance, confocal laser-scanning microscopy exhibited increased signals of claudin-3 and -4 after incubation with cholera toxin, and increased signals of claudin-4 after incubation with theophylline, within tight junction complexes. Morphological analyses revealed no general changes of tight junction complexes, but intercellular spaces were markedly widened after incubation with cholera toxin and theophylline. We conclude that cholera toxin and theophylline have different effects on sealing tight junction proteins in native colon preparations, which may synergistically contribute to transport functions, in vitro.

  6. [Ontogenetic diversity of colonies and intercellular cytoplasmic bridges in the algae of the genuis Volvox].

    PubMed

    Desnitskiĭ, A G

    2014-01-01

    In all representatives of the genus Volvox, cells of cleaving embryos are connected by cytoplasmic bridges, which play an important role in the process of young colony inversion. However, during subsequent development, the intercellular bridges are retained not in all species of Volvox; the occurrence of the bridges in an adult colony correlates withthe small size of mature gonidia (asexual reproductive cells) and with the presence of cell growth in the intervals between divisions. This complex of ontogenetic features is derived and arises independently in three evolutionary lineages of colonial volvocine algae. A putative role of the syncytial state of adult colonies for the evolution of developmental cycles in Volvox is discussed.

  7. Targeting sites of inflammation: intercellular adhesion molecule-1 as a target for novel inflammatory therapies

    PubMed Central

    Hua, Susan

    2013-01-01

    Targeted drug delivery to sites of inflammation will provide effective, precise, and safe therapeutic interventions for treatment of diverse disease conditions, by limiting toxic side effects and/or increasing drug action. Disease-site targeting is believed to play a major role in the enhanced efficacy observed for a variety of drugs when formulated inside lipid vesicles. This article will focus on the factors and mechanisms involved in drug targeting to sites of inflammation and the importance of cell adhesion molecules, in particular intercellular adhesion molecule-1, in this process. PMID:24109453

  8. Seebeck effect in molecular junctions.

    PubMed

    Zimbovskaya, Natalya A

    2016-05-11

    Advances in the fabrication and characterization of nanoscale systems presently allow for a better understanding of their thermoelectric properties. As is known, the building blocks of thermoelectricity are the Peltier and Seebeck effects. In the present work we review results of theoretical studies of the Seebeck effect in single-molecule junctions and similar systems. The behavior of thermovoltage and thermopower in these systems is controlled by several factors including the geometry of molecular bridges, the characteristics of contacts between the bridge and the electrodes, the strength of the Coulomb interactions between electrons on the bridge, and of electron-phonon interactions. We describe the impact of these factors on the thermopower. Also, we discuss a nonlinear Seebeck effect in molecular junctions.

  9. Seebeck effect in molecular junctions

    NASA Astrophysics Data System (ADS)

    Zimbovskaya, Natalya A.

    2016-05-01

    Advances in the fabrication and characterization of nanoscale systems presently allow for a better understanding of their thermoelectric properties. As is known, the building blocks of thermoelectricity are the Peltier and Seebeck effects. In the present work we review results of theoretical studies of the Seebeck effect in single-molecule junctions and similar systems. The behavior of thermovoltage and thermopower in these systems is controlled by several factors including the geometry of molecular bridges, the characteristics of contacts between the bridge and the electrodes, the strength of the Coulomb interactions between electrons on the bridge, and of electron-phonon interactions. We describe the impact of these factors on the thermopower. Also, we discuss a nonlinear Seebeck effect in molecular junctions.

  10. Thermocouple, multiple junction reference oven

    NASA Technical Reports Server (NTRS)

    Leblanc, L. P. (Inventor)

    1981-01-01

    An improved oven for maintaining the junctions of a plurality of reference thermocouples at a common and constant temperature is described. The oven is characterized by a cylindrical body defining a heat sink with axially extended-cylindrical cavity a singularized heating element which comprises a unitary cylindrical heating element consisting of a resistance heating coil wound about the surface of metallic spool with an axial bore defined and seated in the cavity. Other features of the oven include an annular array of radially extended bores defined in the cylindrical body and a plurality of reference thermocouple junctions seated in the bores in uniformly spaced relation with the heating element, and a temperature sensing device seated in the axial bore for detecting temperature changes as they occur in the spool and circuit to apply a voltage across the coil in response to detected drops in temperatures of the spool.

  11. Intracellular cytoskeleton and junction proteins of endothelial cells in the porcine iris microvasculature.

    PubMed

    Yang, Hongfang; Yu, Paula K; Cringle, Stephen J; Sun, Xinghuai; Yu, Dao-Yi

    2015-11-01

    Recently we reported studies of the iris microvasculature and its endothelial cells using intra-luminal micro-perfusion, fixation, and silver staining, suggesting that the iris vascular endothelium may be crucial for maintaining homeostasis in the ocular anterior segment. Here we present information regarding the intracellular structure and cell junctions of the iris endothelium. Thirty-seven porcine eyes were used for this study. The temporal long posterior ciliary artery was cannulated to assess the iris microvascular network and its endothelium using intra-luminal micro-perfusion, fixation, and staining with phalloidin for intracellular cytoskeleton f-actin, and with antibodies against claudin-5 and VE-cadherin for junction proteins. Nuclei were counterstained with Hoechst. The iris was flat-mounted for confocal imaging. The iris microvasculature was studied for its distribution, branch orders and endothelial morphometrics with endothelial cell length measured for each vessel order. Our results showed that morphometrics of the iris microvasculature was comparable with our previous silver staining. Abundant stress fibres and peripheral border staining were seen within the endothelial cells in larger arteries. An obvious decrease in cytoplasmic stress fibres was evident further downstream in the smaller arterioles, and they tended to be absent from capillaries and veins. Endothelial intercellular junctions throughout the iris vasculature were VE-cadherin and claudin-5 immuno-positive, indicating the presence of both adherent junctions and tight junctions between vascular endothelial cells throughout the iris microvasculature. Unevenness of claudin-5 staining was noted along the endothelial cell borders in almost every order of vessels, especially in veins and small arterioles. Our results suggest that significant heterogeneity of intracellular structure and junction proteins is present in different orders of the iris vasculature in addition to vascular diameter

  12. Thermoelectric efficiency of molecular junctions.

    PubMed

    Perroni, C A; Ninno, D; Cataudella, V

    2016-09-21

    Focus of the review is on experimental set-ups and theoretical proposals aimed to enhance thermoelectric performances of molecular junctions. In addition to charge conductance, the thermoelectric parameter commonly measured in these systems is the thermopower, which is typically rather low. We review recent experimental outcomes relative to several junction configurations used to optimize the thermopower. On the other hand, theoretical calculations provide estimations of all the thermoelectric parameters in the linear and non-linear regime, in particular of the thermoelectric figure of merit and efficiency, completing our knowledge of molecular thermoelectricity. For this reason, the review will mainly focus on theoretical studies analyzing the role of not only electronic, but also of the vibrational degrees of freedom. Theoretical results about thermoelectric phenomena in the coherent regime are reviewed focusing on interference effects which play a significant role in enhancing the figure of merit. Moreover, we review theoretical studies including the effects of molecular many-body interactions, such as electron-vibration couplings, which typically tend to reduce the efficiency. Since a fine tuning of many parameters and coupling strengths is required to optimize the thermoelectric conversion in molecular junctions, new theoretically proposed set-ups are discussed in the conclusions.

  13. Squeezed States in Josephson Junctions.

    NASA Astrophysics Data System (ADS)

    Hu, X.; Nori, F.

    1996-03-01

    We have studied quantum fluctuation properties of Josephson junctions in the limit of large Josephson coupling energy and small charging energy, when the eigenstates of the system can be treated as being nearly localized. We have considered(X. Hu and F. Nori, preprints.) a Josephson junction in a variety of situations, e.g., coupled to one or several of the following elements: a capacitor, an inductor (in a superconducting ring), and an applied current source. By solving an effective Shrödinger equation, we have obtained squeezed vacuum (coherent) states as the ground states of a ``free-oscillating'' (linearly-driven) Josephson junction, and calculated the uncertainties of its canonical momentum, charge, and coordinate, phase. We have also shown that the excited states of the various systems we consider are similar to the number states of a simple harmonic oscillator but with different fluctuation properties. Furthermore, we have obtained the time-evolution operators for these systems. These operators can make it easier to calculate the time-dependence of the expectation values and fluctuations of various quantities starting from an arbitrary initial state.

  14. Three-way RNA junctions with remote tertiary contacts: A recurrent and highly versatile fold

    PubMed Central

    de la Peña, Marcos; Dufour, David; Gallego, José

    2009-01-01

    Three-way junction RNAs adopt a recurrent Y shape when two of the helices form a coaxial stack and the third helix establishes one or more tertiary contacts several base pairs away from the junction. In this review, the structure, distribution, and functional relevance of these motifs are examined. Structurally, the folds exhibit conserved junction topologies, and the distal tertiary interactions play a crucial role in determining the final shape of the structures. The junctions and remote tertiary contacts behave as flexible hinge motifs that respond to changes in the other region, providing these folds with switching mechanisms that have been shown to be functionally useful in a variety of contexts. In addition, the juxtaposition of RNA domains at the junction and at the distal tertiary complexes enables the RNA helices to adopt unusual conformations that are frequently used by proteins, RNA molecules, and antibiotics as platforms for specific binding. As a consequence of these properties, Y-shaped junctions are widely distributed in all kingdoms of life, having been observed in small naked RNAs such as riboswitches and ribozymes or embedded in complex ribonucleoprotein systems like ribosomal RNAs, RNase P, or the signal recognition particle. In all cases, the folds were found to play an essential role for the functioning or assembly of the RNA or ribonucleoprotein systems that contain them. PMID:19741022

  15. Sensor assembly

    DOEpatents

    Bennett, Thomas E.; Nelson, Drew V.

    2004-04-13

    A ribbon-like sensor assembly is described wherein a length of an optical fiber embedded within a similar lengths of a prepreg tow. The fiber is ""sandwiched"" by two layers of the prepreg tow which are merged to form a single consolidated ribbon. The consolidated ribbon achieving a generally uniform distribution of composite filaments near the embedded fiber such that excess resin does not ""pool"" around the periphery of the embedded fiber.

  16. Dump assembly

    DOEpatents

    Goldmann, Louis H.

    1986-01-01

    A dump assembly having a fixed conduit and a rotatable conduit provided with overlapping plates, respectively, at their adjacent ends. The plates are formed with openings, respectively, normally offset from each other to block flow. The other end of the rotatable conduit is provided with means for securing the open end of a filled container thereto. Rotation of the rotatable conduit raises and inverts the container to empty the contents while concurrently aligning the conduit openings to permit flow of material therethrough.

  17. Communication via gap junctions underlies early functional and beneficial interactions between grafted neural stem cells and the host.

    PubMed

    Jäderstad, Johan; Jäderstad, Linda M; Li, Jianxue; Chintawar, Satyan; Salto, Carmen; Pandolfo, Massimo; Ourednik, Vaclav; Teng, Yang D; Sidman, Richard L; Arenas, Ernest; Snyder, Evan Y; Herlenius, Eric

    2010-03-16

    How grafted neural stem cells (NSCs) and their progeny integrate into recipient brain tissue and functionally interact with host cells is as yet unanswered. We report that, in organotypic slice cultures analyzed by ratiometric time-lapse calcium imaging, current-clamp recordings, and dye-coupling methods, an early and essential way in which grafted murine or human NSCs integrate functionally into host neural circuitry and affect host cells is via gap-junctional coupling, even before electrophysiologically mature neuronal differentiation. The gap junctions, which are established rapidly, permit exogenous NSCs to influence directly host network activity, including synchronized calcium transients with host cells in fluctuating networks. The exogenous NSCs also protect host neurons from death and reduce such signs of secondary injury as reactive astrogliosis. To determine whether gap junctions between NSCs and host cells may also mediate neuroprotection in vivo, we examined NSC transplantation in two murine models characterized by degeneration of the same cell type (Purkinje neurons) from different etiologies, namely, the nervous and SCA1 mutants. In both, gap junctions (containing connexin 43) formed between NSCs and host cells at risk, and were associated with rescue of neurons and behavior (when implantation was performed before overt neuron loss). Both in vitro and in vivo beneficial NSC effects were abrogated when gap junction formation or function was suppressed by pharmacologic and/or RNA-inhibition strategies, supporting the pivotal mediation by gap-junctional coupling of some modulatory, homeostatic, and protective actions on host systems as well as establishing a template for the subsequent development of electrochemical synaptic intercellular communication.

  18. Communication via gap junctions underlies early functional and beneficial interactions between grafted neural stem cells and the host

    PubMed Central

    Jäderstad, Johan; Jäderstad, Linda M.; Li, Jianxue; Chintawar, Satyan; Salto, Carmen; Pandolfo, Massimo; Ourednik, Vaclav; Teng, Yang D.; Sidman, Richard L.; Arenas, Ernest; Snyder, Evan Y.; Herlenius, Eric

    2010-01-01

    How grafted neural stem cells (NSCs) and their progeny integrate into recipient brain tissue and functionally interact with host cells is as yet unanswered. We report that, in organotypic slice cultures analyzed by ratiometric time-lapse calcium imaging, current-clamp recordings, and dye-coupling methods, an early and essential way in which grafted murine or human NSCs integrate functionally into host neural circuitry and affect host cells is via gap-junctional coupling, even before electrophysiologically mature neuronal differentiation. The gap junctions, which are established rapidly, permit exogenous NSCs to influence directly host network activity, including synchronized calcium transients with host cells in fluctuating networks. The exogenous NSCs also protect host neurons from death and reduce such signs of secondary injury as reactive astrogliosis. To determine whether gap junctions between NSCs and host cells may also mediate neuroprotection in vivo, we examined NSC transplantation in two murine models characterized by degeneration of the same cell type (Purkinje neurons) from different etiologies, namely, the nervous and SCA1 mutants. In both, gap junctions (containing connexin 43) formed between NSCs and host cells at risk, and were associated with rescue of neurons and behavior (when implantation was performed before overt neuron loss). Both in vitro and in vivo beneficial NSC effects were abrogated when gap junction formation or function was suppressed by pharmacologic and/or RNA-inhibition strategies, supporting the pivotal mediation by gap-junctional coupling of some modulatory, homeostatic, and protective actions on host systems as well as establishing a template for the subsequent development of electrochemical synaptic intercellular communication. PMID:20147621

  19. Acetaldehyde disrupts tight junctions in Caco-2 cell monolayers by a protein phosphatase 2A-dependent mechanism.

    PubMed

    Dunagan, Mitzi; Chaudhry, Kamaljit; Samak, Geetha; Rao, R K

    2012-12-15

    Acetaldehyde is accumulated at high concentrations in the colonic lumen following ethanol administration. Previous studies demonstrated that acetaldehyde disrupts intestinal epithelial tight junctions and increases paracellular permeability. In the present study, we investigated the role of PP2A in the acetaldehyde-induced disruption of intestinal epithelial tight junctions. Caco-2 cell monolayers were exposed to 200-600 μM acetaldehyde for varying times, and the epithelial barrier function was evaluated by measuring transepithelial electrical resistance and inulin permeability. Acetaldehyde treatment resulted in a time-dependent increase in inulin permeability and redistribution of occludin and ZO-1 from the intercellular junctions. Treatment of cells with fostriecin (a PP2A-selective inhibitor) or knockdown of PP2A by siRNA blocked acetaldehyde-induced increase in inulin permeability and redistribution of occludin and ZO-1. The effects of fostriecin and acetaldehyde were confirmed in mouse intestine ex vivo. Acetaldehyde-induced tight junction disruption and barrier dysfunction were also attenuated by a PP2A-specific inhibitory peptide, TPDYFL. Coimmunoprecipitation studies showed that acetaldehyde increased the interaction of PP2A with occludin and induced dephosphorylation of occludin on threonine residues. Fostriecin and TPDYFL significantly reduced acetaldehyde-induced threonine dephosphorylation of occludin. Acetaldehyde failed to change the level of the methylated form of PP2A-C subunit. However, genistein (a tyrosine kinase inhibitor) blocked acetaldehyde-induced association of PP2A with occludin and threonine dephosphorylation of occludin. These results demonstrate that acetaldehyde-induced disruption of tight junctions is mediated by PP2A translocation to tight junctions and dephosphorylation of occludin on threonine residues.

  20. Antofine-induced connexin43 gap junction disassembly in rat astrocytes involves protein kinase Cβ.

    PubMed

    Huang, Yu-Fang; Liao, Chih-Kai; Lin, Jau-Chen; Jow, Guey-Mei; Wang, Hwai-Shi; Wu, Jiahn-Chun

    2013-03-01

    Antofine, a phenanthroindolizidine alkaloid derived from Cryptocaryachinensis and Ficusseptica in the Asclepiadaceae milkweed family, is cytotoxic for various cancer cell lines. In this study, we demonstrated that treatment of rat primary astrocytes with antofine induced dose-dependent inhibition of gap junction intercellular communication (GJIC), as assessed by scrape-loading 6-carboxyfluorescein dye transfer. Levels of Cx43 protein were also decreased in a dose- and time-dependent manner following antofine treatment. Double-labeling immunofluorescence microscopy showed that antofine (10ng/ml) induced endocytosis of surface gap junctions into the cytoplasm, where Cx43 was co-localized with the early endosome marker EEA1. Inhibition of lysosomes or proteasomes by co-treatment with antofine and their respective specific inhibitors, NH4Cl or MG132, partially inhibited the antofine-induced decrease in Cx43 protein levels, but did not inhibit the antofine-induced inhibition of GJIC. After 30min of treatment, antofine induced a rapid increase in the intracellular Ca(2+) concentration and activation of protein kinase C (PKC)α/βII, which was maintained for at least 6h. Co-treatment of astrocytes with antofine and the intracellular Ca(2+) chelator BAPTA-AM prevented downregulation of Cx43 and inhibition of GJIC. Moreover, co-treatment with antofine and a specific PKCβ inhibitor prevented endocytosis of gap junctions, downregulation of Cx43, and inhibition of GJIC. Taken together, these findings indicate that antofine induces Cx43 gap junction disassembly by the PKCβ signaling pathway. Inhibition of GJIC by antofine may undermine the neuroprotective effect of astrocytes in CNS.

  1. Ezrin: a regulator of actin microfilaments in cell junctions of the rat testis

    PubMed Central

    Gungor-Ordueri, N Ece; Celik-Ozenci, Ciler; Cheng, C Yan

    2015-01-01

    Ezrin, radixin, moesin and merlin (ERM) proteins are highly homologous actin-binding proteins that share extensive sequence similarity with each other. These proteins tether integral membrane proteins and their cytoplasmic peripheral proteins (e.g., adaptors, nonreceptor protein kinases and phosphatases) to the microfilaments of actin-based cytoskeleton. Thus, these proteins are crucial to confer integrity of the apical membrane domain and its associated junctional complex, namely the tight junction and the adherens junction. Since ectoplasmic specialization (ES) is an F-actin-rich testis-specific anchoring junction-a highly dynamic ultrastructure in the seminiferous epithelium due to continuous transport of germ cells, in particular spermatids, across the epithelium during the epithelial cycle-it is conceivable that ERM proteins are playing an active role in these events. Although these proteins were first reported almost 25 years and have since been extensively studied in multiple epithelia/endothelia, few reports are found in the literature to examine their role in the actin filament bundles at the ES. Studies have shown that ezrin is also a constituent protein of the actin-based tunneling nanotubes (TNT) also known as intercellular bridges, which are transient cytoplasmic tubular ultrastructures that transport signals, molecules and even organelles between adjacent and distant cells in an epithelium to coordinate cell events that occur across an epithelium. Herein, we critically evaluate recent data on ERM in light of recent findings in the field in particular ezrin regarding its role in actin dynamics at the ES in the testis, illustrating additional studies are warranted to examine its physiological significance in spermatogenesis. PMID:25652626

  2. SLO BK Potassium Channels Couple Gap Junctions to Inhibition of Calcium Signaling in Olfactory Neuron Diversification

    PubMed Central

    Schumacher, Jennifer A.; Wang, Xiaohong; Merrill, Sean A.; Millington, Grethel; Bayne, Brittany; Jorgensen, Erik M.; Chuang, Chiou-Fen

    2016-01-01

    The C. elegans AWC olfactory neuron pair communicates to specify asymmetric subtypes AWCOFF and AWCON in a stochastic manner. Intercellular communication between AWC and other neurons in a transient NSY-5 gap junction network antagonizes voltage-activated calcium channels, UNC-2 (CaV2) and EGL-19 (CaV1), in the AWCON cell, but how calcium signaling is downregulated by NSY-5 is only partly understood. Here, we show that voltage- and calcium-activated SLO BK potassium channels mediate gap junction signaling to inhibit calcium pathways for asymmetric AWC differentiation. Activation of vertebrate SLO-1 channels causes transient membrane hyperpolarization, which makes it an important negative feedback system for calcium entry through voltage-activated calcium channels. Consistent with the physiological roles of SLO-1, our genetic results suggest that slo-1 BK channels act downstream of NSY-5 gap junctions to inhibit calcium channel-mediated signaling in the specification of AWCON. We also show for the first time that slo-2 BK channels are important for AWC asymmetry and act redundantly with slo-1 to inhibit calcium signaling. In addition, nsy-5-dependent asymmetric expression of slo-1 and slo-2 in the AWCON neuron is necessary and sufficient for AWC asymmetry. SLO-1 and SLO-2 localize close to UNC-2 and EGL-19 in AWC, suggesting a role of possible functional coupling between SLO BK channels and voltage-activated calcium channels in AWC asymmetry. Furthermore, slo-1 and slo-2 regulate the localization of synaptic markers, UNC-2 and RAB-3, in AWC neurons to control AWC asymmetry. We also identify the requirement of bkip-1, which encodes a previously identified auxiliary subunit of SLO-1, for slo-1 and slo-2 function in AWC asymmetry. Together, these results provide an unprecedented molecular link between gap junctions and calcium pathways for terminal differentiation of olfactory neurons. PMID:26771544

  3. Installation and Assembly, Electrical Ground Support Equipment (GSE), Specification for

    NASA Technical Reports Server (NTRS)

    Denson, Erik C.

    2014-01-01

    This specification covers the general workmanship requirements and procedures for the complete installation and assembly of electrical ground support equipment (EGSE) such as terminal distributors, junction boxes, conduit and fittings, cable trays and accessories, interconnecting cables (including routing requirements), motor-control equipment, and necessary hardware as specified by the applicable contract and drawings.

  4. Ozone concentration in leaf intercellular air spaces is close to zero

    SciTech Connect

    Laisk, A.; Moldau, H. ); Kull, O. )

    1989-07-01

    Transpiration and ozone uptake rates were measured simultaneously in sunflower leaves at different stomatal openings and various ozone concentrations. Ozone uptake rates were proportional to the ozone concentration up to 1500 nanoliters per liter. The leaf gas phase diffusion resistance (stomatal plus boundary layer) to water vapor was calculated and converted to the resistance to ozone multiplying it by the theoretical ratio of diffusion coefficients for water vapor and ozone in air (1.67). The ozone concentration in intercellular air spaces calculated from the ozone uptake rate and diffusion resistance to ozone scattered around zero. The ozone concentration in intercellular air spaces was measured directly bu supplying ozone to the leaf from one side and measuring the equilibrium concentration above the other side, and it was found to be zero. The total leaf resistance to ozone was proportional to the gas phase resistance to water vapor with a coefficient of 1.68. It is concluded that ozone enters the leaf by diffusion through the stomata, and is rapidly decomposed in cell walls and plasmalemma.

  5. Novel microscopy-based screening method reveals regulators of contact-dependent intercellular transfer

    PubMed Central

    Michael Frei, Dominik; Hodneland, Erlend; Rios-Mondragon, Ivan; Burtey, Anne; Neumann, Beate; Bulkescher, Jutta; Schölermann, Julia; Pepperkok, Rainer; Gerdes, Hans-Hermann; Kögel, Tanja

    2015-01-01

    Contact-dependent intercellular transfer (codeIT) of cellular constituents can have functional consequences for recipient cells, such as enhanced survival and drug resistance. Pathogenic viruses, prions and bacteria can also utilize this mechanism to spread to adjacent cells and potentially evade immune detection. However, little is known about the molecular mechanism underlying this intercellular transfer process. Here, we present a novel microscopy-based screening method to identify regulators and cargo of codeIT. Single donor cells, carrying fluorescently labelled endocytic organelles or proteins, are co-cultured with excess acceptor cells. CodeIT is quantified by confocal microscopy and image analysis in 3D, preserving spatial information. An siRNA-based screening using this method revealed the involvement of several myosins and small GTPases as codeIT regulators. Our data indicates that cellular protrusions and tubular recycling endosomes are important for codeIT. We automated image acquisition and analysis to facilitate large-scale chemical and genetic screening efforts to identify key regulators of codeIT. PMID:26271723

  6. Dilated intercellular spaces and chronic cough as an extra-oesophageal manifestation of gastrooesophageal reflux disease.

    PubMed

    Orlando, Roy C

    2011-06-01

    Chronic cough is one of the extra-oesophageal manifestations of gastrooesophageal reflux disease (GORD). It is presumed to occur either directly by microaspiration of acidic gastric contents into the airway or indirectly by a reflex triggered by contact of acidic refluxates with the oesophageal epithelium in GORD. How contact of the oesophageal epithelium with acidic refluxates promotes sensitization for chronic cough is unknown, but like heartburn, which is a necessary accompaniment, it requires acid activation of nociceptors within the oesophageal mucosa. Dilated intercellular spaces within the oesophageal epithelium, a reflection of an increase in paracellular permeability, is a histopathologic feature of both erosive and non-erosive forms of GORD. Since it correlates with the symptom of heartburn, it is hypothesized herein that the increase in paracellular permeability to acid reflected by dilated intercellular spaces in oesophageal epithelium also serves as mediator of the signals that produce the reflex-induced sensitization for cough--a sensitization that can occur centrally within the medullary Nucleus Tractus Solitarius or peripherally within the tracheobronchial tree.

  7. Requirements for the Ca2+-independent component in the initial intercellular adhesion of C2 myoblasts

    PubMed Central

    1988-01-01

    Using a sensitive and quantitative adhesion assay, we have studied the initial stages of the intercellular adhesion of the C2 mouse myoblast line. After dissociation in low levels of trypsin in EDTA, C2 cells can rapidly reaggregate by Ca2+-independent mechanisms to form large multicellular aggregates. If cells are allowed to recover from dissociation by incubation in defined media, this adhesive system is augmented by a Ca2+-dependent mechanism with maximum recovery seen after 4 h incubation. The Ca2+-independent adhesion system is inhibited by preincubation of cell monolayers with cycloheximide before dissociation. Aggregation is also reduced after exposure to monensin, implicating a role for surface-translocated glycoproteins in this mechanism of adhesion. In coaggregation experiments using C2 myoblasts and 3T3 fibroblasts in which the Ca2+-dependent adhesion system was inactivated, no adhesive specificity between the two cell types was seen. Although synthetic peptides containing the RGD sequence are known to inhibit cell-substratum adhesion in various cell types, incubation of C2 myoblasts with the integrin-binding tetrapeptide, RGDS, greatly stimulated the Ca2+-independent aggregation of these cells while control analogs had no effect. These results show that a Ca2+- independent mechanism alone is sufficient to allow for the rapid formation of multicellular aggregates in a mouse myoblast line, and that many of the requirements and perturbants of the Ca2+-independent system of intercellular myoblast adhesion are similar to those of the Ca2+-dependent adhesion mechanisms. PMID:3198689

  8. Evolutionary and molecular analysis of Dof transcription factors identified a conserved motif for intercellular protein trafficking.

    PubMed

    Chen, Huan; Ahmad, Munawar; Rim, Yeonggil; Lucas, William J; Kim, Jae-Yean

    2013-06-01

    · Cell-to-cell trafficking of transcription factors (TFs) has been shown to play an important role in the regulation of plant developmental events, but the evolutionary relationship between cell-autonomous and noncell-autonomous (NCA) TFs remains elusive. · AtDof4.1, named INTERCELLULAR TRAFFICKING DOF 1 (ITD1), was chosen as a representative NCA member to explore this evolutionary relationship. Using domain structure-function analyses and swapping studies, we examined the cell-to-cell trafficking of plant-specific Dof TF family members across Arabidopsis and other species. · We identified a conserved intercellular trafficking motif (ITM) that is necessary and sufficient for selective cell-to-cell trafficking and can impart gain-of-function cell-to-cell movement capacity to an otherwise cell-autonomous TF. The functionality of related motifs from Dof members across the plant kingdom extended, surprisingly, to a unicellular alga that lacked plasmodesmata. By contrast, the algal homeodomain related to the NCA KNOX homeodomain was either inefficient or unable to impart such cell-to-cell movement function. · The Dof ITM appears to predate the evolution of selective plasmodesmal trafficking in the plant kingdom, which may well have acted as a molecular template for the evolution of Dof proteins as NCA TFs. However, the ability to efficiently traffic for KNOX homeodomain (HD) proteins may have been acquired during the evolution of early nonvascular plants.

  9. Electrotonic potentials in Aloe vera L.: Effects of intercellular and external electrodes arrangement.

    PubMed

    Volkov, Alexander G; Nyasani, Eunice K; Tuckett, Clayton; Scott, Jessenia M; Jackson, Mariah M Z; Greeman, Esther A; Greenidge, Ariane S; Cohen, Devin O; Volkova, Maia I; Shtessel, Yuri B

    2017-02-01

    Electrostimulation of plants can induce plant movements, activation of ion channels, ion transport, gene expression, enzymatic systems activation, electrical signaling, plant-cell damage, enhanced wound healing, and influence plant growth. Here we found that electrical networks in plant tissues have electrical differentiators. The amplitude of electrical responses decreases along a leaf and increases by decreasing the distance between polarizing Pt-electrodes. Intercellular Ag/AgCl electrodes inserted in a leaf and extracellular Ag/AgCl electrodes attached to the leaf surface were used to detect the electrotonic potential propagation along a leaf of Aloe vera. There is a difference in duration and amplitude of electrical potentials measured by electrodes inserted in a leaf and those attached to a leaf's surface. If the external reference electrode is located in the soil near the root, it changes the amplitude and duration of electrotonic potentials due to existence of additional resistance, capacitance, ion channels and ion pumps in the root. The information gained from this study can be used to elucidate extracellular and intercellular communication in the form of electrical signals within plants.

  10. Ozone Concentration in Leaf Intercellular Air Spaces Is Close to Zero 1

    PubMed Central

    Laisk, Agu; Kull, Olevi; Moldau, Heino

    1989-01-01

    Transpiration and ozone uptake rates were measured simultaneously in sunflower leaves at different stomatal openings and various ozone concentrations. Ozone uptake rates were proportional to the ozone concentration up to 1500 nanoliters per liter. The leaf gas phase diffusion resistance (stomatal plus boundary layer) to water vapor was calculated and converted to the resistance to ozone multiplying it by the theoretical ratio of diffusion coefficients for water vapor and ozone in air (1.67). The ozone concentration in intercellular air spaces calculated from the ozone uptake rate and diffusion resistance to ozone scattered around zero. The ozone concentration in intercellular air spaces was measured directly by supplying ozone to the leaf from one side and measuring the equilibrium concentration above the other side, and it was found to be zero. The total leaf resistance to ozone was proportional to the gas phase resistance to water vapor with a coefficient of 1.68. It is concluded that ozone enters the leaf by diffusion through the stomata, and is rapidly decomposed in cell walls and plasmalemma. PMID:16666867

  11. Rab8a/Rab11a regulate intercellular communications between neural cells via tunneling nanotubes

    PubMed Central

    Zhu, Hui; Xue, Chengbin; Xu, Xi; Guo, Yibing; Li, Xiaohong; Lu, Jingjing; Ju, Shaoqing; Wang, Yongjun; Cao, Zheng; Gu, Xiaosong

    2016-01-01

    Tunneling nanotubes (TNTs) are F-actin-based membrane tubes, and can form between cultured cells and within vital tissues. TNTs mediate intercellular communications that range from electrical signaling to the transfer of organelles. Following peripheral nerve injury, the orchestrated intercellular communications among neural and non-neural cells are required for effective nerve regeneration. It remains unknown whether TNTs exist between neural cells in the peripheral nerve system and how TNTs affect neural regeneration. To address these interesting questions, we investigated the transfer of neurotropic factors, membrane protein, cytoplasmic protein, mitochondria and RNA in functional TNTs formed between cultured Schwann cells (SCs). TNT-like structures were increased not only in cultured SCs after exposure to serum depletion but also in longitudinal sections of proximal sciatic nerve stump harvested after rat peripheral nerve transection. Meanwhile, downregulation of Rab8a or Rab11a in cultured SCs inhibited the formation of functional TNTs and vesicle transfer and led to decrease in cell migration, increase in SCs apoptosis. Likewise, knockdown of Rab8a or Rab11a in primary SCs also suppressed axonal outgrowth from co-cultured dorsal root ganglion (DRG) neurons. Overall, our results suggested that the gene of Rab8a or Rab11a might be involved in the formation of TNTs structures in the peripheral nerve system, while TNTs structures were likely to affect peripheral nerve regeneration through the regulation of neural cell communications. PMID:28005071

  12. Exploring the role of lipids in intercellular conduits: breakthroughs in the pipeline.

    PubMed

    Delage, Elise; Zurzolo, Chiara

    2013-12-10

    It has been known for more than a century that most of the plant cells are connected to their neighbors through membranous pores perforating the cell wall, namely plasmodesmata (PDs). The recent discovery of tunneling nanotubes (TNTs), thin membrane bridges established between distant mammalian cells, suggests that intercellular communication mediated through cytoplasmic continuity could be a conserved feature of eukaryotic organisms. Although TNTs differ from PDs in their formation and architecture, both are characterized by a continuity of the plasma membrane between two cells, delimiting a nanotubular channel supported by actin-based cytoskeleton. Due to this unusual membrane organization, lipids are likely to play critical roles in the formation and stability of intercellular conduits like TNTs and PDs, but also in regulating the transfer through these structures. While it is crucial for a better understanding of those fascinating communication highways, the study of TNT lipid composition and dynamics turned out to be extremely challenging. The present review aims to give an overview of the recent findings in this context. We will also discuss some of the promising imaging approaches, which might be the key for future breakthroughs in the field and could also benefit the research on PDs.

  13. Microvesicles as mediators of intercellular communication in cancer--the emerging science of cell